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H-Ras is degraded by Wnt/ -catenin signaling via -TrCP-mediated polyubiquitylation
... RAS elevation also occurs in HCCs; this elevation is associated with poor prognosis in patients [12][13][14][15] . Stabilization of RAS proteins constitutively activates downstream signaling pathways associated with tumorigenesis [6][7][8][16][17][18][19][20] . Particularly, in CRC, RAS stabilization via the Wnt/β-catenin pathway, especially by the APC mutations that are found in~90% of human CRCs, plays important roles in the tumorigenesis [6][7][8] . ...
... Particularly, in CRC, RAS stabilization via the Wnt/β-catenin pathway, especially by the APC mutations that are found in~90% of human CRCs, plays important roles in the tumorigenesis [6][7][8] . In the resting state, RAS proteins are maintained at low levels due to proteasomal degradation by GSK3β-mediated phosphorylation and subsequent recruitment of the β-TrCP E3 linker protein 7,17 . In the case of aberrant Wnt/β-catenin signaling activation (e.g., caused by APC loss), RAS proteins and β-catenin are stabilized by inactivation of GSK3β, which results in enhancement of the colorectal tumorigenesis 7,21 . ...
... Although, activities of the RAS proteins have been mainly known to be regulated by the GTP and GDP loading switch and membrane localization, emerging evidences are suggesting that stability regulation of RAS also plays important roles in pathophysiology [6][7][8][16][17][18]21,43,44 . Recently, it has been reported that Nedd4-1 targets RAS proteins for their degradation and suppresses tumorigenesis, but activation of RAS evades the Nedd4-1-mediated degradation, leading to potential initiation of tumorigenesis 20 . ...
Stability regulation of RAS that can affect its activity, in addition to the oncogenic mutations, occurs in human cancer. However, the mechanisms for stability regulation of RAS involved in their activity and its roles in tumorigenesis are poorly explored. Here, we identify WD40-repeat protein 76 (WDR76) as one of the HRAS binding proteins using proteomic analyses of hepatocellular carcinomas (HCC) tissue. WDR76 plays a role as an E3 linker protein and mediates the polyubiquitination-dependent degradation of RAS. WDR76-mediated RAS destabilization results in the inhibition of proliferation, transformation, and invasion of liver cancer cells. WDR76−/− mice are more susceptible to diethylnitrosamine-induced liver carcinogenesis. Liver-specific WDR76 induction destabilizes Ras and markedly reduces tumorigenesis in HRasG12V mouse livers. The clinical relevance of RAS regulation by WDR76 is indicated by the inverse correlation of their expressions in HCC tissues. Our study demonstrates that WDR76 functions as a tumor suppressor via RAS degradation.
... The RAS/ERK pathway is regulated by Wnt/b-catenin signaling, and stability regulation of RAS is one of the key events for signaling cross-talk [9][10][11]. RAS proteins are subjected to glycogen synthase kinase 3 beta (GSK3b)-mediated polyubiquitination-dependent proteasomal degradation; GSK3b, the key component of the bcatenin destruction complex, phosphorylates RAS at the threonine (Thr) 144 and Thr-148 residues that are conserved among the RAS isoforms, which leads to the recruitment of the b-TrCP E3 linker [12,13]. Costabilization of b-catenin and RAS, particularly the KRAS mutant form, by APC mutations synergistically enhances the growth of CRC, and elevated levels of b-catenin and RAS are observed in CRC patient tissues, which indicates their pathological significance [12,14]. ...
... In a resting status, RAS proteins are subjected to GSK3bmediated phosphorylation at Thr-144 and Thr-148 residues, followed by the recruitment of the b-TrCP E3 linker, which leads to polyubiquitin-dependent proteasomal degradation [12,13]. Our previous observation that GSK3b-mediated RAS degradation is absent in CRC cells that express nondegradable MT b-catenin led us to identify a novel mechanism for the b-catenin-dependent sequential degradations of b-catenin and RAS via Wnt/b-catenin signaling. ...
