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NF-??B Controls Cell Growth and Differentiation through Transcriptional Regulation of Cyclin D1
Abstract
Accumulating evidence implicates the transcription factor NF-κB as a positive mediator of cell growth, but the molecular mechanism(s) involved in this process remains largely unknown. Here we use both a skeletal muscle differentiation model and normal diploid fibroblasts to gain insight into how NF-κB regulates cell growth and differentiation. Results obtained with the C2C12 myoblast cell line demonstrate that NF-κB functions as an inhibitor of myogenic differentiation. Myoblasts generated to lack NF-κB activity displayed defects in cellular proliferation and cell cycle exit upon differentiation. An analysis of cell cycle markers revealed that NF-κB activates cyclin D1 expression, and the results showed that this regulatory pathway is one mechanism by which NF-κB inhibits myogenesis. NF-κB regulation of cyclin D1 occurs at the transcriptional level and is mediated by direct binding of NF-κB to multiple sites in the cyclin D1 promoter. Using diploid fibroblasts, we demonstrate that NF-κB is required to induce cyclin D1 expression and pRb hyperphosphorylation and promote G
1
-to-S progression. Consistent with results obtained with the C2C12 differentiation model, we show that NF-κB also promotes cell growth in embryonic fibroblasts, correlating with its regulation of cyclin D1. These data therefore identify cyclin D1 as an important transcriptional target of NF-κB and reveal a mechanism to explain how NF-κB is involved in the early phases of the cell cycle to regulate cell growth and differentiation.
... NF-κB is an inducible nuclear transcription factor involved in immune responses, cell proliferation, and apoptosis. NF-κB is strongly activated in response to TNFα and contributes to cell survival and proliferation [13,23,24,43,44]. NF-κB controls the expression of several genes encoding anti-apoptotic proteins, such as c-FLIP, Bcl-2, Bcl-x L , and cIAP2 [45], as well as proteins involved in proliferation, such as cyclin D1 [43,44]. ...
... NF-κB is strongly activated in response to TNFα and contributes to cell survival and proliferation [13,23,24,43,44]. NF-κB controls the expression of several genes encoding anti-apoptotic proteins, such as c-FLIP, Bcl-2, Bcl-x L , and cIAP2 [45], as well as proteins involved in proliferation, such as cyclin D1 [43,44]. Disruption of NF-κB activity either by genetic manipulations or by chemical inhibition of IKK kinase, which activates this transcription factor, renders the cells highly susceptible to TNFα-induced cell death [22][23][24]. ...
... The lower NF-κB nuclear localisation in cells treated with a combination of TNFα and ML-60218 may also partially explain the phenotypes observed in our current work. Firstly, NF-κB is known to positively regulate cyclin D1 through direct binding within the CCND1 gene promoter [43,44]. Thus, NF-κB inactivation may prevent cyclin D1 upregulation and cell cycle acceleration, a phenomenon observed by us and previously by others [43,44]. ...
Simple Summary
Tumour necrosis factor alpha (TNFα) is a cytokine that plays an important role in apoptosis, cell survival, as well as in inflammation and immunity. Although named for its antitumor properties, TNFα has tumour-promoting properties. TNFα is often present in large quantities in tumours, and cancer cells frequently acquire resistance to this cytokine. Identifying the means to sensitise the cancer cells to TNFα would have therapeutical benefits. We, therefore, sought to determine whether inhibition of RNA polymerase III (Pol III), which synthesises several essential components of the protein biosynthetic machinery, would affect the response of cancer cells to TNFα. Here we show that Pol III inhibition augments the cytotoxic and cytostatic effects of TNFα. Our data suggest that targeting Pol III may be a potential therapeutic intervention to treat colorectal cancer.
Abstract
Tumour necrosis factor alpha (TNFα) is a multifunctional cytokine that plays a pivotal role in apoptosis, cell survival, as well as in inflammation and immunity. Although named for its antitumor properties, TNFα also has tumour-promoting properties. TNFα is often present in large quantities in tumours, and cancer cells frequently acquire resistance to this cytokine. Consequently, TNFα may increase the proliferation and metastatic potential of cancer cells. Furthermore, the TNFα-driven increase in metastasis is a result of the ability of this cytokine to induce the epithelial-to-mesenchymal transition (EMT). Overcoming the resistance of cancer cells to TNFα may have a potential therapeutic benefit. NF-κB is a crucial transcription factor mediating inflammatory signals and has a wide-ranging role in tumour progression. NF-κB is strongly activated in response to TNFα and contributes to cell survival and proliferation. The pro-inflammatory and pro-survival function of NF-κB can be disrupted by blocking macromolecule synthesis (transcription, translation). Consistently, inhibition of transcription or translation strongly sensitises cells to TNFα-induced cell death. RNA polymerase III (Pol III) synthesises several essential components of the protein biosynthetic machinery, such as tRNA, 5S rRNA, and 7SL RNA. No studies, however, directly explored the possibility that specific inhibition of Pol III activity sensitises cancer cells to TNFα. Here we show that in colorectal cancer cells, Pol III inhibition augments the cytotoxic and cytostatic effects of TNFα. Pol III inhibition enhances TNFα-induced apoptosis and also blocks TNFα-induced EMT. Concomitantly, we observe alterations in the levels of proteins related to proliferation, migration, and EMT. Finally, our data show that Pol III inhibition is associated with lower NF-κB activation upon TNFα treatment, thus potentially suggesting the mechanism of Pol III inhibition-driven sensitisation of cancer cells to this cytokine.
... Whereas the increase in pNFκBp65_Ser536 was higher in the Comb-NMES (+147%) vs. (+88%) in the control group. Studies have shown that the p65 subunit is associated with the promotion of myogenesis, indicating a positive regulatory role in skeletal muscle differentiation [44][45][46]. Additionally, the p65 subunit has been identified as containing relatively abundant DNA binding activities in skeletal muscles, suggesting its involvement in the formation and/or function of skeletal muscles [44]. ...
... Studies have shown that the p65 subunit is associated with the promotion of myogenesis, indicating a positive regulatory role in skeletal muscle differentiation [44][45][46]. Additionally, the p65 subunit has been identified as containing relatively abundant DNA binding activities in skeletal muscles, suggesting its involvement in the formation and/or function of skeletal muscles [44]. The reduction in TLR4 could indicate an adaptation of the muscle cells to the Comb-NMES intervention, resulting in the resolution of the initial inflammatory response. ...
In individuals with spinal cord injury (SCI), rapid skeletal muscle atrophy and metabolic dysfunction pose profound rehabilitation challenges, often resulting in substantial loss of muscle mass and function. This study evaluates the effect of combined neuromuscular electrical stimulation (Comb-NMES) on skeletal muscle cross-sectional area (CSA) and inflammatory signaling within the acute phase of SCI. We applied a novel Comb-NMES regimen, integrating both high-frequency resistance and low-frequency aerobic protocols on the vastus lateralis muscle, to participants early post-SCI. Muscle biopsies were analyzed for CSA and inflammatory markers pre-and post-intervention. The results revealed a preservation of muscle CSA in the Comb-NMES group compared to a control group receiving transcutaneous electrical nerve stimulation and passive movements. Inflammatory signaling proteins such as TLR4 and Atrogin-1 were downregulated, whereas markers associated with muscle repair and growth were modulated beneficially in the Comb-NMES group. The study's findings suggest that early application of Comb-NMES post-SCI can attenuate inflammatory pathways linked to muscle atrophy and potentially promote muscle repair, advocating for its inclusion in early rehabilitation efforts. However, the small sample size and variability in injury characteristics emphasize the need for further research to corroborate these results across a more diverse and extensive SCI population.
... Neurotrophin signaling mediated through p75NTR has been shown to activate the nuclear factor-kB (NF-kB) and Jun kinase signaling pathways (including JNK pathway) ( 57 ,58 ). While the NF-kB and JNK pathways play an important role in neural development in many aspects, it is also known that they regulate the expression and activation of cyclins, including Cyclin G1 (CCNG1) (59)(60)(61). More specifically, the NF-kB signaling pathway was shown to regulate Cyclin D1 (CCND1) expression, whereas the JNK signaling pathway was shown to regulate the expression of both CCND1 and CCNG1 (59)(60)(61). ...
... While the NF-kB and JNK pathways play an important role in neural development in many aspects, it is also known that they regulate the expression and activation of cyclins, including Cyclin G1 (CCNG1) (59)(60)(61). More specifically, the NF-kB signaling pathway was shown to regulate Cyclin D1 (CCND1) expression, whereas the JNK signaling pathway was shown to regulate the expression of both CCND1 and CCNG1 (59)(60)(61). CCNG1 is a non-canonical cyclin that was initially discovered as a novel member of the cyclin family ( 62 ). Although the exact mechanism is not yet clear, CCNG1 is known to play a crucial role in cell proliferation and cell growth (63)(64)(65). ...
The versatility of cellular response arises from the communication, or crosstalk, of signaling pathways in a complex network of signaling and transcriptional regulatory interactions. Understanding the various mechanisms underlying crosstalk on a global scale requires untargeted computational approaches. We present a network-based statistical approach, MuXTalk, that uses high-dimensional edges called multilinks to model the unique ways in which signaling and regulatory interactions can interface. We demonstrate that the signaling-regulatory interface is located primarily in the intermediary region between signaling pathways where crosstalk occurs, and that multilinks can differentiate between distinct signaling-transcriptional mechanisms. Using statistically over-represented multilinks as proxies of crosstalk, we infer crosstalk among 60 signaling pathways, expanding currently available crosstalk databases by more than five-fold. MuXTalk surpasses existing methods in terms of model performance metrics, identifies additions to manual curation efforts, and pinpoints potential mediators of crosstalk. Moreover, it accommodates the inherent context-dependence of crosstalk, allowing future applications to cell type- and disease-specific crosstalk.
... Additionally, NF-κB maintains myoblasts in a proliferative state by binding to the cyclin D1 promoter and directing its transcription, thereby stimulating the progression of the cells into the S phase. Guttridge et al. demonstrated that in myogenesis, this process is regulated via the classical p50/p65 heterodimer [25]. It was observed that NF-κB inhibits myogenesis via a different pathway by inducing the transcription factor YY-1, a component of the Polycomb repressive complex [26]. ...
... Currently, the chronic activity of NF-κB is known as a hallmark of this disease, showing its detrimental influence on skeletal muscle [37]. Moreover, it has been observed that the activation of NF-κB through the conventional pathway enhances myoblast proliferation (through activation of cyclin D1) and inhibits muscle regeneration (by interrupting MyoD expression) [25]. It has been discovered that the conventional NF-κB dimer composed of p50/p65 is triggered in mdx mouse (model of DMD) muscles, with p65 serving as the transcriptionally active subunit responsible for muscle pathology [38]. ...
The NF-κB-signaling pathway plays a crucial role in cancer progression, including muscle-derived cancers such as rhabdomyosarcoma or sarcoma. Several natural compounds have been studied for their ability to alter NF-κB signaling in these types of cancers. This review paper summarizes the current knowledge on the effects of natural compounds, including curcumin, resveratrol, quercetin, epigallocatechin-3-gallate, and berberine, on NF-κB signaling in muscle-derived cancers. These compounds have been shown to inhibit NF-κB signaling in rhabdomyosarcoma cells through various mechanisms, such as inhibiting the activation of the IKK complex and the NF-κB transcription factor. These findings suggest that natural compounds could be potential therapeutic agents for muscle-derived cancers. However, further research is needed to fully understand their mechanisms of action and potential clinical applications.
... NF-κB, it is plausible that this glucocorticoid-mediated effect occurs via PARP1. Furthermore, NF-κB has also been implicated in differentiation programs of other tissues, including myogenesis [38,39,51]. In this regard, PARP1 and its PARylating activity exert influence over osteoclast differentiation and bone remodeling via NF-κB dependent transcription of IL-1β [39], and glucocorticoidinduced osteoporosis is a consequence of the inhibition of IL-1 production [52]. ...
... In this regard, PARP1 and its PARylating activity exert influence over osteoclast differentiation and bone remodeling via NF-κB dependent transcription of IL-1β [39], and glucocorticoidinduced osteoporosis is a consequence of the inhibition of IL-1 production [52]. Furthermore, as with PARP1, the suppression of NF-κB is fundamental for driving myogenesis [38,51], while dexamethasone treatment at the myoblast stage enhances myogenesis [20][21][22]. Concurrently, we demonstrate that application of dexamethasone during differentiation induction of myoblasts reduces PAR levels (Fig. 2D) indicating glucocorticoids have impacts on PARP1-mediated PARylating activity. ...
The ADP-ribosyltransferase, PARP1 enzymatically generates and applies the post-translational modification, ADP-Ribose (ADPR). PARP1 roles in genome maintenance are well described, but recent work highlights roles in many fundamental processes including cellular identity and energy homeostasis. Herein, we show in both mouse and human skeletal muscle cells that PARP1-mediated PARylation is a regulator of the myogenic program and the muscle transcriptional response to steroid hormones. Chemical PARP1 modulation impacts the expression of major myocellular proteins, including troponins, key in dictating muscle contractile force. Whilst PARP1 in absence of DNA damage is often assumed to be basally inactive, we show PARylation to be acutely sensitive to extracellular glucose concentrations and the steroid hormone class, glucocorticoids which exert considerable authority over muscle tissue mass. Specifically, we find during myogenesis, a transient and significant rise in PAR. This early-stage differentiation event, if blocked with PARP1 inhibition, reduced the abundance of important muscle proteins in the fully differentiated myotubes. This suggests that PAR targets during early-stage differentiation are central to the proper development of the muscle contractile unit. We also show that reduced PARP1 in myoblasts impacts a variety of metabolic pathways in line with the recorded actions of glucocorticoids. Currently, as both regulators of myogenesis and muscle mass loss, glucocorticoids represent a clinical conundrum. Our work goes on to identify that PARP1 influences transcriptional activation by glucocorticoids of a subset of genes critical to human skeletal muscle pathology. These genes may therefore signify a regulatory battery of targets through which selective glucocorticoid modulation could be achieved. Collectively, our data provide clear links between PARP1-mediated PARylation and skeletal muscle homeostatic mechanisms crucial to tissue mass maintenance and endocrine response.
... NF-kB is an important player in host response against infections since it induces the expression of inflammatory cytokines, chemokines, and their receptors. Beyond their role in immunity, all of these factors contribute to keeping the level of NF-kB activation high, and persistent inducing and amplifying side effects of NF-kB such as cyclin D1 and c-MYC transcription [25][26][27], which in turn, play important roles in cancer progression. Finally, NF-kB induces the production of host-deaminase as the APOBEC proteins and activation-induced cytidine deaminase (AID) enzymes, two families of proteins mainly involved in host-defense against viral infections that, however, can also mutate host DNA and contribute to cancer development [28]. ...
... As above reported, NF-kB plays an important role in HR-HPV induced cancer [24], since this pathway also leads to cyclin D1 and c-MYC transcription [25][26][27], which in turn also plays an important role in cancer progression. ...
Vaginal dysbiosis is characterized by a decrease in the relative abundance of Lactobacillus species in favor of other species. This condition facilitates infections by sexually transmitted pathogens including high risk (HR)-human papilloma viruses (HPVs) involved in the development of cervical cancer. Some vaginal dysbiosis bacteria contribute to the neoplastic progression by inducing chronic inflammation and directly activating molecular pathways involved in carcinogenesis. In this study, SiHa cells, an HPV-16-transformed epithelial cell line, were exposed to different representative vaginal microbial communities. The expression of the HPV oncogenes E6 and E7 and the production of relative oncoproteins was evaluated. The results showed that Lactobacillus crispatus and Lactobacillus gasseri modulated the basal expression of the E6 and E7 genes of SiHa cells and the production of the E6 and E7 oncoproteins. Vaginal dysbiosis bacteria had contrasting effects on E6/E7 gene expression and protein production. The expression of the E6 and E7 genes and the production of the relative oncoproteins was increased by strains of Gardnerella vaginalis and, to a lesser extent, by Megasphaera micronuciformis. In contrast, Prevotella bivia decreased the expression of oncogenes and the production of the E7 protein. A decreased amount of p53 and pRb was found in the cultures of SiHa cells with M. micronuciformis, and accordingly, in the same cultures, a higher percentage of cells progressed to the S-phase of the cell cycle compared to the untreated or Lactobacillus-stimulated cultures. These data confirm that L. crispatus represents the most protective component of the vaginal microbiota against neoplastic progression of HR-HPV infected cells, while M. micronuciformis and, to a lesser extent, G. vaginalis may directly interfere in the oncogenic process, inducing or maintaining the production of viral oncoproteins.
... Inflammation is associated with 343 activation of the NF-kappa B pathway (73). In general, inflammation and the NF-kappa B pathway can 344 cause tumor suppression and destruction of altered cells (74) and on the other hand, help the development 345 of cancer (73,74,(74)(75)(76)(77)(78)(79). NF-kappaB also regulates 500 genes involved in inflammation, proliferation, cell 346 transformation, angiogenesis and metastasis (80,81) and NF-kappaB activation and abnormal expression of 347 its subunits have been observed in breast cancer, which contributes to cancer progression and 348 development (82)(83)(84)(85)(86)(87)(88)(89). ...
Breast cancer (BC) is one of most important mortality factors among women therefore to find important genes in BC can help early diagnosis, treatment or prevention. TNFSF or tumor necrosis factor Superfamily have an important role in various cancers. In BC, some of studies have found dual roles for these genes. In this research, we conducted a comprehensive and detailed bioinformatics study on this family. UALCAN, TNMplot, UCSC Xena, GEPIA, The Human Protein Atlas, Kaplan–Meier plotter, bc-GenExMiner, cBioPortal, STRING, GeneMANIA, Enrichr, TIMER and shinyDepMap were used for analysis. We found that these genes play their role through the immune system and the high expression of eight FASLG, LTB, TNF, TNFSF8, TNFSF10, TNFSF11, TNFSF12, TNFSF13 genes were positively associated with OS and RFS. Overall, our data showed that these genes can be considered as prognostic biomarkers. Further, our results suggest that this family has anti-tumor activity.
... Cytoplasmic expression of these subunits appeared unchanged suggesting that inhibiting O-GlcNAcylation impairs their ability to translocate to the nucleus. Furthermore, OSMI-1 treatment also inhibited the expression of NF-kB target genes c-Myc, Cyclin D1, and Cyclin E1 (58)(59)(60) showing that inhibiting O-GlcNAcylation negatively regulates NF-kB transcriptional activity ( Figure 7B). Since the regulation of c-Myc, Cyclin D1, and Cyclin E1 largely account for the role of NF-kB in promoting cell growth (61), we studied the effect of O-GlcNAcylation inhibition on cell cycle progression in OCI-AML3 cells. ...
Introduction
Acute myeloid leukemia (AML) is the most common acute leukemia in adults with an overall poor prognosis and high relapse rate. Multiple factors including genetic abnormalities, differentiation defects and altered cellular metabolism contribute to AML development and progression. Though the roles of oxidative phosphorylation and glycolysis are defined in AML, the role of the hexosamine biosynthetic pathway (HBP), which regulates the O-GlcNAcylation of cytoplasmic and nuclear proteins, remains poorly defined.
Methods
We studied the expression of the key enzymes involved in the HBP in AML blasts and stem cells by RNA sequencing at the single-cell and bulk level. We performed flow cytometry to study OGT protein expression and global O-GlcNAcylation. We studied the functional effects of inhibiting O-GlcNAcylation on transcriptional activation in AML cells by Western blotting and real time PCR and on cell cycle by flow cytometry.
Results
We found higher expression levels of the key enzymes in the HBP in AML as compared to healthy donors in whole blood. We observed elevated O-GlcNAc Transferase (OGT) and O-GlcNAcase (OGA) expression in AML stem and bulk cells as compared to normal hematopoietic stem and progenitor cells (HSPCs). We also found that both AML bulk cells and stem cells show significantly enhanced OGT protein expression and global O-GlcNAcylation as compared to normal HSPCs, validating our in silico findings. Gene set analysis showed substantial enrichment of the NF-κB pathway in AML cells expressing high OGT levels. Inhibition of O-GlcNAcylation decreased NF-κB nuclear translocation and the expression of selected NF-κB-dependent genes controlling cell cycle. It also blocked cell cycle progression suggesting a link between enhanced O-GlcNAcylation and NF-κB activation in AML cell survival and proliferation.
Discussion
Our study suggests the HBP may prove a potential target, alone or in combination with other therapeutic approaches, to impact both AML blasts and stem cells. Moreover, as insufficient targeting of AML stem cells by traditional chemotherapy is thought to lead to relapse, blocking HBP and O-GlcNAcylation in AML stem cells may represent a novel promising target to control relapse.
... The NF-κB signaling pathway regulates cell proliferation in a variety of cell types [35,36]. Here, we determined the role of A1CF in two renal cancer cell lines (OSRC-2 and 786-O). ...
Apobec-1 complementation factor (A1CF) functions as an RNA-binding cofactor for APO-BEC1-mediated C-to-U conversion during RNA editing and as a hepatocyte-specific regulator in the alternative pre-mRNA splicing of metabolic enzymes. Its role in RNA editing has not been clearly established. Western blot, co-immunoprecipitation (Co-IP), immunofluorescence (IF), methyl thiazolyl tetrazolium (MTT), and 5-ethynyl-2′-deoxyuridine (EdU) assays were used to examine the role of A1CF beyond RNA editing in renal carcinoma cells. We demonstrated that A1CF interacts with NKRF, independent of RNA and DNA, without affecting its expression or nuclear translocation; however, it modulates p65(Ser536) phosphorylation and IFN-β levels. Truncation of A1CF or deletion on NKRF revealed that the RRM1 domain of A1CF and the p65 binding motif of NKRF are required for their interaction. Deletion of RRM1 on A1CF abrogates NKRF binding, and the decrease in IFN-β expression and p65(Ser536) phosphorylation was induced by A1CF. Moreover, full-length A1CF, but not an RRM1 deletion mutant, promoted cell proliferation in renal carcinoma cells. Perturbation of A1CF levels in renal carcinoma cells altered anchorage-independent growth and tumor progression in nude mice. Moreover, p65(Ser536) phosphorylation and IFN-β expression were lower, but ki67 was higher in A1CF-overexpressing tumor tissues of a xenograft mouse model. Notably, primary and metastatic samples from renal cancer patients exhibited high A1CF expression, low p65(Ser536) phosphorylation, and decreased IFN-β levels in renal carcinoma tissues compared with the corresponding paracancerous tissues. Our results indicate that A1CF-decreased p65(Ser536) phosphorylation and IFN-β levels may be caused by A1CF competitive binding to the p65-combined site on NKRF and demonstrate the direct binding of A1CF independent of RNA or DNA in signal pathway regulation and tumor promotion in renal carcinoma cells.
... With regard to NF-κB's effect on cell proliferation and survival in lung cancer, NF-κB signaling activates cyclin D1 by binding to its promoter, promoting cell proliferation [97]. Cyclin-dependent kinases (CDKs) are key in regulating the cell cycle, as they interact with cyclin proteins [106]. ...
Simple Summary
Despite massive strides taken across the board in oncology, there remain gaps in understanding the relationship between cancer cells and the body’s immune system, tissues, and signaling pathways. This review explores some of the recent steps made toward filling these gaps and understanding how certain cytokine signals create a tumor microenvironment that facilitates the growth and survival of cancer cells. New treatment approaches targeting these facilitators have been developed as potential disruptors of tumor growth. Lastly, a discussion of current gaps in the research can help navigate new directions to continue life-saving research for lung cancer treatments.
Abstract
The intricate interplay between inflammatory processes and the tumor microenvironment (TME) in lung cancer has garnered increasing attention due to its implications for both oncogenesis and therapeutic strategies. In this review, we explore recent advances in understanding the paracrine regulation and immune system pathways within the inflammatory TME of lung cancer. We delve into the molecular mechanisms underpinning oncogenesis, highlighting the role of immune cell populations, cancer-associated fibroblasts, and endothelial cells, as well as their interactions through immune system pathways regulated in a paracrine pattern. Additionally, we discuss emerging immunotherapeutic strategies with a specific focus on the potential of leveraging the inflammatory TME through these pathways to enhance treatment efficacy in lung cancer.
... Next, since activation of NF-κB pathway can promote proliferation (Guttridge et al., 1999), we questioned whether Y401D-transfected cells had a defective NF-κB response. Expectedly, we observed more than 50% attenuation of NF-κB activity in Y401D-transfected cells compared to the ones transfected with wild-type TNFR1 (Figure 5d). ...
Background/aim
Tumor necrosis factor alpha (TNFα, a.k.a. TNF) is a pleiotropic cytokine that exerts most of its effects through type 1 TNF receptor (TNFR1). Following TNF binding, TNFR1 recruits TRADD (tumor necrosis factor receptor type 1-associated DEATH domain). This interaction triggers formation of signalosome complexes which have been claimed to induce apoptosis (via downstream caspase activations), inflammation (via NF-kappaB) and stress pathways (JNK & p38). However, the mechanism underlying TNF-induced ERK and AKT activation is not completely revealed. TNFR1 is known to constitutively bind c-Src and JAK2, and these enzymes were previously demonstrated to modulate TNF signaling. Therefore, we hypothesized that TNFR1 could be tyrosine phosphorylated by JAK2 and/or c-Src and TNF-induced ERK and Akt activation may be mediated by this phosphorylation.
Materials and methods
Site-directed mutagenesis (SDM) was performed to substitute the two putative Tyrosine phosphorylation sites on TNFR1 (Y360 and Y401) with alanine (A) or with aspartic acid (D), to inhibit or mimic constitutive phosphorylation, respectively. In 293T cells transfected with mutated or wild type TNFR1, ERK and Akt activations were determined by western blot. TNFR1 interaction with c-Src, JAK2, p85 and Grb2 was examined by co-IP. NF-kB activation was measured by luciferase assay, while proliferation was measured by MTT and apoptosis was evaluated by colorimetric caspase 8/3 assays. For determination of necrosis rates, cellular DNA fragmentation ELISA was performed.
Results
In this report, we show that TNFR1 is phosphorylated by JAK2 tyrosine kinase at Y401 and by c-Src at Y360 and Y401. Phosphorylation of Y360 and Y401 augments the interaction of Grb2 and PI3Kp85 with TNFR1. We also demonstrate that phosphomimetic mutations of Y360D and Y401D enhance ERK and Akt activation.
Conclusion
TNFR1 is tyrosine phosphorylated by both c-Src and JAK2, triggering a “noncanonical” pathway, that activates ERK and Akt.
... One mechanism through which cyclin D1 expression is regulated is by direct binding of the nuclear factor kappa B (NFκB) transcription factor family to this promoter, thereby controlling gene transcription [10]. The NFκB signaling pathway is involved in many cellular processes, including immune/inflammatory responses, cell proliferation, cell growth, cell survival, and migration [11]. ...
Molecular targeted therapy using a drug that suppresses the growth and spread of cancer cells via inhibition of a specific protein is a foundation of precision medicine and treatment. High expression of the proto-oncogene Bcl-3 promotes the proliferation and metastasis of cancer cells originating from tissues such as the colon, prostate, breast, and skin. The development of novel drugs targeting Bcl-3 alone or in combination with other therapies can cure these patients or prolong their survival. As a proof of concept, in the present study, we focused on metastatic melanoma as a model system. High-throughput screening and in vitro experiments identified BCL3ANT as a lead molecule that could interfere with Bcl-3-mediated cyclin D1 expression and cell proliferation and migration in melanoma. In experimental animal models of melanoma, it was demonstrated that the use of a Bcl-3 inhibitor can influence the survival of melanoma cells. Since there are no other inhibitors against Bcl-3 in the clinical pipeline for cancer treatment, this presents a unique opportunity to develop a highly specific drug against malignant melanoma to meet an urgent clinical need.
Supplementary Information
The online version contains supplementary material available at 10.1186/s12885-023-11663-y.
... Cell cycle arrest results from the suppression of the production of cyclinD1, a crucial regulator of the G1-S cell cycle transition [23]. Moreover, cyclinD1 is a target protein of the NF-κB pathway, and when the NF-κB pathway is inhibited, cyclinD1 is down-regulated causing the cells to block in G1 phase [24]. We therefore suggest that the reason for this discrepancy may be that overexpression of keap1 in hucMSCs down-regulates IKKβ and inhibits the activation of the NF-κB pathway, thereby inhibiting cell cycle progression. ...
Background
HucMSCs had shown promising efficacy in treating childhood diseases, but oxidative stress induced by the poor microenvironment at the site of damage resulted in low cell survival after transplantation, thus preventing the cells from maximizing therapeutic efficacy. Therefore, this study aimed to investigate the role and mechanism of keap1 in oxidative stress injury of human umbilical cord mesenchymal stem cells (hucMSCs), and to provide theoretical support for improving the efficacy of stem cell therapy.
Methods
The hucMSCs were treated with hypoxic low-sugar-free serum (GSDH) to mimic the damaged site microenvironment after implantation. Adenoviral overexpression of keap1 gene of hucMSCs was performed in vitro, and cell proliferation ability was detected by CCK8 assay, crystal violet staining assay, and cell cycle assay. Cellular redox level was assessed by Amplex Red, MDA, and GSH/GSSG kit. Mitochondrial morphology was evaluated by mitotracker Red staining. ATP production was estimated by ATP detection kit. The mRNA and protein expression levels were tested by western blotting and RT-qPCR.
Results
GSDH treatment substantially upregulated keap1 expression. Subsequently, we found that overexpression of keap1 notably inhibited cell proliferation and caused cells to stagnate in G1 phase. At the same time, overexpression of keap1 induced the production of large amounts of H2O2 and the accumulation of MDA, but suppressed the GSH/GSSG ratio and the expression of antioxidant proteins NQO1 and SOD1, which caused oxidative stress damage. Overexpression of keap1 induced cells to produce a large number of dysfunctional mitochondria resulting in reduced ATP production. Moreover, Overexpression of keap1 significantly decreased the IKKβ protein level, while upregulating IkB mRNA levels and downregulating P50 mRNA levels.
Conclusions
Overexpression of keap1 may induce oxidative stress injury in hucMSCs by down-regulating IKKβ expression and inhibiting NF-κB pathway activation. This implies the importance of keap1 in hucMSCs and it may be a potential gene for genetic modification of hucMSCs.
... That is, it might repress transcription under normal conditions, but induce transcription under stress conditions by the recruitment of as-yet unknown cofactors. To clarify this hypothetical, we cite the activation of NF-κB to maintain myogenic cells in an undifferentiated state, whereas it acts as a repressor during myocyte differentiation [43,44]. In another example, the role of Myc protein in the transcriptional activation and repression of target genes may depend on the unique factors it recruits, such as the constellation of other transcription factors proximal to the binding site, or even the chromatin context of the target gene [45][46][47]. ...