... Horseradish peroxidase-conjugated anti-mouse (Cell Signaling Technology, #7076; 1:4,000) or anti-rabbit (Bio-Rad; 1:4,000) secondary antibodies were used. Immunoprecipitation and in vivo ubiquitination assays were performed as previously described [13]. Briefly, cells were washed in ice-cold PBS and lysed with RIPA buffer. ...
RAS proteins play critical roles in various cellular processes, including growth and transformation. RAS proteins are subjected to protein stability regulation via the Wnt/β-catenin pathway, and glycogen synthase kinase 3 beta (GSK3β) is a key player for the phosphorylation-dependent RAS degradation through proteasomes. GSK3β-mediated RAS degradation does not occur in cells that express a nondegradable mutant (MT) β-catenin. Here, we show that β-catenin directly interacts with RAS at the α-interface region that contains the GSK3β phosphorylation sites, threonine 144 and threonine 148 residues. Exposure of these sites by prior β-catenin degradation is required for RAS degradation. The introduction of a peptide that blocks the β-catenin-RAS interaction by binding to β-catenin rescues the GSK3β-mediated RAS degradation in colorectal cancer (CRC) cells that express MT β-catenin. The coregulation of β-catenin and RAS stabilities by the modulation of their interaction provides a mechanism for Wnt/β-catenin and RAS-ERK pathway cross-talk and the synergistic transformation of CRC by both APC and KRAS mutations.
... Recently, we found that Ras protein is stabilized via the activated Wnt/β-catenin signaling and stabilization of oncogenic Ras by aberrant mutational activation of Wnt/β-catenin pathway genes related with colorectal tumorigenesis [16][17][18][19][20][21]. However, other signaling pathways regulating Ras stability and its involvement with human cancer have not been characterized. ...
... The half-life of pan-Ras, as determined by treatment with the protein synthesis inhibitor cycloheximide (CHX), was approximately 9 hours and was mostly blocked by E 2 treatment ( Figure 1C and 1D). The Ras proteins were subjected to the degradation by polyubiquitylation-dependent proteasomes [19,20], but E 2 reduced the level of Ras polyubiquitylation ( Figure 1E). ...
... Recently, the stabilization of Ras protein has been shown as an alternative mechanism for Ras activity control. Ras protein stability increased by the Wnt/βcatenin signaling has been shown to be directly involved in colorectal tumorigenesis and cancer stem cell activation [19][20][21]. In addition, up-regulation of Ras has been observed in several cancers including colorectal and breast [31,32], and its increased expression has been associated with neoplastic transformation. ...
Estrogens are considered as a major risk factor of endometrial cancer. In this study, we identified a mechanism of tumorigenesis in which K-Ras protein is stabilized via estrogen signaling through the ER-α36 receptor. PKCδ was shown to stabilize K-Ras specifically via estrogen signaling. Estrogens stabilize K-Ras via inhibition of polyubiquitylation-dependent proteasomal degradation. Estrogen-induced cellular transformation was abolished by either K-Ras or PKCδ knockdown. The role of PKCδ in estrogen-induced tumorigenesis was confirmed in a mouse xenograft model by reduction of tumors after treatment with rottlerin, a PKCδ inhibitor. Finally, levels of PKCδ correlated with that of Ras in human endometrial tumor tissues. Stabilization of K-Ras by estrogen signaling involving PKCδ up-regulation provides a potential therapeutic approach for treatment of endometrial cancer.
... Increasingly, it is becoming recognized that ubiquitin-mediated protein modifications of RAS family members (H-, N-, and KRAS) play critical roles in protein abundance, maintenance of their activity, and association with downstream signaling molecules. Particularly, in the case of mutant KRAS, mono-/bi-ubiquitination enhances its GTP binding and its association with downstream signaling molecules [26,27], whereas polyubiquitination mediated through an F-box family E3, β-transducing repeat containing protein 1 (β-TrCP1), induces RAS degradation [28,29]. ...