ENDOU-1 encodes an endoribonuclease that overcomes the inhibitory upstream open reading frame (uORF)-trap at 5′-untranslated region (UTR) of the CHOP transcript, allowing the downstream coding sequence of CHOP be translated during endoplasmic reticulum (ER) stress. However, transcriptional control of ENDOU-1 remains enigmatic. To address this, we cloned an upstream 2.1 kb (−2055~+77 bp) of human ENDOU-1 (pE2.1p) fused with reporter luciferase (luc) cDNA. The promoter strength driven by pE2.1p was significantly upregulated in both pE2.1p-transfected cells and pE2.1p-injected zebrafish embryos treated with stress inducers. Comparing the luc activities driven by pE2.1p and −1125~+77 (pE1.2p) segments, we revealed that cis-elements located at the −2055~−1125 segment might play a critical role in ENDOU-1 upregulation during ER stress. Since bioinformatics analysis predicted many cis-elements clustered at the −1850~−1250, we further deconstructed this segment to generate pE2.1p-based derivatives lacking −1850~−1750, −1749~−1650, −1649~−1486, −1485~−1350 or −1350~−1250 segments. Quantification of promoter activities driven by these five internal deletion plasmids suggested a repressor binding element within the −1649~−1486 and an activator binding element within the −1350~−1250. Since luc activities driven by the −1649~−1486 were not significantly different between normal and stress conditions, we herein propose that the stress-inducible activator bound at the −1350~−1250 segment makes a major contribution to the increased expression of human ENDOU-1 upon ER stresses.
... The NF-κB signaling pathway controls the expression of proinflammatory cytokines and anti-infective factors, including TNF-α, interleukin-1 (IL-1), IL-6, IL-8, adhesion molecules, and cc chemokine ligand 5 (CCL5) [89]. In addition, NF-κB signaling pathway governs cellular processes such as cell proliferation, differentiation, and apoptosis [90,91]. ...
DEAD-box decapping enzyme 20 (DDX20) is a putative RNA-decapping enzyme that can be identified by the conserved motif Asp–Glu–Ala–Asp (DEAD). Cellular processes involve numerous RNA secondary structure alterations, including translation initiation, nuclear and mitochondrial splicing, and assembly of ribosomes and spliceosomes. DDX20 reportedly plays an important role in cellular transcription and post-transcriptional modifications. On the one hand, DDX20 can interact with various transcription factors and repress the transcriptional process. On the other hand, DDX20 forms the survival motor neuron complex and participates in the assembly of snRNP, ultimately affecting the RNA splicing process. Finally, DDX20 can potentially rely on its RNA-unwinding enzyme function to participate in microRNA (miRNA) maturation and act as a component of the RNA-induced silencing complex. In addition, although DDX20 is not a key component in the innate immune system signaling pathway, it can affect the nuclear factor kappa B (NF-κB) and p53 signaling pathways. In particular, DDX20 plays different roles in tumorigenesis development through the NF-κB signaling pathway. This process is regulated by various factors such as miRNA. DDX20 can influence processes such as viral replication in cells by interacting with two proteins in Epstein–Barr virus and can regulate the replication process of several viruses through the innate immune system, indicating that DDX20 plays an important role in the innate immune system. Herein, we review the effects of DDX20 on the innate immune system and its role in transcriptional and post-transcriptional modification processes, based on which we provide an outlook on the future of DDX20 research in innate immunity and viral infections.
... Cell cycle arrest results from the suppression of the production of cyclinD1, a crucial regulator of the G1-S cell cycle transition 23 . Moreover, cyclinD1 is a target protein of the NF-κB pathway, and when the NF-κB pathway is inhibited, cyclinD1 is down-regulated causing the cells to block in G1 phase 24 . We therefore suggest that the reason for this discrepancy may be that overexpression of keap1 in hucMSCs down-regulates IKKβ and inhibits the activation of the NF-κB pathway, thereby inhibiting cell cycle progression. ...
Background: HucMSCs had shown promising efficacy in treating childhood diseases, but oxidative stress induced by the poor microenvironment at the site of damage resulted in low cell survival after transplantation, thus preventing the cells from maximizing therapeutic efficacy. Therefore, this study aimed to investigate the role and mechanism of keap1 in oxidative stress injury of human umbilical cord mesenchymal stem cells (hucMSCs), and to provide theoretical support for improving the efficacy of stem cell therapy.
Methods: The hucMSCs were treated with hypoxic low-sugar-free serum (GSDH) to mimic the damaged site microenvironment after implantation. Adenoviral overexpression of keap1 gene of hucMSCs was performed in vitro, and cell proliferation ability was detected by CCK8 assay, crystal violet staining assay, and cell cycle assay; cellular redox level was assessed by Amplex Red, MDA, and GSH/GSSG kit; mitochondrial morphology was evaluated by mitotracker Red staining; ATP production was estimated by ATP detection kit; and the mRNA expression level of related proteins and related genes was tested by western blotting and RT-qPCR.
Results: GSDH treatment substantially upregulated keap1 mRNA levels and protein levels. Subsequently, we found that overexpression of keap1 notably inhibited cell proliferation and caused cells to stagnate in G1 phase. At the same time, overexpression of keap1 induced the production of large amounts of H2O2 and the accumulation of MDA, but suppressed the GSH/GSSG ratio and the expression of antioxidant proteins NQO1 and SOD1, which caused oxidative stress damage. Overexpression of keap1 induced cells to produce a large number of dysfunctional mitochondria resulting in reduced ATP production. Moreover, Overexpression of keap1 significantly decreased the IKKβ protein level，while upregulating IkB mRNA levels and downregulating P50 mRNA levels.
Conclusions: Overexpression of keap1 may induce oxidative stress injury in hucMSCs by down-regulating IKKβ expression and inhibiting NF-κB pathway activation. This implies the importance of keap1 in hucMSCs and it may be a potential gene for genetic modification of hucMSCs.
... Finally, NF-kB promotes cell growth: activated NF-kB enhances the expression of cyclin D1, an important regulator of cell cycle progression. NF-kB upregulated Myc and cycle regulator proteins, cyclin D1, and cyclin-dependent kinase 2, which further promoted melanoma growth (37)(38)(39)(40). Activation of NF-kB enhances tumor cell survival and proliferation and helps to transform tumorassociated macrophages into tumor-promoting phenotypes (41). ...
Background and objective
Existing cross-sectional and retrospective studies were unable to establish a causal relationship between psoriasis and cutaneous melanoma (CM). We sought to evaluate the causal role between psoriasis and CM.
Methods
We performed a bidirectional two-sample Mendelian randomization analysis using summary statistics from genome-wide association studies of psoriasis and CM among individuals of predominantly European ancestry. Mendelian randomization–Egger regression, inverse variance weighting, Mendelian Randomization Pleiotropy RESidual Sum and Outlier, weighted mode, and weighted median were used to examine the causal effect between psoriasis and CM.
Results
Genetically predicted psoriasis was a significant risk factor for CM (odds ratio, 1.69; 95% confidence interval, 1.15–2.48; P = 0.025). In contrast, no association was observed between genetically predicted CM and psoriasis.
Conclusion
Our findings corroborated the existence of genetically predicted psoriasis increases risk of CM. Enhanced early screening of cutaneous melanoma in patients with psoriasis may improve clinical burden. However, we did not find evidence for a causal link from CM to psoriasis, so further studies are required to elucidate the effect of CM activity on psoriasis.
... Cells 2023, 12, 1792 2 of 17 data. Indeed, most authors reported that ROS can cause inhibition of myogenic differentiation [3-8] by (i) NF-kB activation, which in turn causes a reduction in MyoD levels [9]; (ii) increased cyclin D1 transcription and cell proliferation [10]; and (iii) enhanced expression of YY1, a transcriptional repressor of myogenic genes [11]. Moreover, it has also been demonstrated that higher amounts of ROS might target the mitochondria and mitochondrial DNA, inducing the block of myogenesis [12,13]. ...
Reactive oxygen species (ROS) are currently recognized as a key driver of several physiological processes. Increasing evidence indicates that ROS levels can affect myogenic differentiation, but the molecular mechanisms still need to be elucidated. Protein kinase C (PKC) epsilon (PKCe) promotes muscle stem cell differentiation and regeneration of skeletal muscle after injury. PKCs play a tissue-specific role in redox biology, with specific isoforms being both a target of ROS and an up-stream regulator of ROS production. Therefore, we hypothesized that PKCe represents a molecular link between redox homeostasis and myogenic differentiation. We used an in vitro model of a mouse myoblast cell line (C2C12) to study the PKC–redox axis. We demonstrated that the transition from a myoblast to myotube is typified by increased PKCe protein content and decreased ROS. Intriguingly, the expression of the antioxidant enzyme superoxide dismutase 2 (SOD2) is significantly higher in the late phases of myogenic differentiation, mimicking PKCe protein content. Furthermore, we demonstrated that PKCe inhibition increases ROS and reduces SOD2 protein content while SOD2 silencing did not affect PKCe protein content, suggesting that the kinase could be an up-stream regulator of SOD2. To support this hypothesis, we found that in C2C12 cells, PKCe interacts with Nrf2, whose activation induces SOD2 transcription. Overall, our results indicate that PKCe is capable of activating the antioxidant signaling preventing ROS accumulation in a myotube, eventually promoting myogenic differentiation.
... Testing this hypothesis will require additional studies. NF-κB also directly regulates c-MYC, which plays important roles in cell growth and proliferation [43,44]. Thus, our previous observation that c-MYC is downregulated by FABP5 knockdown in CRC cells [20] could be partly explained by a reduction in NF-κB signaling resulting from suppression of the NF-κB-FABP5 pathway. ...
Fatty acid-binding proteins (FABPs) are intracellular lipid-binding proteins that play roles in fatty acid transport and the regulation of gene expression. Dysregulated FABP expression and/or activity have been associated with cancer pathogenesis; in particular, epidermal-type FABP (FABP5) is upregulated in many types of cancer. However, the mechanisms regulating FABP5 expression and its involvement in cancer remain largely unknown. Here, we examined the regulation of FABP5 gene expression in non-metastatic and metastatic human colorectal cancer (CRC) cells. We found that FABP5 expression was upregulated in metastatic compared with non-metastatic CRC cells as well as in human CRC tissues compared with adjacent normal tissue. Analysis of the DNA methylation status of the FABP5 promoter showed that hypomethylation correlated with the malignant potential of the CRC cell lines. Moreover, FABP5 promoter hypomethylation also correlated with the expression pattern of splice variants of the DNA methyltransferase DNMT3B. ChIP assays and luciferase reporter assays demonstrated that the transcription factor nuclear factor-kappa B (NF-κB) was involved in regulating FABP5 expression. FABP5 expression could be upregulated in metastatic CRC cells by sequential promotion of DNA demethylation followed by activation of NF-κB. We also found that upregulated FABP5 in turn controlled NF-κB activity through IL-8 production. Collectively, these findings suggest the existence of a DNA methylation-dependent NF-κB /FABP5 positive feed-forward loop that may lead to constitutive activation of NF-κB signaling pathway and play a crucial role in CRC progression.
... Moreover, the persistent infection can induce mutations in molecules, such as EGFR or RAS, which are upstream in the cascade of NF-κB, affecting its function. This altered activity induces the expression of genes, such as telomerase and c-myc, among others, which induce cell immortalization and proliferation, as well as metastasis (epithelial-mesenchymal transition) and angiogenesis [318,328]. Its reactivation induces expression of the AID/APOBEC (activation-induced cytokine deaminase) protein family known to participate in cancer development by causing genomic damage [329]. ...
The Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathway constitutes a rapid signaling module from the cell surface to the nucleus, and activates different cellular responses, such as proliferation, survival, migration, invasion, and inflammation. When the JAK/STAT pathway is altered, it contributes to cancer progression and metastasis. STAT proteins play a central role in developing cervical cancer, and inhibiting the JAK/STAT signaling may be necessary to induce tumor cell death. Several cancers show continuous activation of different STATs, including cervical cancer. The constitutive activation of STAT proteins is associated with a poor prognosis and overall survival. The human papillomavirus (HPV) oncoproteins E6 and E7 play an essential role in cervical cancer progression, and they activate the JAK/STAT pathway and other signals that induce proliferation, survival, and migration of cancer cells. Moreover, there is a crosstalk between the JAK/STAT signaling cascade with other signaling pathways, where a plethora of different proteins activate to induce gene transcription and cell responses that contribute to tumor growth. Therefore, inhibition of the JAK/STAT pathway shows promise as a new target in cancer treatment. In this review, we discuss the role of the JAK/STAT pathway components and the role of the HPV oncoproteins associated with cellular malignancy through the JAK/STAT proteins and other signaling pathways to induce tumor growth.
... The cyclin D1 and Cyclin D, and CDK4 and CDK6 are overexpressed in BC. Guttridge et al. [94] determined the essence of the association between NF-κB and cyclin D, whereby NF-κB regulates growth and progression via cyclin D1 (Fig. 4). It was also shown that RANK (receptor activator of NF-κB) induced the NF-κB activation in mammary epithelial cells, where cyclin D1 appearance is significantly upregulated [95]. ...
Breast cancer (BC) is the second most fatal disease and is the prime cause of cancer allied female deaths. BC is caused by aberrant tumor suppressor genes and oncogenes regulated by transcription factors (TFs) like NF-κB. NF-κB is a pro-inflammatory TF that crucially alters the expressions of various genes associated with inflammation, cell progression, metastasis, and apoptosis and modulates a network of genes that underlie tumorigenesis. Herein, we focus on NF-κB signaling pathways, its regulators, and the rationale for targeting NF-κB. This review also includes TFs that maintain NF-κB crosstalk and their roles in promoting angiogenesis and metastasis. In addition, we discuss the importance of combination therapies, resistance to treatment, and potential novel therapeutic strategies including nanomedicine that targets NF-κB.
... The NF-κB signaling pathway is one of the important pathways involved in cell cycle regulation (37). NF-κB can activate the transcription of cell cycle-related genes, including cyclin D1, cyclin E1, CDK2, CDK2 and Myc, thereby modulating cell cycle distribution (38)(39)(40)(41). The NF-κB pathway also contributes to maintaining CSC-like properties in multiple types of cancer (42)(43)(44). ...
Cervical cancer (CC) is the most common human papillomavirus-related disease. Continuous activation of the NF-κB signaling pathway has been observed in CC. SHC binding and spindle associated 1 (SHCBP1) contributes to tumorigenesis and activation of the NF-κB pathway in multiple cancer types, while its function in CC remains unclear. In the present study, three Gene Expression Omnibus datasets were used to identify differentially expressed genes (DEGs) in CC. Loss- and gain-of-function experiments were performed using stable SHCBP1-silenced and SHCBP1-overexpressing CC cells. To further explore the molecular mechanism of SHCBP1 in CC, small interfering RNA targeting eukaryotic translation initiation factor 5A (EIF5A) was transfected into stable SHCBP1-overexpressing CC cells. The results demonstrated that SHCBP1 was an upregulated DEG in CC tissues compared with healthy control cervical tissues. Functional experiments revealed the pro-proliferative and pro-stemness role of SHCBP1 in CC cells (CaSki and SiHa cells), in vitro. Furthermore, the NF-κB signaling pathway in CC cells was activated by SHCBP1. Increases in cell proliferation, stemness and activation of NF-κB, induced by SHCBP1 overexpression in CC cells, were reversed by EIF5A knockdown. Taken together, the results indicated that SHCBP1 serves an important role in regulation of CC cell proliferation, self-renewal and activation of NF-κB via EIF5A. The present study demonstrated a potential molecular mechanism underlying the progression of CC.
... Nuclear factor kappa B (NF-κB) was first discovered by Ranjan Sen and David Baltimore in 1986 [6]. As its name suggests, NF-κB binds to the immunoglobulin κ light chain enhancer of activated B cells, regulating the induction of several target genes involved in the cell cycle, differentiation, activation, and apoptosis [7,8]. There are five different NF-κB subtypes: RelA (p65), RelB, c-Rel, p105 (which can be further cleaved and matured into p50), and p100 [9]. ...
... Because activation of RAGE, IL-33, p-p65, PCNA, and osteopontin play critical roles in promoting cell proliferation of osteoclasts and fibroblasts, and immune cell activation and infiltration. [25][26][27][28][29][30] The resveratrol-induced inhibition of these gene expression might result in reduced proliferation of osteoclasts and fibroblasts and suppressed myometrial infiltration, leading to attenuating the progression of adenomyosis. ...
Adenomyosis is a uterine condition in which endometrial glands and stroma are commonly pathologically observed in the myometrium. In this study, we sought to determine the effect of resveratrol on the progression of adenomyosis. Adenomyosis was induced in mice given tamoxifen neonatally. All mice were subjected to body weight measurement and hotplate testing every four weeks beginning four weeks after birth. All mice with adenomyosis were randomly separated into 3 groups at 16 weeks: untreated, low-dose resveratrol (25 mg/kg), and high-dose resveratrol (50 mg/kg). After 3 weeks of treatment, final hotplate test and body weight measurement were performed, and the uterine horn blood samples were collected. Adenomyosis in mice caused body weight loss and uterine weight gain, reduced hotplate latency, and progression of endometrial fibrosis. The underlying biological process could be coupled with the overexpression of many cells' proliferation and immune-regulation-related genes. Resveratrol treatment could slow the progression of adenomyosis by enhancing hotplate latency, lowering endometrial fibrosis, and restoring cell proliferation- and immune-regulation-associated gene expression levels in endometrium and plasma. However, resveratrol treatment also reduced the body weight and uterine weight. In conclusion, Resveratrol might be a potential compound for treating patients with adenomyosis.
... Nuclear factor kappa B (NF-κB) was first discovered by Ranjan Sen and David Baltimore in 1986 [6]. As its name suggests, NF-κB binds to the immunoglobulin κ light chain enhancer of activated B cells, regulating the induction of several target genes involved in the cell cycle, differentiation, activation, and apoptosis [7,8]. There are five different NF-κB subtypes: RelA (p65), RelB, c-Rel, p105 (which can be further cleaved and matured into p50), and p100 [9]. ...
Nuclear factor kappa B (NF-κB) signaling pathways progress inflammation and immune cell differentiation in the host immune response; however, the uncontrollable stimulation of NF-κB signaling is responsible for several inflammatory illnesses regardless of whether the conditions are acute or chronic. Innate immune cells, such as macrophages, microglia, and Kupffer cells, secrete pro-inflammatory cytokines, such as TNF-α, IL-6, and IL-1β, via the activation of NF-κB subunits, which may lead to the damage of normal cells, including neurons, cardiomyocytes, hepatocytes, and alveolar cells. This results in the occurrence of neurodegenerative disorders, cardiac infarction, or liver injury, which may eventually lead to systemic inflammation or cancer. Recently, ginsenosides from Panax ginseng, a historical herbal plant used in East Asia, have been used as possible options for curing inflammatory diseases. All of the ginsenosides tested target different steps of the NF-κB signaling pathway, ameliorating the symptoms of severe illnesses. Moreover, ginsenosides inhibit the NF-κB-mediated activation of cancer metastasis and immune resistance, significantly attenuating the expression of MMPs, Snail, Slug, TWIST1, and PD-L1. This review introduces current studies on the therapeutic efficacy of ginsenosides in alleviating NF-κB responses and emphasizes the critical role of ginsenosides in severe inflammatory diseases as well as cancers.
... In recent years, the activation of the NF-κB pathway signaling in cell growth has been well documented. Moreover, two NF-κB binding sites at the CCNB1 promoter have been identified [25,26]. Actually, the regulatory effect of GRPEL2 on the NF-κB signaling pathway was also observed in our study. ...
Hepatocellular carcinoma (HCC) continues to cause severe burden worldwide. The limited options especially toward HCC with metastasis prompts us to identify novel molecules for either diagnostic/prognostic or therapeutic purposes. GRPEL2 is well defined in maintaining mitochondrial homeostasis, which is critical to multiple biological processes for cancer survival. However, its role in HCC progression was not investigated before. In our analysis using data from The Cancer Genome Atlas Liver Hepatocellular Carcinoma (TCGA LIHC) dataset and tissue microarray, higher expression levels of GRPEL2 were obseved in HCC tissues compared to in normal liver tissues, and indicated higher tumor grade, higher tumor stage, and shorter overall survival (OS). Consistent with the results of above analyses, the functional experiments validated that GRPEL2 acted as a tumor-promoting factor in HCC progression. GRPEL2 knockdown suppressed cell growth, migration, and invasion in vitro, as well as inhibited tumor growth in vivo. Moreover, GRPEL2 deficiency also accelerated reactive oxygen species (ROS) production and increased mitochondrial membrane potential (MMP), leading to cell apoptosis. In addition, we found that the cell cycle and NF-κB signaling pathways were responsible for GRPEL2-induced HCC progression, based on the results of Gene Set Enrichment Analysis (GSEA) and subsequent experimental validation. Our study, for the first time, identified the role of GRPEL2 in HCC development and provided a compelling biomarker for targted therapy in HCC treatment.
... La proliferación celular es un factor fundamental en el proceso celular, y el factor NF-κB también participa en esta actividad al activar genes que regulan la proliferación celular, como las ciclinas D1/D2/D3 (25) . Además, el factor NF-κB regula la proliferación celular por inducción de enzimas críticas, como la expresión de la proteína ubiquitina ligasa E3 Mdm-2, la cual es dependiente de NF-κB y afecta la estabilidad de p53 y la proliferación celular (26) . ...
The nuclear factor kappa B (NF-κB) family of transcription factors, which regulates a large range of genes in various immunological and inflammatory response pathways, is of utmost importance. This family consists of five structurally similar members that can activate target genes by attaching to particular regions of deoxyribonucleic acid (DNA). A class of inhibitory proteins usually keep NF-κB proteins in the cytoplasm; however, different oncogenic signaling pathways can activate them and cause malignant phenotypes in the appropriate cells. The main goal of this review article is to understand the regulatory mechanisms of NF-κB transcription factor, its pathogenesis and its potential cancer therapies. From the year 2000 to December 2022, several databases, including PubMed, Scopus and SciELO, were consulted. Finally, the review was developed by searching bibliographic references looking for the keywords related to NF-κB and cancer. The NF-κB transcription factor plays a key role in numerous cell signaling pathways, is involved in a number of biological functions, and its mutations have been linked to cancer and immunological disorders, among other pathologies. Since NF-κB is expressed in all cell types and tissues, many oncogenic mutations can activate NF-κB in tumor cells, opening up new research possibilities for the treatment of cancer. The canonical pathway and the alternative (non-canonical) pathway are two distinct NF-κB signaling pathways with various activation methods. NF-κB is involved in a variety of oncogenic pathways, including chronic inflammation, proliferation, apoptosis, angiogenesis, effect on cancer stem cells, metastasis, metabolic control and other related mechanisms. In conclusion, there are still many unanswered questions regarding the mechanisms and functions of NF-κB in the cellular context. A complete blockade of NF-κB does not appear to be a feasible strategy for the treatment of cancer at this time due to the variety of significant physiological actions that are altered by its blockade. Future research on NF-κB should focus on preventing cancer promotion while preserving the body’s natural physiological processes.
... It has been shown that IL-1β increases the expression of HIF-1α in lung cancer cells through an NF-κB-COX-2 pathway and by silencing miR-101 [45,54]. It's well known that NF-κB is a positive regulator of cell growth and proliferation in lung cells, through the upregulation of cyclin D1 and consequently enhanced transition from G1 to S phase [55,56]. In addition, it has been shown that HIF-1α is essential for the proliferation of NSCLC cells [57], once HIF-1α overexpression is correlated with c-Myc, a transcriptional factor associated with proliferation, overexpression in lung cancer specimens [58]. ...
Lung cancer is the leading cause of cancer-related deaths worldwide. The main risk factor for lung cancer is exposure to chemicals present in cigarettes and atmospheric pollutants, which, among other mechanisms, can increase the risk of cancer by inducing pulmonary inflammation. Among the complex features of inflammatory processes, the role of inflammasomes has attracted increasing attention due to their role in different stages of carcinogenesis. Inflammasomes are intracellular multiprotein complexes that when activated promote the maturation of interleukin-1beta (IL-1β) and IL-18, pro-inflammatory cytokines involved in the promotion, progression, epithelial-mesenchymal transition, metastasis, and resistance to therapy of lung cancer. In this way, this review summarizes the recent findings of inflammasome research in different stages of lung cancer, with a focus on non-small cell lung carcinoma (NSCLC), and highlights these multiprotein complexes as promising targets for cancer therapy.
... 62 Besides, NF-κB targets two other genes, MyoD and cyclin D, responsible for muscle cell differentiation. 63 During the post-burn flow phase, an inflammatory stress response is activated, and release of pro-inflammatory cytokines commences. Elevated concentrations of cytokines TNFα, IL-6 and IL-1 and interferon-γ (IFN-γ) increase expression of NF-κB and thus expression of atrogenes after burns ( Figure 5). ...
After a severe burn injury, a systemic stress response activates metabolic and inflammatory derangements that, among other, leads to muscle mass loss (muscle wasting). These negative effects on skeletal muscle continue for several months or years and are aggravated by short-term and long-term disuse. The dynamic balance between muscle protein synthesis and muscle protein breakdown (proteolysis) is regulated by complex signalling pathways that leads to an overall negative protein balance in skeletal muscle after a burn injury. Research concerning these molecular mechanisms is still scarce and inconclusive, understanding of which, if any, molecular mechanisms contribute to muscle wasting is of fundamental importance in designing of therapeutic interventions for burn patients as well. This review not only summarizes our present knowledge of the molecular mechanisms that underpin muscle protein balance but also summarizes the effects of exercise on muscle wasting post-burn as promising strategy to counteract the detrimental effects on skeletal muscle. Future research focusing on the pathways causing post-burn muscle wasting and the different effects of exercise on them is needed to confirm this hypothesis and to lay the foundation of therapeutic strategies.
... RNA-seq analysis identified that 476 genes were significantly up-regulated (> twofold change) and 716 genes were down-regulated (< 0.5-fold change) ( Fig. 3A and Additional file 4: Figure S4A). NF-κB signaling pathway caught our attention in KEGG analysis of pathways significantly under-represented in ZNF24 overexpressed lung cancer cells (Fig. 3B), as this pathway has been extensively reported to be involved in cell cycling [30][31][32]. Importantly, ZNF24 overexpression significantly inhibited transcription of target genes of NF-κB pathway, including IL-1β, IL-6 and TNFα in A549i and Hop62i cells (Fig. 3C, D), while its knockdown significantly activated their expression in EKVX cells (Fig. 3E). ...
Systemic identification of tumor suppressor genes (TSGs) and elucidation of their signaling provide a new angle for understanding of tumorigenesis, which is important for developing successful treatment for lung cancer patients. In our current work, we conducted an in vivo screen for lung cancer TSGs through CRISPR/Cas9 mediated knockout of genes at genome-wide scale. We found that ZNF24 was a potent and clinically relevant TSG of lung cancer. Ectopic expression of ZNF24 arrested lung cancer cells in S phase. Mechanistically, ZNF24 bound to promoter region of P65 to negatively regulate its transcription and thereby the signaling activity of NF-κB pathway. This signaling cascade is clinically relevant. Importantly, we found that combinational inhibition of KRAS, NF-κB, and PD-1 effectively shrank autochthonous Kras G12D / ZNF24 −/− lung cancers in transgenic mouse model. Our current work thus revealed an important role played by loss of function of ZNF24 in lung tumorigenesis and shed new light in precision medicine for a portion of lung cancer patients.
... The more the NF-κB repressed the more the activities of antioxidants enzymes in scavenging the free radicals increased and thereby reduced tumor formation. [43] As observed in this study, there is an increase in fragmented DNA in the mice exposed to DMBA only but there is reduction in the level of fragmented DNA in the group administered with Ocimum basilicum essential oil, increase in the level of fragmented DNA give an insight that the DNA damage as well as cell cycle might be dysregulated due to the increase in the number of fragmented DNA. ...
Background: Breast cancer is one of the most common cancers worldwide.7,12-dimethylbenz[a]anthracene (DMBA) is one of the xenobiotics that are implicated in various toxicological conditions including carcinogenesis. This study was conducted to assess the protective effect of Ocimum basilicum essential oil on Dimethylbenz[a]anthracene-induced breast tumor in mice.
Method: Sixty (60) adult female mice were randomized into five groups (A-E) each containing twelve (12) mice: Group A- Control received distilled water, Group B- DMBA induced only, Group C- DMBA induced+100mg/kg bodyweight (bwt) essential oil, Group D- DMBA induced+200mg/kg bwt essential oil and Group E- DMBA induced + Tamoxifen, respectively. The animals were treated with Ocimum basilicum essential oil for 14 days. At the end of 14 days, assays for serum catalase, superoxide dismutase, malondialdehyde, reduced glutathione and glutathione-s- transferase also IL-1β, TNF-α, NF-κB and fragmented DNA were done.
Result: Histopathology analysis of the skin of the mammary gland shows that DMBA-induced mice possessed enlarged skin tissue with epidermal necrosis and disruption of the entire layers of the skin, this suggesting tissue damage and cancer while positive control and group administered with Ocimum basilicum essential oil showed preserved epidermal, dermal and subcutaneous layers. There is increase in levels of IL-1β, TNF-α, NF-κB, fragmented DNA, SOD,CAT, GST activities and GSH level in the group administered with Ocimum basilicum essential oil when compared with the negative control.
... The Nuclear Factor "kappa-light-chain-enhancer" of activated B-cells (NF-κB) is an important immune defense transcription factor that mediates cytoplasmic/ nuclear signaling pathways [4] and regulates the gene expression of various cytokines and cytokine receptors and adhesion molecules in the inflammatory nuclear immune response [5]. It is also associated with apoptotic pathways, cell proliferation and differentiation, and resistance to chemotherapy/radiotherapy in tumors [6,7]. Therefore, targeting the NF-κB pathway may be an effective means of preventing cancer progression and improving chemotherapy sensitivity. ...
Background
The upregulated expression of CXCL1 has been validated in colorectal cancer patients. As a potential biotherapeutic target for colorectal cancer, the mechanism by which CXCL1 affects the development of colorectal cancer is not clear.
Methods
Expression data of CXCL1 in colorectal cancer were obtained from the GEO database and verified using the GEPIA database and the TIMER 2.0 database. Knockout and overexpression of CXCL1 in colorectal cancer cells by CRISPR/Cas and "Sleeping Beauty" transposon-mediated gene editing techniques. Cell biological function was demonstrated by CCK-8, transwell chamber and Colony formation assay. RT-qPCR and Western Blot assays measured RNA and protein expression. Protein localization and expression were measured by immunohistochemistry and immunofluorescence.
Results
Bioinformatics analysis showed significant overexpression of CXCL1 in the colorectal cancer tissues compared to normal human tissues, and identified CXCL1 as a potential therapeutic target for colorectal cancer. We demonstrate that CXCL1 promotes the proliferation and migration of colon cancer cells and has a facilitative effect on tumor angiogenesis. Furthermore, CXCL1 elevation promoted the migration of M2-tumor associated macrophages (TAMs) while disrupting the aggregation of CD4+ and CD8+ T cells at tumor sites. Mechanistic studies suggested that CXCL1 activates the NF-κB pathway. In the in vivo colon cancer transplantation tumor model, treatment with the P300 inhibitor C646 significantly inhibited the growth of CXCL1-overexpressing colon cancer.
Conclusion
CXCL1 promotes colon cancer development through activation of NF-κB/P300, and that CXCL1-based therapy is a potential novel strategy to prevent colon cancer development.