... (This is in contrast to other RAS family members such as H-or N-RAS, which undergo proteasomal degradation.) [28,29] Furthermore, in HEK293 cells overexpressing KRAS-G12V, the protein half-life decreased from >12 hours to ∼3.5 hours following SMURF2 (CA) overexpression [ Figure 2, B (right panel) and C], suggesting the importance of SMURF2 ligase activity on mutant KRAS protein stability. To determine the half-life of the endogenous KRAS protein on Smurf2 knockdown, we transfected lung cancer cells carrying either wild type (H2347) or KRAS-G12V (H441) with either control or Smurf2 siRNA. ...
... Recently, β-TrCP, an F-box ubiquitin ligase, has been implicated in RAS (including KRAS) ubiquitination and degradation [28,29]. As reported earlier, we also observed that siRNA-mediated downregulation of β-TrCP1 in mutant KRAS-driven H358 and H441 cells enhances KRAS steady-state level ( Figure W3A); overexpression of β-TrCP1 causes degradation of KRAS ( Figure W3B), and immunoprecipitation studies show interaction between KRAS and β-TrCP1 only when KRAS degradation was inhibited by a lysosomal inhibitor, 3-MA ( Figure W3C ). ...
Attempts to target mutant KRAS have been unsuccessful. Here, we report the identification of Smad ubiquitination regulatory factor 2 (SMURF2) and UBCH5 as a critical E3:E2 complex maintaining KRAS protein stability. Loss of SMURF2 either by small interfering RNA/short hairpin RNA (siRNA/shRNA) or by overexpression of a catalytically inactive mutant causes KRAS degradation, whereas overexpression of wild-type SMURF2 enhances KRAS stability. Importantly, mutant KRAS is more susceptible to SMURF2 loss where protein half-life decreases from >12 hours in control siRNA-treated cells to <3 hours on Smurf2 silencing, whereas only marginal differences were noted for wild-type protein. This loss of mutant KRAS could be rescued by overexpressing a siRNA-resistant wild-type SMURF2. Our data further show that SMURF2 monoubiquitinates UBCH5 at lysine 144 to form an active complex required for efficient degradation of a RAS-family E3, β-transducing repeat containing protein 1 (β-TrCP1). Conversely, β-TrCP1 is accumulated on SMURF2 loss, leading to increased KRAS degradation. Therefore, as expected, β-TrCP1 knockdown following Smurf2 siRNA treatment rescues mutant KRAS loss. Further, we identify two conserved proline (P) residues in UBCH5 critical for SMURF2 interaction; mutation of either of these P to alanine also destabilizes KRAS. As a proof of principle, we demonstrate that Smurf2 silencing reduces the clonogenic survival in vitro and prolongs tumor latency in vivo in cancer cells including mutant KRAS-driven tumors. Taken together, we show that SMURF2:UBCH5 complex is critical in maintaining KRAS protein stability and propose that targeting such complex may be a unique strategy to degrade mutant KRAS to kill cancer cells.
... The roles of these additional regulatory mechanisms in cancer are poorly understood. The E3 ligase substrate recognition protein b-TrCP contributes to H-Ras degradation (8). H-Ras is also subject to monoand di-ubiquitylation, and these posttranslational modifications affect the cellular trafficking of H-Ras (6); how this influences H-Ras activity and the physiological relevance of this modification is not clear. ...
... These studies indicated a potential mechanism for tumor promotion by mutational synergistic activation of the Wnt/b-catenin and Ras pathways. The ERK pathway inhibition through negative Wnt/b-catenin signaling occurred by proteasomal degradation of Ras mediated by b-TrCP-E3 ligase (8). However, the detailed mechanism, including molecular events triggering the recruitment of b-TrCP-E3 ligase to Ras, was not determined. ...