This book series invites all the Specialists, Professors, Doctors, Scientists, Academicians, Healthcare professionals, Nurses, Students, Researchers, Business Delegates, and Industrialists across the globe to publish their insights and convey recent developments in the field of Nursing, Pharmaceutical Research and Innovations in Pharma Industry. Book series on Pharmacy and Nursing covers research work in a set of clinical sciences and medicine.
The Ras–mitogen-activated protein kinase (MAPK) pathway is a major target for cancer treatment. To better understand the genetic pathways that modulate cancer cell sensitivity to MAPK pathway inhibitors, we performed a CRISPR knockout screen with MAPK pathway inhibitors on a colorectal cancer (CRC) cell line carrying mutant KRAS. Genetic deletion of the catalytic subunit of protein phosphatase 6 (PP6), encoded by PPP6C , rendered KRAS - and BRAF -mutant CRC and BRAF -mutant melanoma cells more resistant to these inhibitors. In the absence of MAPK pathway inhibition, PPP6C deletion in CRC cells decreased cell proliferation in two-dimensional (2D) adherent cultures but accelerated the growth of tumor spheroids in 3D culture and tumor xenografts in vivo. PPP6C deletion enhanced the activation of nuclear factor κB (NF-κB) signaling in CRC and melanoma cells and circumvented the cell cycle arrest and decreased cyclin D1 abundance induced by MAPK pathway blockade in CRC cells. Inhibiting NF-κB activity by genetic and pharmacological means restored the sensitivity of PPP6C -deficient cells to MAPK pathway inhibition in CRC and melanoma cells in vitro and in CRC cells in vivo. Furthermore, a R264 point mutation in PPP6C conferred loss of function in CRC cells, phenocopying the enhanced NF-κB activation and resistance to MAPK pathway inhibition observed for PPP6C deletion. These findings demonstrate that PP6 constrains the growth of KRAS - and BRAF -mutant cancer cells, implicates the PP6–NF-κB axis as a modulator of MAPK pathway output, and presents a rationale for cotargeting the NF-κB pathway in PPP6C -mutant cancer cells.
Diabetes represents a major public health concern with a considerable impact on human life and healthcare expenditures. It is now well established that diabetes is characterized by a severe skeletal muscle pathology that limits functional capacity and quality of life. Increasing evidence indicates that diabetes is also one of the most prevalent disorders characterized by impaired skeletal muscle regeneration, yet underlying mechanisms and therapeutic treatments remain poorly established. In this review, we describe the cellular and molecular alterations currently known to occur during skeletal muscle regeneration in people with diabetes and animal models of diabetes, including its associated comorbidities, e.g., obesity, hyperinsulinemia, and insulin resistance. We describe the role of myogenic and non-myogenic cell types on muscle regeneration in conditions with or without diabetes. Therapies for skeletal muscle regeneration and gaps in our knowledge are also discussed, while proposing future directions for the field.
NF-κB signaling has been discovered for nearly 40 years. Initially, NF-κB signaling was identified as a pivotal pathway in mediating inflammatory responses. However, with extensive and in-depth investigations, researchers have discovered that its role can be expanded to a variety of signaling mechanisms, biological processes, human diseases, and treatment options. In this review, we first scrutinize the research process of NF-κB signaling, and summarize the composition, activation, and regulatory mechanism of NF-κB signaling. We investigate the interaction of NF-κB signaling with other important pathways, including PI3K/AKT, MAPK, JAK-STAT, TGF-β, Wnt, Notch, Hedgehog, and TLR signaling. The physiological and pathological states of NF-κB signaling, as well as its intricate involvement in inflammation, immune regulation, and tumor microenvironment, are also explicated. Additionally, we illustrate how NF-κB signaling is involved in a variety of human diseases, including cancers, inflammatory and autoimmune diseases, cardiovascular diseases, metabolic diseases, neurological diseases, and COVID-19. Further, we discuss the therapeutic approaches targeting NF-κB signaling, including IKK inhibitors, monoclonal antibodies, proteasome inhibitors, nuclear translocation inhibitors, DNA binding inhibitors, TKIs, non-coding RNAs, immunotherapy, and CAR-T. Finally, we provide an outlook for research in the field of NF-κB signaling. We hope to present a stereoscopic, comprehensive NF-κB signaling that will inform future research and clinical practice.
Advanced glycation end products (AGEs) stochastically accrue in skeletal muscle and on collagen over an individual's lifespan, stiffening the muscle and modifying the stem cell (MuSC) microenvironment while promoting proinflammatory, anti-regenerative signaling via the receptor for advanced glycation end products (RAGE). In the present study, a novel in vitro model was developed of this phenomenon by cross-linking a 3D collagen scaffold with AGEs and investigating how myoblasts responded to such an environment. Briefly, collagen scaffolds were incubated with D-ribose (0, 25, 40, 100, or 250 mM) for 5 days at 37° C. C2C12 immortalized mouse myoblasts were grown on the scaffolds for 6 days in growth conditions for proliferation, and 12 days for differentiation and fusion. Human primary myoblasts were also used to confirm the C2C12 data. AGEs aberrantly extended the DNA production stage of C2C12s (but not in human primary myoblasts) which is known to delay differentiation in myogenesis, and this effect was prevented by RAGE inhibition. Further, the differentiation and fusion of myoblasts were disrupted by AGEs, which were associated with reductions in integrins and suppression of RAGE. The addition of S100b (RAGE agonist) recovered the differentiation and fusion of myoblasts, and the addition of RAGE inhibitors (FPS-ZM1 and Azeliragon) inhibited the differentiation and fusion of myoblasts. Our results provide novel insights into the role of the AGE-RAGE axis in skeletal muscle aging, and future work is warranted on the potential application of S100b as a pro-regenerative factor in aged skeletal muscle.
Objectives: Pulmonary tuberculosis (PTB) is a leading infectious disease and a major public health concern. Out of susceptible genes, nuclear factor kappa B ( NF-κB ) gene influences inflammation and hence the progression of the disease. A single nucleotide polymorphism of functional NF-κB1 promoter −94 ins/del ATTG (rs28362491) has a regulatory effect on the NF-κB1 gene. The aim of our study was to ascertain the bacterial load and to find the association with NF-κB gene polymorphism. Methodology : One hundred two diagnosed and microbiologically confirmed cases of PTB patients between 18 and 69 years of age and 102 healthy age, sex, and socioeconomic status–matched volunteers were taken as controls. Sputum from cases and blood samples from cases and controls were collected. Bacterial load estimation was done from the sputum sample. Genomic deoxyribonucleic acid (DNA) was extracted for NF-κB gene. Polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) were performed. Results: The genotypic frequency of ins/del was found to be 54.9% in PTB patients which was more than that of the other genotype frequencies in the case group and also from the control genotype frequencies, although it was found to be insignificant ( P value = 0.098). On analysis of bacterial load with NF-κB gene polymorphism, a significant association was found more particularly for ins/del versus del/del. Conclusion: The bacterial load showed significant variation among the del/del genotype, ins/del genotype, and ins/ins genotype which was statistically significant. The del/del genotype exhibited the highest bacterial load in and greater statistical significance was found between ins/del and del/del genotypes. This implies the association between NF-κB gene polymorphism and bacterial load.
Background:
New biomarkers of progression in patients with prostate cancer (PCa) are needed to improve their classification and clinical management. This systematic review investigated the relationship between single nucleotide polymorphisms (SNPs) and PCa progression.
Methods:
A keyword search was performed in Pubmed, EMBASE, Scopus, Web of Science, and Cochrane for publications between 2007 and 2022. We included articles with adjusted and significant associations, a median follow-up greater than or equal to 24 months, patients taken to radical prostatectomy (RP) as a first therapeutic option, and results presented based on biochemical recurrence (BCR).
Results:
In the 27 articles selected, 73 SNPs were identified in 39 genes, organized in seven functional groups. Of these, 50 and 23 SNPs were significantly associated with a higher and lower risk of PCa progression, respectively. Likewise, four haplotypes were found to have a significant association with PCa progression.
Conclusion:
This article highlights the importance of SNPs as potential markers of PCa progression and their possible functional relationship with some genes relevant to its development and progression. However, most variants were identified only in cohorts from two countries; no additional studies reproduce these findings.
Tumors in dogs and humans share many similar molecular and genetic features, incentivizing a better understanding of canine neoplasms not only for the purpose of treating companion animals, but also to facilitate research of spontaneously developing tumors with similar biologic behavior and treatment approaches in an immunologically competent animal model. Multiple tumor types of both species have similar dysregulation of signal transduction through phosphatidylinositol 3-kinase (PI3K), protein kinase B (PKB; AKT), and mechanistic target of rapamycin (mTOR), collectively known as the PI3K-AKT-mTOR pathway. This review aims to delineate the pertinent aspects of the PI3K-AKT-mTOR signaling pathway in health and in tumor development. It will then present a synopsis of current understanding of PI3K-AKT-mTOR signaling in important canine cancers and advancements in targeted inhibitors of this pathway.
Hsa‐microRNA‐215 (hsa‐miR‐215) plays multiple roles in carcinogenesis through regulating its target genes. Genetic variants in hsa‐miR‐215 target sites thus may affect hsa‐miR‐215‐mRNA interactions, result in altered expression of target genes and even influence cancer susceptibility. This study aimed to investigate the associations of genetic variants which located in the binding sites of hsa‐miR‐215 with non‐small cell lung cancer (NSCLC) susceptibility in the Chinese population and reveal the potential regulatory mechanism of functional variants in NSCLC development. The candidate genetic variants were predicted and screened through bioinformatics analysis based on the degree of complementarity of hsa‐miR‐215 sequences. The potential effects of genetic variants on the binding ability of hsa‐miR‐215 and target genes were also predicted. A case–control study with 932 NSCLC patients and 1036 healthy controls was conducted to evaluate the association of candidate genetic variants with NSCLC susceptibility, and an independent case–control study with 552 NSCLC cases and 571 controls were used to further validate the promising associations. Dual luciferase reporter gene assay was applied to explore the regulation of the genetic variants on transcription activity of target gene. Cell phenotyping experiments in vitro and RNA sequencing (RNA‐seq) were then carried out to preliminarily explore the potential regulatory mechanisms of the target genes in NSCLC. A total of five candidate genetic variants located in the binding sites of hsa‐miR‐215 were screened. The two‐stage case–control study showed that a variant rs1854268 A > T, which located in the 3' untranslated (3'UTR) region of NDUFAF4 gene, was associated with decreased risk of NSCLC (additive model, odds ratio [OR] = 0.83, 95% confidence interval [CI]: 0.75−0.92, p < 0.001). Functional annotation displayed that rs1854268 A > T might downregulate the expression of NDUFAF4 by enhancing the binding affinity of hsa‐miR‐215‐5p to NDUFAF4 mRNA. Additionally, transient knockdown of the NDUFAF4 could inhibit lung cancer cell migration and promote lung cancer cell apoptosis. Further RNA‐seq analysis revealed that the knockdown of NDUFAF4 may affect NSCLC development by downregulating the nuclear factor kappa B (NF‐κB) and phosphoinositide 3 kinase‐AKT (PI3K‐AKT) signaling pathways. Moreover, the overexpression of CCND1 could partially attenuate the effects of NDUFAF4 knock down on lung cancer cell migration and apoptosis, indicating that CCND1 may be involved in the tumor‐promoting effects of NDUFAF4 as a downstream molecule of NDUFAF4 gene. In conclusion, the genetic variant rs1854268 (A > T) on NDUFAF4 confers NSCLC susceptibility by altering the binding affinity of hsa‐miR‐215‐5p, thus regulating the expression of NDUFAF4 and subsequently influencing downstream tumor molecules and pathways such as CCND1 , NF kappa B, and PI3K‐AKT signaling pathways.
Zika virus (ZIKV) infection during the first trimester of the pregnancy may lead to Congenital zika syndrome in the neonates. The viral infection hampers foetal brain development and causes microcephaly. Human neural progenitor cells (hNPCs) play an important role in brain development, however they are highly susceptible to ZIKV infection. In this study, we elucidated the molecular mechanisms that lead to cellular alterations in hNPCs due to ZIKV E-protein. We investigated proliferation, differentiation, migration and inflammation in hNPCs, which may lead to microcephaly. In our study, we found that ZIKV E-protein causes cell cycle arrest, decrease in proliferation and increase in mitotic length of the dividing hNPCs. We observed CyclinD1 and upstream molecules (p21 and p53) of the pathway are dysregulated, and intracellular calcium at basal level as well as upon ATP stimulation were reduced following over expression of ZIKV E-protein. ZIKV E-protein transfected hNPCs exhibited pre-mature differentiation with pro-neural genes upregulated. Furthermore, ZIKV E-protein disrupted migrational properties of hNPCs and caused elevated levels of inflammatory chemokines and cytokines. To gain insights into molecular mechanisms of these effects on hNPCs, we explored the possible involvement of long non coding RNAs in ZIKV neuropathogenesis. We have shortlisted lncRNAs associated with differentially expressed genes from publicly available transcriptomic data and found some of those lncRNAs are differentially expressed upon E-protein transfection of hNPCs. Gene ontology analysis suggest these lncRNAs play an important role in regulation of viral life cycle, host's defence response and cell proliferation.
Colorectal cancer is the cancer of the colon, located at the lower part of the digestive system. Although the role of STAT3 in cancer is known, its role particularly in colon cancer is largely unknown. STAT3 is a cytoplasmic transcription factor that involves extracellular signaling to the nucleus regulating fundamental functions, like cell proliferation, apoptosis, differentiation, and angiogenesis. STAT3 is a key regulator; abrogated activation leads to several diseases, including cancer. Aberrant interleukin (IL)-6-mediated JAK/STAT3 signaling pathway is closely related to the advancement of several human solid tumors including colorectal cancer. With other upstream regulators, IL-6/JAK signaling can activate STAT3, and its role appears to be critical in various types of cancer. STAT3 has been traditionally recognized as an oncogene; more recently the dual role of STAT3 in cancer, either tumor inductive or suppressive, has been appreciated. This chapter describes the potential role of STAT3 in colon cancer based on in vitro, in vivo, and patient studies. Furthermore, we will discuss the mechanism of action and roles of the IL-6/JAK/STAT3 pathway in colorectal cancer and exploit current therapeutic strategies, to treat colorectal cancer. Understanding the complexity of STAT3 function in colorectal cancer has the potential to elucidate important molecular aspects of colorectal cancer with significant therapeutic implications.
Today, treatment options for cancer patients typically include surgery, radiation therapy, immunotherapy, and chemotherapy. While these therapies have saved lives and reduced pain and suffering, cancer still takes millions of lives every year around the world. Researchers are now developing advanced therapeutic strategies such as immunotherapy, targeted therapy, and combination nanotechnology for drug delivery. In addition, the identification of new biomarkers will potentiate early-stage diagnosis. Molecular Targets and Cancer presents information about cancer diagnosis and therapy in a simple way. It covers several aspects of the topic with updated information on par with medical board levels. The book features contributions from experts and includes an overview of cancer from basic biology and pathology, classifications, surveillance, prevention, diagnosis, types of cancer, treatment and prognosis. The first part of this book introduces the reader to cancer epidemiology, genetic alterations in cancer, exogenous and endogenous factors in carcinogenesis, roles for growth factors in cancer progression, cell signaling in cancer, transcription factors in cancer, and cancer genetics and epigenetics. This comprehensive guide is a valuable resource for oncologists, researchers, and all medical professionals who work in cancer care and research.
N6-methyladenosine (m6A) modification controls cell fate determination. Here, we show that liquid-liquid phase separation (LLPS) of YTH N6-methyladenosine RNA binding protein 1 (YTHDF1), a pivotal m6A "reader" protein, promotes the transdifferentiation of spermatogonial stem cells (SSCs) into neural stem cell-like cells by activating the IκB-nuclear factor κB (NF-κB)-CCND1 axis. The inhibition of IκBα/β mRNA translation mediated by YTHDF1 LLPS is the key to the activation of the IκB-NF-κB-CCND1 axis. Disrupting either YTHDF1 LLPS or NF-κB activation inhibits transdifferentiation efficiency. Moreover, overexpression of the YTH domain of YTHDF1 inhibits the activation of the IκB-NF-κB-CCND1 axis by promoting IκBα/β mRNA translation. Overexpression of the tau-YTH fusion protein reactivates IκB-NF-κB-CCND1 axis by inhibiting the translation of IκBα/β mRNAs, and tau LLPS is observed, which can restore transdifferentiation efficiency. Our findings demonstrate that the protein-RNA LLPS plays essential roles in cell fate transition and provide insights into translational medicine and the therapy of neurological diseases.
The success story of cisplatin spans over six decades now and yet it continues to be the key player in most chemotherapeutic regimens. Numerous efforts have been made to improve...
Insulin-like growth factor 1 receptor (IGF-1R) is an influential growth receptor in mammalian cells. Majority of cancers in humans show an active IGF-1R signaling pathway. Malus domestica apple peel extracts were investigated for their potential mechanisms of action against regulating cell proliferation in human breast cancer MDA-MB-231 cells under IGF-1 induction. Apple peel extracts significantly inhibited cellular proliferation in breast cancer cells in a dose-dependent manner at a dose ≥ 10 mg/mL. No cytotoxicity was observed in the human breast cancer MDA-MB-231 cells at the doses below 50.2 mg/mL. The IGF-1R/PI3K/Akt pathway expression was suppressed by apple peel extracts treatment. Expressions of cyclin D1-stimulating proteins NF-κ B and GSK-3β were inhibited, whereas p21 expression was upregulated. Apple peel extracts reduced the expression of cyclin D1, stagnating MDA-MB-231 cells in a G1 cell cycle arrest. Apple peel extracts stimulated PTEN, a tumor suppressor protein and an inhibitor of PI3K/Akt signaling pathway. The results suggested that apple peel extracts can induce inhibitory effects through downregulating IGF-1R/PI3K/Akt signal transduction pathway in human breast cancer MDA-MB-231 cells. This study shows the efficacy of apple peel extracts in reversing the IGF-1R signal transduction pathway, thereby hindering tumorigenesis in breast cancer cells.
To investigate the function of NF-κB RelA (p65), we generated mice deficient in this NF-κB family member by homologous recombination. Mice lacking RelA showed liver degeneration and died around embryonic day 14.5. To elucidate the role of RelA in lymphocyte development and function, we transplanted fetal liver cells of 13.5-day embryos from heterozygote matings into irradiated SCID mice. Within 4 weeks, both T and B cells had developed in the SCID mice receiving relA−/− fetal liver transplants, similar to the relA+/+ and +/− cases. T cells were found to mature to Thy-1+/TCRαβ+/CD3+/CD4+ or CD8+, while B cells had the ability to differentiate to IgM+/B220+ and to secrete immunoglobulins. However, the secretion of IgG1 and IgA was reduced in RelA-deficient B cells. Furthermore, both T and B cells lacking RelA showed marked reduction in proliferative responses to stimulation with Con A, anti-CD3, anti-CD3+anti-CD28, LPS, anti-IgM, and PMA+calcium ionophore. The results indicate that RelA plays a critical role in production of specific Ig isotypes and also in signal transduction pathways for lymphocyte proliferation.
Ectopic overexpression of v-H-Ras protein in NIH 3T3 cells resulted in cellular transformation and an
acceleration of G1 progression of these cells. A shortened G1 phase was found to be associated with an
increased level of cyclin D1 but not cyclin E protein. Using an antisense blocking method, reduced synthesis
of cyclin D1 in v-H-Ras transformants resulted in a slower G1 progression rate of these cells. Although
constitutive overexpression of cyclin D1 in NIH 3T3 cells accelerated G1 progression, cells remained untransformed.
Furthermore, inhibition of cyclin D1 synthesis greatly impaired the soft-agar cloning efficiency of
v-H-Ras transformants. These results suggest that increased expression of cyclin D1 is necessary but not
sufficient for the transforming activity of v-H-Ras. Similar effect on cell cycle progression was also observed in
Raf-transformed cells. In addition to cyclin D1, cyclin E protein was found to be elevated in Src transformants.
This may account for the further shortening of the G1 phase of these cells. Activation of an additional
Ras-independent pathway was suggested to be responsible for the further acceleration of the G1 phase in Src
transformants.
The nfkbl and nfkb2 genes encode closely related products regulating immune and inflammatory responses1−3. Their role during development and differentiation remains unclear. The generation of nfkb1 null mice (p50-/-) resulted in altered immune responses, but had no effect on development4. Similarly, nfkb2 knockout mice (p52-/-) did not show developmental defects (J.C. et al., manuscript submitted). We have investigated the potential for in vivo compensatory functions of these genes by generating double-knockout mice. The surprising result was that the animals developed osteopetrosis because of a defect in osteoclast differentiation, suggesting redundant functions of NF-B1 and NF-B2 proteins in the development of this cell lineage. The osteopetrotic phenotype was rescued by bone marrow transplantation, indicating that the hematopoietic component was impaired. These results define a new mouse osteopetrotic mutant and implicate NF-B proteins in bone development, raising new directions in the treatment of bone disorders.
Several different oncogenes and growth factors promote G1 phase progression. Cyclin D1, the regulatory subunit of several cyclin-dependent kinases, is required for, and capable of
shortening, the G1 phase of the cell cycle. The present study demonstrates that transforming mutants of p21 (Ras Val-12, Ras Leu-61) induce the cyclin D1 promoter in human trophoblasts (JEG-3), mink lung epithelial (Mv1.Lu), and
in Chinese hamster ovary fibroblast cell lines. Site-directed mutagenesis of AP-1-like sequences at −954 abolished p21-dependent activation of cyclin D1 expression. The AP-1-like sequences were also required for activation of the cyclin D1
promoter by c-Jun. In electrophoretic mobility shift assays using nuclear extracts from cultured cells and primary tissues,
several AP-1 proteins (c-Jun, JunB, JunD, and c-Fos) bound the cyclin D1 −954 region. Cyclin D1 promoter activity was stimulated
by overexpression of mitogen-activated protein kinase (p41) or c-Ets-2 through the proximal 22 base pairs. Expression of plasmids encoding either dominant negative MAPK (p41) or dominant negatives of ETS activation (Ets-LacZ), antagonized MAPK-dependent induction of cyclin D1 promoter activity.
Epidermal growth factor induction of cyclin D1 transcription, through the proximal promoter region, was antagonized by either
p41 or Ets-LacZ, suggesting that ETS functions downstream of epidermal growth factor and MAPK in the context of the cyclin D1
promoter. The activation of cyclin D1 transcription by p21provides evidence for cross-talk between the p21 and cell cycle regulatory pathways.
Expression of the fos family of transcription factors is stimulated by growth factors that induce quiescent cells to reenter the cell cycle, but the cellular targets of the Fos family that regulate cell cycle reentry have not been identified. To address this issue, mice that lack two members of the fos family, c-fos and fosB, were derived. The fosB−/− c-fos−/− mice are similar in phenotype to c-fos−/− mice but are 30% smaller. This decrease in size is consistent with an abnormality in cell proliferation. Fibroblasts derived from fosB−/− c-fos−/− mice were found to have a defect in proliferation that results at least in part from a failure to induce cyclin D1 following serum-stimulated cell cycle reentry. Although definitive evidence that c-Fos and FosB directly induce cyclin D1 transcription will require further analysis, these findings raise the possibility that c-Fos and FosB are either direct or indirect transcriptional regulators of the cyclin D1 gene and may function as a critical link between serum stimulation and cell cycle progression.
The nuclear form of the NF-kappa B transcription factor binds to DNA as a heterodimer of a 50 kDa (p50) and 65 kDa (p65) polypeptide. The two polypeptides are encoded by different genes but share a long region of homology, the NRD motif, encompassing domains required for DNA binding and dimerization. In this study we have analysed the contribution of the two subunits to the strong transactivating potential of NF-kappa B. Transient expression of the p65 subunit alone resulted in a potent transactivation of a CAT reporter construct under the control of two NF-kappa B binding sites in monkey COS and mouse L cells. The strongly DNA binding p50 subunit showed only very weak, if any, induction of gene expression. Co-expression of p50 suppressed the transactivation by p65 presumably by competitive DNA binding of transcriptionally inactive p50 dimers (KBF1). Fusion of p65 sequences to DNA binding domain of the yeast GAL4 transcription factor allowed detection of the principal transactivation domain of p65 (TA1) in the C-terminal 30 amino acid sequence. TA1 is likely to adopt an amphipathic alpha-helical structure which clusters serine residues on the hydrophilic surface, a structural feature conserved between human, mouse and Xenopus p65. The unique C-terminal third of p65 contained at least one more activation domain, TA2, within a 90 amino acid sequence directly adjacent to TA1. In two mammalian cell lines, TA1 and TA2 acted separately, while in an insect cell line, the two domains were inactive after their separation. Our study suggests that the p50 subunit in NF-kappa B might only serve a helper function in DNA binding whereas the p65 subunit is responsible for initiating transcription. Homodimers of p50 seem to have the potential of down-regulating kappa B-specific gene expression.
Differentiation of skeletal muscle involves withdrawal of myoblasts from the cell cycle, fusion to form myotubes, and the coordinate expression of a variety of muscle-specific gene products. Fibroblast growth factor and type beta transforming growth factor specifically inhibit myogenesis; however, the transmembrane signaling pathways responsible for suppression of differentiation by these growth factors remain elusive. Because ras proteins have been implicated in the transduction of growth factor signals across the plasma membrane, we used DNA-mediated gene transfer to investigate the potential involvement of this family of regulatory proteins in the control of myogenesis. Transfection of the mouse skeletal muscle cell line C2 with the oncogenic forms of H-ras or N-ras completely suppressed both myoblast fusion and induction of the muscle-specific gene products nicotinic acetylcholine receptor and creatine kinase. Inhibition of differentiation by activated ras genes occurred at the level of muscle-specific mRNA accumulation. In contrast, proto-oncogenic forms of N-ras or H-ras had no apparent effects on the ability of C2 cells to differentiate. Myoblasts transfected with activated ras genes exhibited normal growth properties and ceased proliferating in the absence of mitogens, indicating that ras inhibited differentiation through a mechanism independent of cell proliferation. These results demonstrate that activated ras gene products mimic the inhibitory effects of fibroblast growth factor and type beta transforming growth factor on myogenic differentiation and suggest that each of these regulators of myogenesis may operate through a common intracellular pathway.
The c-rel proto-oncogene, which is expressed predominantly in hemopoietic cells encodes a subunit of the NF-kappa B-like family of transcription factors. In mice with an inactivated c-rel gene, whereas development of cells from all hemopoietic lineages appeared normal, humoral immunity was impaired and mature B and T cells were found to be unresponsive to most mitogenic stimuli. Phorbol ester and calcium ionophore costimulation, in contrast to certain membrane receptor-mediated signals, overcame the T cell-proliferative defect, demonstrating that T cell proliferation occurs by Rel-dependent and -independent mechanisms. The ability of exogenous interleukin-2 to restore T Cell, but not B cell, proliferation indicates that Rel regulates the expression of different genes in B and T cells that are crucial for cell division and immune function.
The eukaryotic transcription factor NF-kappa B plays a central role in the induced expression of human immunodeficiency virus type 1 and in many aspects of the genetic program mediating normal T-cell activation and growth. The nuclear activity of NF-kappa B is tightly regulated from the cytoplasmic compartment by an inhibitory subunit called I kappa B alpha. This cytoplasmic inhibitor is rapidly phosphorylated and degraded in response to a diverse set of NF-kappa B-inducing agents, including T-cell mitogens, proinflammatory cytokines, and viral transactivators such as the Tax protein of human T-cell leukemia virus type 1. To explore these I kappa B alpha-dependent mechanisms for NF-kappa B induction, we identified novel mutants of I kappa B alpha that uncouple its inhibitory and signal-transducing functions in human T lymphocytes. Specifically, removal of the N-terminal 36 amino acids of I kappa B alpha failed to disrupt its ability to form latent complexes with NF-kappa B in the cytoplasm. However, this deletion mutation prevented the induced phosphorylation, degradative loss, and functional release of I kappa B alpha from NF-kappa B in Tax-expressing cells. Alanine substitutions introduced at two serine residues positioned within this N-terminal regulatory region of I kappa B alpha also yielded constitutive repressors that escaped from Tax-induced turnover and that potently inhibited immune activation pathways for NF-kappa B induction, including those initiated from antigen and cytokine receptors. In contrast, introduction of a phosphoserine mimetic at these sites rectified this functional defect, a finding consistent with a causal linkage between the phosphorylation status and proteolytic stability of this cytoplasmic inhibitor. Together, these in vivo studies define a critical signal response domain in I kappa B alpha that coordinately controls the biologic activities of I kappa B alpha and NF-kappa B in response to viral and immune stimuli.
The terminal differentiation of C2C12 skeletal muscle cells involves the activation of unique sets of genes and an irreversible withdrawal from the cell cycle. This process is associated with a decrease in cdk2 activity in cell extracts. The decrease in cdk2 activity correlates with diminished levels of cdk2 and cyclin A and with a marked induction of the p21 cyclin-dependent kinase (cdk) inhibitor. The upregulation of p21 occurred at the levels of mRNA and protein, and p21 formed a complex with the cyclin kinases in myotubes. Further, the immunodepletion of p21 from myotube extracts neutralized the heat-stable cdk2 inhibitory activity that was induced upon myogenic differentiation. The levels of p21 mRNA, protein, and activity remained constant in myotubes when they were reexposed to mitogen-rich growth medium, indicating that permanent changes in the cell's genetic program contribute to its sustained expression following terminal differentiation. Indeed, 10T1/2 fibroblasts transformed with the myogenic factor MyoD, but not the parental multipotent cells, upregulated p21 transcript levels when induced to differentiate by serum withdrawal, demonstrating that the upregulation is an integral feature of myogenic commitment and differentiation. The functional consequences of this upregulation were indicated by ectopically expressing p21 in myoblasts; this was sufficient for cell cycle arrest in mitogen-rich growth medium. The induction and sustained expression of p21 appears to be a contributory mechanism by which myocytes irreversibly exit the cell cycle upon terminal differentiation.
Ectopic overexpression of v-H-Ras protein in NIH 3T3 cells resulted in cellular transformation and an acceleration of G1 progression
of these cells. A shortened G1 phase was found to be associated with an increased level of cyclin D1 but not cyclin E protein.
Using an antisense blocking method, reduced synthesis of cyclin D1 in v-H-Ras transformants resulted in a slower G1 progression
rate of these cells. Although constitutive overexpression of cyclin D1 in NIH 3T3 cells accelerated G1 progression, cells
remained untransformed. Furthermore, inhibition of cyclin D1 synthesis greatly impaired the soft-agar cloning efficiency of
v-H-Ras transformants. These results suggest that increased expression of cyclin D1 is necessary but not sufficient for the
transforming activity of v-H-Ras. Similar effect on cell cycle progression was also observed in Raf-transformed cells. In
addition to cyclin D1, cyclin E protein was found to be elevated in Src transformants. This may account for the further shortening
of the G1 phase of these cells. Activation of an additional Ras-independent pathway was suggested to be responsible for the
further acceleration of the G1 phase in Src transformants.