... In a previous study, H-Ras was recognized by b-TrCP (8). Because b-TrCP recognizes b-catenin phosphorylated by GSK3b (11,12), we investigated whether GSK3b phosphorylated H-Ras and contributed to its ubiquitylation. ...
Although the guanosine triphosphate/guanosine diphosphate loading switch is a major regulatory mechanism that controls the activity of the guanosine triphosphatase Ras, we report a distinct mechanism for regulating Ras activity through phosphorylation-mediated degradation and describe the role of this second regulatory mechanism in the suppression of cellular transformation and tumors induced by Ras mutations. We found that negative regulators of Wnt/β-catenin signaling contributed to the polyubiquitin-dependent degradation of Ras after its phosphorylation by glycogen synthase kinase 3β (GSK3β) and the subsequent recruitment of β-TrCP-E3 ligase. We found a positive association between tumorigenesis and Ras stabilization resulting from the aberrant activation of Wnt/β-catenin signaling in adenomas from two mouse models of colon cancer, human colonic tumors from various stages, and colon polyps of patients with familial adenomatous polyposis. Our results indicated that GSK3β plays an essential role in Ras degradation and that inhibition of this degradation pathway by aberrant Wnt/β-catenin signaling may contribute to Ras-induced transformation in colorectal tumorigenesis.
... Therefore, β-catenin and Ras, which are aberrantly stabilized in CRC, could serve as good targets for the development of anti-CRC drugs. Based on our studies, which identified the mechanism of Ras degradation via inhibition of the Wnt/β-catenin pathway [7,16,17], we recently identified and characterized small molecules destabilizing both β-catenin and Ras by screening a library of chemicals that inhibit the Wnt/β-catenin pathway [18]. KY1220 and its functionally improved analog KYA1797K specifically bind to the RGS domain of Axin, activate GSK3β via a conformational change enhancing β-catenin complex assembly, and subsequently degrade both β-catenin and Ras via proteasomal degradation [18]. ...
... Immunoprecipitation and ubiquitination assays were performed as previously described [17]. Briefly, cells were washed in ice-cold PBS (Gibco) and lysed with RIPA buffer. ...
APC (80-90%) and K-Ras (40-50%) mutations frequently occur in human colorectal cancer (CRC) and these mutations cooperatively accelerate tumorigenesis including metastasis. In addition, both β-catenin and Ras levels are highly increased in CRC, especially in metastatic CRC (mCRC). Therefore, targeting both the Wnt/β-catenin and Ras pathways could be an ideal therapeutic approach for treating mCRC patients. In this study, we characterized the roles of KY1022, a small molecule that destabilizes both β-catenin and Ras via targeting the Wnt/β-catenin pathway, in inhibiting the cellular events, including EMT, an initial process of metastasis, and apoptosis. As shown by in vitro and in vivo studies using APCMin/+/K-RasG12DLA2 mice, KY1022 effectively suppressed the development of mCRC at an early stage of tumorigenesis. A small molecular approach degrading both β-catenin and Ras via inhibition of the Wnt/β-catenin signaling would be an ideal strategy for treatment of mCRC.
... Recently, we discovered that Ras proteins can be degraded by inhibition of the Wnt/β-catenin pathway via its negative regulators such as APC, axin and glycogen synthase kinase 3β (GSK3β) [16][17][18][19] . When the Wnt/β-catenin signaling is negatively regulated, the Ras degradation occurs together with the β-catenin degradation; Ras is phosphorylated by GSK3β, and β-TrCP E3 linker is subsequently recruited for ultimate proteasomal degradation 16 . ...
... Immunoprecipitation and ubiquitination assays. Immunoprecipitation and in vivo ubiquitination assays were performed as previously described 19 . Briefly, cells were washed in ice-cold PBS and lysed with RIPA buffer. ...