NF-kappa B, a heterodimeric transcription factor composed of p50 and p65 subunits, can be activated in many cell types and is thought to regulate a wide variety of genes involved in immune function and development. Mice lacking the p50 subunit of NF-kappa B show no developmental abnormalities, but exhibit multifocal defects in immune responses involving B lymphocytes and nonspecific responses to infection. B cells do not proliferate in response to bacterial lipopolysaccharide and are defective in basal and specific antibody production. Mice lacking p50 are unable effectively to clear L. monocytogenes and are more susceptible to infection with S. pneumoniae, but are more resistant to infection with murine encephalomyocarditis virus. These data support the role of NF-kappa B as a vital transcription factor for both specific and nonspecific immune responses, but do not indicate a developmental role for the factor.
A DNA-binding factor with properties of NF-kappa B and another similar activity are rapidly induced when growth-arrested BALB/c 3T3 cells are stimulated with serum growth factors. Induction of these DNA-binding activities is not inhibited by pretreatment of quiescent cells with the protein synthesis inhibitor cycloheximide. Interestingly, the major NF-kappa B-like activity is not detected in nuclear extracts of proliferating cells, and thus its expression appears to be limited to the G0-to-G1 transition in 3T3 cells. These DNA-binding activities bind many of the expected NF-kappa B target sequences, including elements in the class I major histocompatibility complex and human immunodeficiency virus enhancers, as well as a recently identified NF-kappa B binding site upstream of the c-myc gene. Furthermore, both the class I major histocompatibility complex and c-myc NF-kappa B binding sites confer inducibility on a minimal promoter in 3T3 cells stimulated with serum growth factors. The results demonstrate that NF-kappa B-like activities are immediate-early response proteins in 3T3 cells and suggest a role for these factors in the G0-to-G1 transition.
Conditional overexpression of human cyclins B1, D1, and E was accomplished by using a synthetic cDNA expression system based on the Escherichia coli tetracycline repressor. After induction of these cyclins in asynchronous Rat-1 fibroblasts, a decrease in the length of the G1 interval was observed for cyclins D1 and E, consistent with an acceleration of the G1/S phase transition. We observed, in addition, a compensatory lengthening of S phase and G2 so that the mean cell cycle length in populations constitutively expressing these cyclins was unchanged relative to those of their uninduced counterparts. We found that expression of cyclin B1 had no effect on cell cycle dynamics, despite elevated levels of cyclin B-associated histone H1 kinase activity. Induction of cyclins D1 and E also accelerated entry into S phase for synchronized cultures emerging from quiescence. However, whereas cyclin E exerted a greater effect than cyclin D1 in asynchronous cycling cells, cyclin D1 conferred a greater effect upon stimulation from quiescence, suggesting a specific role for cyclin D1 in the G0-to-G1 transition. Overexpression of cyclins did not prevent cells from entering into quiescence upon serum starvation, although a slight delay in attainment of quiescence was observed for cells expressing either cyclin D1 or cyclin E. These results suggest that cyclins D1 and E are rate-limiting activators of the G1-to-S phase transition and that cyclin D1 might play a specialized role in facilitating emergence from quiescence.
Differentiation of skeletal muscle involves withdrawal of myoblasts from the cell cycle, fusion to form myotubes, and the coordinate expression of a variety of muscle-specific gene products. Fibroblast growth factor and type beta transforming growth factor specifically inhibit myogenesis; however, the transmembrane signaling pathways responsible for suppression of differentiation by these growth factors remain elusive. Because ras proteins have been implicated in the transduction of growth factor signals across the plasma membrane, we used DNA-mediated gene transfer to investigate the potential involvement of this family of regulatory proteins in the control of myogenesis. Transfection of the mouse skeletal muscle cell line C2 with the oncogenic forms of H-ras or N-ras completely suppressed both myoblast fusion and induction of the muscle-specific gene products nicotinic acetylcholine receptor and creatine kinase. Inhibition of differentiation by activated ras genes occurred at the level of muscle-specific mRNA accumulation. In contrast, proto-oncogenic forms of N-ras or H-ras had no apparent effects on the ability of C2 cells to differentiate. Myoblasts transfected with activated ras genes exhibited normal growth properties and ceased proliferating in the absence of mitogens, indicating that ras inhibited differentiation through a mechanism independent of cell proliferation. These results demonstrate that activated ras gene products mimic the inhibitory effects of fibroblast growth factor and type beta transforming growth factor on myogenic differentiation and suggest that each of these regulators of myogenesis may operate through a common intracellular pathway.
Studies on mice deficient in nuclear factor kappa B (NF-κB) subunits have shown that this transcription factor is important for lymphocyte responses to antigens and cytokine-inducible gene expression. In particular, the RelA (p65) subunit is required for induction of tumor necrosis factor-α (TNF-α)-dependent genes. Treatment of RelA-deficient (RelA^(−/−)) mouse fibroblasts and macrophages with TNF-α resulted in a significant reduction in viability, whereas RelA^(+/+) cells were unaffected. Cytotoxicity to both cell types was mediated by TNF receptor 1. Reintroduction of RelA into RelA^(−/−) fibroblasts resulted in enhanced survival, demonstrating that the presence of RelA is required for protection from TNF-α. These results have implications for the treatment of inflammatory and proliferative diseases.
The NF-kappa B/Rel family of at least five transcription factor polypeptides is thought to function both as a developmental regulator in B cells and as a rapid response system in all cells. To examine this notion in more detail, we determined the protein contents of both the inducible and constitutive NF-kappa B/Rel activities in a pre-B-cell line, 70Z/3, and a mature B-cell line, WEHI 231. NF-kappa B p50/p65 is the major inducible nuclear complex after lipopolysaccharide or phorbol myristate acetate treatment of 70Z/3 cells. The constitutive and inducible complexes in WEHI 231 cells are mainly composed of p50 and Rel. The constitutive or induced activities are all sensitive to I kappa B-alpha, but this inhibitor is very short-lived in WEHI 231 cells, suggesting that the balance between synthesis and degradation of I kappa B-alpha determines whether a particular cell lineage has constitutive activity. A patterned expression of the NF-kappa B/Rel activator proteins emerges from an analysis of other B-lineage cell lines and splenic B cells: mainly p50 and p65 in pre-B (and non-B) cells, a predominance of Rel and p50 in mature B cells, and expression of p52 and RelB in plasmacytoma lines. This ordered pattern of regulators may reflect the requirement for expression of different genes during terminal B-cell differentiation because different combinations of NF-kappa B/Rel family members preferentially activate distinct kappa B sites in reporter constructs.
NF-κB, which consists of two polypeptides, p50 (M_r 50K) and p65/RelA (M_r 65K), is thought to be a key regulator of genes involved in responses to infection, inflammation and stress. Indeed, although developmentally normal, mice deficient in p50 display functional defects in immune responses. Here we describe the generation of mice deficient in the RelA subunit of NF-κB. Disruption of the relA locus leads to embryonic lethality at 15–16 days of gestation, concomitant with a massive degeneration of the liver by programmed cell death or apoptosis. Embryonic fibroblasts from RelA-deficient mice are defective in the tumour necrosis factor (TNF)-mediated induction of messenger RNAs for IκBα and granulocyte/macrophage colony stimulating factor (GM-CSF), although basal levels of these transcripts are unaltered. These results indicate that RelA controls inducible, but not basal, transcription in NF-κB-regulated pathways.
The ras proto-oncogene is frequently mutated in human tumors and functions to chronically stimulate signal transduction cascades resulting in the synthesis or activation of specific transcription factors, including Ets, c-Myc, c-Jun, and nuclear factor kappa B (NF-κB). These Ras-responsive transcription factors are required for transformation, but the mechanisms by which these proteins facilitate oncogenesis have not been fully established. Oncogenic Ras was shown to initiate a p53-independent apoptotic response that was suppressed through the activation of NF-κB. These results provide an explanation for the requirement of NF-κB for Ras-mediated oncogenesis and provide evidence that Ras-transformed cells are susceptible to apoptosis even if they do not express the p53 tumor-suppressor gene product.
Guidelines for submitting commentsPolicy: Comments that contribute to the discussion of the article will be posted within approximately three business days. We do not accept anonymous comments. Please include your email address; the address will not be displayed in the posted comment. Cell Press Editors will screen the comments to ensure that they are relevant and appropriate but comments will not be edited. The ultimate decision on publication of an online comment is at the Editors' discretion. Formatting: Please include a title for the comment and your affiliation. Note that symbols (e.g. Greek letters) may not transmit properly in this form due to potential software compatibility issues. Please spell out the words in place of the symbols (e.g. replace “α” with “alpha”). Comments should be no more than 8,000 characters (including spaces ) in length. References may be included when necessary but should be kept to a minimum. Be careful if copying and pasting from a Word document. Smart quotes can cause problems in the form. If you experience difficulties, please convert to a plain text file and then copy and paste into the form.
Almost exactly ten years following the first publication on NF-κB (Sen and Baltimore 1986), researchers working on transcriptional regulation by NF-κB/Rel and IκB proteins gathered for the third time to discuss recent developments in the field (Madrid, July 8-10, 1996). The first meeting of its kind was a Howard Hughes workshop at the NIH in November 1992 and the second one a Banbury Conference held at Cold Spring Harbor in October 1993. This year's meeting was organized by R. Bravo (Bristol-Myers Squibb, Princeton) and P. S. Lazo (Universidad de Oviedo) and held at the Juan March Foundation in Madrid, Spain.
Activation of the transcription factor NF-κB by tumor necrosis factor (TNF) and interleukin-1 (IL-1) requires the NF-κB-inducing kinase (NIK). In a yeast two-hybrid screen for NIK-interacting proteins, we have identified a protein kinase previously known as CHUK. Overexpression of CHUK activates a NF-κB-dependent reporter gene. A catalytically inactive mutant of CHUK is a dominant-negative inhibitor of TNF-, IL-1-, TRAF-, and NIK-induced NF-κB activation. CHUK associates with the NF-κB inhibitory protein, IκB-α, in mammalian cells. CHUK specifically phosphorylates IκB-α on both serine 32 and serine 36, modifications that are required for targeted degradation of IκB-α via the ubiquitin-proteasome pathway. This phosphorylation of IκB-α is greatly enhanced by NIK costimulation. Thus, CHUK is a NIK-activated IκB-α kinase that links TNF- and IL-1-induced kinase cascades to NF-κB activation.
Activation of muscle gene transcription in differentiating skeletal myoblasts requires their withdrawal from the cell cycle.
The effects of ectopic cyclin expression on activation of muscle gene transcription by myogenic basic helix-loop-helix (bHLH)
regulators were investigated. Ectopic expression of cyclin D1, but not cyclins A, B1, B2, C, D3, and E, inhibited transcriptional
activation of muscle gene reporter constructs by myogenic bHLH regulators in a dose-dependent manner. Ectopic expression of
cyclin D1 inhibited the activity of a myogenic bHLH regulator mutant lacking the basic region protein kinase C site, indicating
that phosphorylation of this site is not relevant to the mechanism of inhibition. Analysis of cyclin D1 mutants revealed that
the C-terminal acidic region was required for inhibition of myogenic bHLH regulator activity, whereas an intact N-terminal
pRb binding motif was not essential. Together, these results implicate expression of cyclin D1 as a central determinant of
a putatively novel mechanism that links positive control of cell cycle progression to negative regulation of genes expressed
in differentiated myocytes.
Recently we purified a 900 kDa cytokine-responsive IκB kinase complex (IKK) and molecularly cloned one of its subunits, IKKα, a serine kinase. We now describe the molecular cloning and characterization of IKKβ, a second subunit of the IKK complex. IKKβ is 50% identical to IKKα and like it contains a kinase domain, a leucine zipper, and a helix-loop-helix. Although IKKα and IKKβ can undergo homotypic interaction, they also interact with each other and the functional IKK complex contains both subunits. The catalytic activities of both IKKα and IKKβ make essential contributions to IκB phosphorylation and NF-κB activation. While the interactions between IKKα and IKKβ may be mediated through their leucine zipper motifs, their helix-loop-helix motifs may be involved in interactions with essential regulatory subunits.
In Drosophila, the Dorsal protein establishes the embryonic dorso- ventral axis during development. Here we show that the vertebrate homo1ogue of Dorsal, nuclear factor-kappa B (NF-κB), is vital for the formation of the proximo-distal organizer of the developing limb bud, the apical ectodermal ridge (AER). Transcription of the HF-κB proto-oncogene c-rel is regulated, in part, during morphogenesis of the limb bud by AER-derived signals such as fibroblast growth factors. Interruption of NF-κB activity using viral- mediated delivery of an inhibitor results in a highly dysmorphic AER, reduction in overall limb size, loss of distal elements and reversal in the direction of limb outgrowth. Furthermore, inhibition of NF-κB activity in limb mesenchyme leads to a reduction in expression of Sonic hedgehog and T~ but derepresses expression of the bone morphogenetic protein-4 gene. These results are the first evidence that vertebrate NF-κB proteins act to transmit growth factor signals between the ectoderm and the underlying mesenchyme during embryonic limb formation.
Bcr-Abl is a chimeric oncoprotein that is strongly implicated in acute lymphoblastic (ALL) and chronic myelogenous leukemias (CML). This deregulated tyrosine kinase selectively causes hematopoietic disorders resembling human leukemias in animal models and transforms fibroblasts and hematopoietic cells in culture. Bcr-Abl also protects cells from death induced on cytokine deprivation or exposure to DNA damaging agents. In addition, the antiapoptotic function of Bcr-Abl is thought to play a necessary role in hematopoietic transformation and potentially in leukemogenesis. The transcription factor NF-kappaB has been identified recently as an inhibitor of apoptosis and as a potential regulator of cellular transformation. This study shows that expression of Bcr-Abl leads to activation of NF-kappaB-dependent transcription by causing nuclear translocation of NF-kappaB as well as by increasing the transactivation function of the RelA/p65 subunit of NF-kappaB. Importantly, this activation is dependent on the tyrosine kinase activity of Bcr-Abl and partially requires Ras. The ability of Bcr-Abl to protect cytokine-dependent 32D myeloid cells from death induced by cytokine deprivation or DNA damage does not, however, require functional NF-kappaB. However, using a super-repressor form of IkappaBalpha, we show that NF-kappaB is required for Bcr-Abl-mediated tumorigenicity in nude mice and for transformation of primary bone marrow cells. This study implicates NF-kappaB as an important component of Bcr-Abl signaling. NF-kappaB-regulated genes, therefore, likely play a role in transformation by Bcr-Abl and thus in Bcr-Abl-associated human leukemias.
In muscle cells, as in a variety of cell types, proliferation and differentiation are mutually exclusive events controlled by a balance of opposing cellular signals. Members of the MyoD family of muscle-specific helix-loop-helix proteins which, in collaboration with ubiquitous factors, activate muscle differentiation and inhibit cell proliferation function at the nexus of the cellular circuits that control proliferation and differentiation of muscle cells. The activities of these myogenic regulators are negatively regulated by peptide growth factors and activated oncogenes whose products transmit growth signals from the membrane to the nucleus. Recent studies have revealed multiple mechanisms through which intracellular growth factor signals may interfere with the functions of the myogenic regulators. When expressed at high levels, members of the MyoD family can override mitogenic signals and can cause growth arrest independent of their effects on differentiation. The ability of these myogenic regulators to inhibit proliferation of normal as well as transformed cells from multiple lineages suggests that they interact with conserved components of the cellular machinery involved in cell cycle progression and that similar types of regulatory factors participate in differentiation and cell cycle control in diverse cell types.
NF-kappa B is an inducible transcription factor comprised of a 50-kD (p50) and a 65-kD (p65) subunit. Induction of NF-kappa B activity, which is a critical event in many signal transduction pathways, involves release from a cytoplasmic inhibitory protein, I kappa B, followed by translocation of the active transcription factor complex into the nucleus. Earlier studies suggested that I kappa B targets the p65 subunit of NF-kappa B. However, we demonstrate by in vitro and in vivo methods that the recently cloned I kappa B/MAD-3 interacts with both the p50 and p65 subunits of NF-kappa B, as well as c-Rel. Furthermore, an alternatively spliced, dimerization-deficient transforming variant of p65 (p65 delta) interacts extremely weakly with I kappa B/MAD-3, suggesting that dimerization is important for interaction. We demonstrate that the conserved nuclear localization sequences (NLSs) of NF-kappa B and c-Rel are the targets for I kappa B/MAD-3 interaction. Indirect immunofluorescence experiments demonstrate that I kappa B/MAD-3 expression retains both p65 and p50 in the cytoplasm. Furthermore, and most important, a p65 that contains an SV40 large T antigen NLS in addition to its own NLS is no longer retained in the cytoplasm in the presence of I kappa B/MAD-3. We propose that I kappa B/MAD-3 masks the NLSs of NF-kappa B and c-Rel and that this constitutes the mechanism for cytoplasmic retention of these proteins.
We report the molecular characterization of two novel rat helix-loop-helix (HLH) proteins, designated HES-1 and HES-3, that show structural homology to the Drosophila hairy and Enhancer of split [E(spl)] proteins, both of which are required for normal neurogenesis. HES-1 mRNA, expressed in various tissues of both embryos and adults, is present at a high level in the epithelial cells, including the embryonal neuroepithelial cells, as well as in the mesoderm-derived tissues such as the embryonal muscle. In contrast, HES-3 mRNA is produced exclusively in cerebellar Purkinje cells. HES-1 represses transcription by binding to the N box, which is a recognition sequence of E(spl) proteins. Interestingly, neither HES-1 nor HES-3 alone interacts efficiently with the E box, but each protein decreases the transcription induced by E-box-binding HLH activators such as E47. Furthermore, HES-1 also inhibits the functions of MyoD and MASH1 and effectively diminishes the myogenic conversion of C3H10T1/2 cells induced by MyoD. These results suggest that HES-1 may play an important role in mammalian development by negatively acting on the two different sequences while HES-3 acts as a repressor in a specific type of neurons.
Three mouse cyclin-like (CYL) genes were isolated, two of which are regulated by colony-stimulating factor 1 (CSF-1) during the G1 phase of the macrophage cell cycle. CSF-1 deprivation during G1 leads to rapid degradation of CYL proteins (p36CYL) and correlates with failure to initiate DNA synthesis. However, after entering S phase, macrophages no longer require CSF-1 and can complete cell division without expressing CYL genes. During G1, p36CYL is phosphorylated and associates with a polypeptide antigenically related to p34cdc2. The timing of p36CYL expression, its rapid turnover in the absence of CSF-1, and its phosphorylation and transient binding to a cdc2-related polypeptide suggest that CYL genes may function during S phase commitment.
A cDNA library prepared from a human glioblastoma cell line has been introduced into a budding yeast strain that lacks CLN1 and CLN2 and is conditionally deficient for CLN3 function. We rescued a gene that we call cyclin D1. It is related to A-, B-, and CLN-type cyclins, but appears to define a new subclass within the cyclin gene family. Transcription of the cyclin D1 gene gives rise to two major transcripts through alternative polyadenylation. The cyclin D1 gene transcript and its 34 kd product are both abundant in the glioblastoma cell line of origin.
A DNA-binding factor with properties of NF-kappa B and another similar activity are rapidly induced when growth-arrested BALB/c
3T3 cells are stimulated with serum growth factors. Induction of these DNA-binding activities is not inhibited by pretreatment
of quiescent cells with the protein synthesis inhibitor cycloheximide. Interestingly, the major NF-kappa B-like activity is
not detected in nuclear extracts of proliferating cells, and thus its expression appears to be limited to the G0-to-G1 transition
in 3T3 cells. These DNA-binding activities bind many of the expected NF-kappa B target sequences, including elements in the
class I major histocompatibility complex and human immunodeficiency virus enhancers, as well as a recently identified NF-kappa
B binding site upstream of the c-myc gene. Furthermore, both the class I major histocompatibility complex and c-myc NF-kappa
B binding sites confer inducibility on a minimal promoter in 3T3 cells stimulated with serum growth factors. The results demonstrate
that NF-kappa B-like activities are immediate-early response proteins in 3T3 cells and suggest a role for these factors in
the G0-to-G1 transition.
We have isolated a cDNA clone encoding a novel helix-loop-helix (HLH) protein, Id. Id is missing the basic region adjacent to the HLH domain that is essential for specific DNA binding in another HLH protein, MyoD. An in vitro translation product of Id can associate specifically with at least three HLH proteins (MyoD, E12, and E47) and attenuate their ability to bind DNA as homodimeric or heterodimeric complexes. Id is expressed at varying levels in all cell lines tested. In three cell lines that can be induced to undergo terminal differentiation, Id RNA levels decrease upon induction. Transfection experiments indicate that over-expression of Id inhibits the trans-activation of the muscle creatine kinase enhancer by MyoD. Based on these findings, we propose that HLH proteins lacking a basic region may negatively regulate other HLH proteins through the formation of nonfunctional heterodimeric complexes.
Transformation of myoblasts by activated ras inhibits myogenic differentiation. We demonstrate that this oncogene inhibits expression of the muscle regulatory factors MyoD1 and myogenin. Expression of retroviral-encoded MyoD1 in ras-transformed myoblasts leads to the re-expression of both terminal differentiation markers and lineage markers expressed in proliferating myoblasts (including endogenous MyoD1 and myogenin), suggesting that ras inhibits myogenic differentiation in a manner dependent on the loss of MyoD1 expression. In addition, we show that fos transformation of myoblasts inhibits muscle differentiation by a similar mechanism.
Skeletal muscle development is regulated by a complex series of genetic and environmental cues that control the establishment of the myogenic lineage and the differentiation of determined myoblasts. Numerous agents, including growth factors and oncogene products, have been shown to inhibit skeletal muscle development, possibly by affecting the pattern of signal transduction that is required for myogenesis. Among the eukaryotic G proteins that have been implicated as mediators of signal transduction are the protein products of the mammalian ras genes (p21s). In this study, we demonstrate that expression of a transfected, oncogenic, human H-ras gene in C3H10T1/2-derived myoblasts has dramatic, yet varied, effects on skeletal myogenesis. While some H-ras transformed myoblast clones are differentiation-defective, other clones are inhibited from morphologically differentiating but retain a limited ability to biochemically differentiate. The H-ras induced inhibition of differentiation usually is associated with a decreased expression of the myogenic determination gene, MyoD1. Introduction of a MyoD1 cDNA expression vector into differentiation-defective H-ras expressing myoblasts partially restores the myogenic potential in these cells. Our results suggest that activated H-ras p21 inhibits the terminal differentiation of myoblasts by producing a general reduction in the differentiation competence of cells which, in the most extreme case, is a consequence of the down-regulation of the MyoD1 determination gene.
Several observations have suggested that NF-kappa B transcription factors could be involved in carcinogenesis. To investigate the possibility that members of the NF-kappa B family participate in the molecular control of the transformed phenotype, we examined the expression of these proteins in human breast cancer cell lines as well as in primary tumors. Western Immunoblots demonstrated high expression of the p52 precursor p100 (NFKB2) in several breast cancer cell lines while human mammary epithelial cells express this protein only faintly. Eighteen primary breast tumors out of 24 displayed significant expression of the p100/p52 protein. In MDA-MB-435 cells, overexpressed p100 and p52 are predominantly cytoplasmic and coimmunoprecipitation experiments demonstrated that p100 sequesters the heterodimer p50/p65 in the cytoplasm. We demonstrate that most p65 protein is complexed with p100 in these cells while it is complexed predominantly with I kappa B-alpha in cell lines expressing less p100. Our data strengthen the hypothesis that NF-kappa B could be involved in carcinogenesis and suggest that the p100/p52 NF-kappa B subunit could play a role in the development of human breast cancers, possibly by sequestering other NF-kappa B-related proteins in the cytoplasm.
Guidelines for submitting commentsPolicy: Comments that contribute to the discussion of the article will be posted within approximately three business days. We do not accept anonymous comments. Please include your email address; the address will not be displayed in the posted comment. Cell Press Editors will screen the comments to ensure that they are relevant and appropriate but comments will not be edited. The ultimate decision on publication of an online comment is at the Editors' discretion. Formatting: Please include a title for the comment and your affiliation. Note that symbols (e.g. Greek letters) may not transmit properly in this form due to potential software compatibility issues. Please spell out the words in place of the symbols (e.g. replace “α” with “alpha”). Comments should be no more than 8,000 characters (including spaces ) in length. References may be included when necessary but should be kept to a minimum. Be careful if copying and pasting from a Word document. Smart quotes can cause problems in the form. If you experience difficulties, please convert to a plain text file and then copy and paste into the form.
Using homologous recombination, mice lacking cyclin D1 were generated by replacing most of the first exon of the Cyl-1 gene with sequences encoding neomycin resistance. Cyl-1(-1-) mice were viable and fertile but consistently smaller than their heterozygous or wild-type littermates. The nullizygous animals also showed two distinctive abnormalities: a severe retinopathy caused by impaired development of all layers of the retina and, in the mammary gland during pregnancy, a marked reduction in acinar development accompanied by a failure to lactate. Approximately 50% of animals also had a malformation of the jaw that manifested itself as a misalignment of the incisor teeth. Mouse embryo fibroblasts isolated from 14 day nullizygous, heterozygous, or wild-type embryos and grown under standard conditions showed similar cell-cycle and growth characteristics. Thus although cyclin D1 kinase activity may facilitate G1 progression, it is not essential for the development of most tissues and organs, and only a few specialized cell lineages are demonstrably sensitive to its absence.
NF-kappa B, which consists of two polypeptides, p50 (M(r) 50K) and p65/RelA (M(r) 65K), is thought to be a key regulator of genes involved in responses to infection, inflammation and stress. Indeed, although developmentally normal, mice deficient in p50 display functional defects in immune responses. Here we describe the generation of mice deficient in the RelA subunit of NF-kappa B. Disruption of the relA locus leads to embryonic lethality at 15-16 days of gestation, concomitant with a massive degeneration of the liver by programmed cell death or apoptosis. Embryonic fibroblasts from RelA-deficient mice are defective in the tumour necrosis factor (TNF)-mediated induction of messenger RNAs for I kappa B alpha and granulocyte/macrophage colony stimulating factor (GM-CSF), although basal levels of these transcripts are unaltered. These results indicate that RelA controls inducible, but not basal, transcription in NF-kappa B-regulated pathways.
D-type cyclins couple extracellular signals to the biochemical machinery that governs progression through G1 phase of the mammalian cell division cycle. Induced by growth factor stimulation, D-type cyclins assemble with cyclin-dependent kinases CDK4 and CDK6 to form holoenzymes that facilitate exit from G1 by phosphorylating key substrates, including the retinoblastoma protein. Activation of the holoenzymes is antagonized by polypeptide inhibitors of CDK activity, which are induced by antiproliferative signals. Once cells pass a late G1 restriction point, cyclin-D-dependent kinases are unnecessary for completion of the cell cycle, implying that their primary role is to sense the cell's readiness to replicate DNA and to enforce the commitment to enter S phase.
Mice lacking cyclin D1 have been generated by gene targeting in embryonic stem cells. Cyclin D1-deficient animals develop to term but show reduced body size, reduced viability, and symptoms of neurological impairment. Their retinas display a striking reduction in cell number due to proliferative failure during embryonic development. In situ hybridization studies of normal mouse embryos revealed an extremely high level of cyclin D1 in the retina, suggesting a special dependence of this tissue on cyclin D1. In adult mutant females, the breast epithelial compartment fails to undergo the massive proliferative changes associated with pregnancy despite normal levels of ovarian steroid hormones. Thus, steroid-induced proliferation of mammary epithelium during pregnancy may be driven through cyclin D1.
PRAD1 (previously D11S287) is a putative proto-oncogene at 11q13, activated by overexpression through gene rearrangement or gene amplification in several types of human tumors including parathyroid adenomas, centrocytic lymphomas and other B-cell tumors with t(11;14), and breast cancers. PRAD1 (also CCND1) encodes cyclin D1, which may regulate the G1-S phase transition in the cell cycle. Here, we report the cloning and characterization of the chromosomal PRAD1/cyclin D1 gene and the sequence of its promoter region. The gene spans about 15 kb and has 5 exons; its promoter region has Sp1 binding sites and no obvious TATA box, characteristics of housekeeping genes and growth-regulating genes. Furthermore, an E2F binding motif present close to the major transcription start site may be involved in cell cycle-dependent expression of this gene. We also report the sequence of DNAs spanning joining regions of a reciprocal parathyroid hormone/PRAD1 gene rearrangement in a parathyroid adenoma. Comparison with normal sequences suggests that the rearrangement was not a simple break-and-ligate event, but rather involved multiple steps, including two microdeletions and a microinversion. Very short sequences conserved near the breakpoints and symmetrical elements in the eventually inverted DNA segment might have played a role in this illegitimate complex recombination, which may have similarities with a constitutional translocation in Duchenne muscular dystrophy.
The transcription factor NF-kappa B, shown to be essential for expression of the immunoglobulin C kappa gene, is a key regulatory component in pre-B to B-cell differentiation. While previous studies have used lymphoid cell line models, here we examine the expression and subunit composition of rel/NF-kappa B complexes in normal murine pre-B and B lymphocytes. Two major NF-kappa B complexes are detected in pre-B and B cells. A high mobility complex, found in pre-B (Cb) and B cells (C beta) is a homodimer of the NF-kappa B subunit p50. In pre-B cells, the slower migrating complex (Ca), which is predominantly cytoplasmic, is largely comprised of p50 and p65, whereas in B cells, a nuclear and cytoplasmic complex (C alpha) of identical mobility to Ca mainly consists of p50 and p75c-rel. While p50 and p65 levels do not change during pre-B to B-cell differentiation, p75c-rel is 5- to 6-fold more abundant in B cells compared to pre-B cells, a finding consistent with the switch in NF-kappa B subunit usage. During lipopolysaccharide-induced B-cell proliferation, transient up-regulation of both the nuclear p50 homodimer and p75c-rel containing complex is mirrored by a concurrent increase in c-rel and p105 but not p65 mRNA expression, a finding consistent with rel-NF-kappa B expression in B cells being controlled by an autoregulatory mechanism.
Liver regeneration after two-thirds partial hepatectomy (PH) is a process in which quiescent, fully differentiated hepatocytes rapidly reenter the cell cycle and eventually divide until the original liver mass is restored. Although the exact nature of the growth-initiating signals is unknown, enhanced expression of growth-related genes has been detected during the first hour after operation. This suggests that activation of transcriptional and posttranscriptional regulatory factors is likely to be a very early event in liver regeneration. Here we report the rapid, transient induction of DNA binding by nuclear factor (NF)-kappa B (p50/p65 heterodimer) and p50 homodimers within 30 min after PH. We also detected binding of post-hepatectomy factor. NF-kappa B binding peaks at 1 h after PH before declining and is not induced by sham operation. Liver cell separation studies indicated that the binding activation occurs in hepatocytes, a conclusion further supported by cell culture studies using the hepatocyte cell line AML-12. Furthermore, studies with the liver epithelial cell line LE-6 indicated that these DNA-binding activities are mitogen inducible. One-third hepatectomy, a procedure which primes hepatocytes to respond to growth factors, also induced NF-kappa B binding. We also found that tumor necrosis factor alpha, which may be involved in the control of liver regeneration, rapidly induced NF-kappa B DNA-binding activities in intact animals, similar to those induced by PH. These results suggest that NF-kappa B binding may play a role in making hepatocytes competent to proliferate.