Both the Wnt/β-catenin and Ras pathways are aberrantly activated in most human colorectal cancers (CRCs) and interact cooperatively in tumor promotion. Inhibition of these signaling may therefore be an ideal strategy for treating CRC. We identified KY1220, a compound that destabilizes both β-catenin and Ras, via targeting the Wnt/β-catenin pathway, and synthesized its derivative KYA1797K. KYA1797K bound directly to the regulators of G-protein signaling domain of axin, initiating β-catenin and Ras degradation through enhancement of the β-catenin destruction complex activating GSK3β. KYA1797K effectively suppressed the growth of CRCs harboring APC and KRAS mutations, as shown by various in vitro studies and by in vivo studies using xenograft and transgenic mouse models of tumors induced by APC and KRAS mutations. Destabilization of both β-catenin and Ras via targeting axin is a potential therapeutic strategy for treatment of CRC and other type cancers activated Wnt/β-catenin and Ras pathways.
... Our previous work has shown that valproic acid, a drug used to treat epilepsy and bipolar disorder, increases pan-Ras levels during neuronal differentiation of NSCs (Jung et al., 2008). In addition, we have investigated regulatory mechanisms of Ras protein stability in normal and cancer cells (Jeong et al., 2012;Kim et al., 2009;Moon et al., 2014). ...
... The following plasmids were used in these experiments and have been described previously (Jeon et al., 2007;Kim et al., 2009;Park et al., 2006): pcDNA3.1, pcDNA3.1-HRas, ...
Ras signaling is tightly regulated during neural stem cell (NSC) differentiation, and defects in this pathway result in aberrant brain development. However, the mechanism regulating Ras signaling during NSC differentiation was unknown. Here, we show that stabilized HRas specifically induces neuronal differentiation of NSCs. Lentivirus-mediated HRas overexpression and knockdown resulted in stimulation and inhibition, respectively, of NSC differentiation into neuron in the ex vivo embryo. Retinoic acid (RA), an active metabolite of vitamin A, promoted neuronal differentiation of NSCs by stabilizing HRas, and HRas knockdown blocked the RA effect. Vitamin A-deficient mice displayed abnormal brain development with reduced HRas levels and reduced thickness of the postmitotic region containing differentiated neurons. All of these abnormal phenotypes were rescued with the restoration of HRas protein levels by RA-supplemented diet. In summary, this study shows that RA stabilizes HRas protein during neurogenesis, and that required for the NSC differentiation into neuron and murine brain development.
... An interesting dichotomy exists because Ras has been reported to exhibit both positive and negative effects on the pathway (4 -6). Moreover, although synergistic activation of transformation by Ras and Wnt/-catenin has been reported, other reports have identified an antagonistic role in transformation (5,6). How Ras impacts the Wnt/-catenin pathway is still not fully understood, nor is the balance of cellular factors that dictate whether a net positive or negative effect is observed for Ras on Wnt/-catenin pathway signaling. ...
... Over a decade of study has revealed multiple, subtle links between the Ras and Wnt/-catenin pathways. The links are complex and remain poorly understood because they invoke both synergistic and antagonistic relationships (6). Indeed, Ras has been reported to suppress -catenin levels but also to promote -catenin transcription (31,32). ...
Ras is the most frequently activated oncogene found in human cancer, but its mechanisms of action remain only partially understood.
Ras activates multiple signaling pathways to promote transformation. However, Ras can also exhibit a potent ability to induce
growth arrest and death. NORE1A (RASSF5) is a direct Ras effector that acts as a tumor suppressor by promoting apoptosis and
cell cycle arrest. Expression of NORE1A is frequently lost in human tumors, and its mechanism of action remains unclear. Here
we show that NORE1A forms a direct, Ras-regulated complex with β-TrCP, the substrate recognition component of the SCFβ-TrCP ubiquitin ligase complex. This interaction allows Ras to stimulate the ubiquitin ligase activity of SCFβ-TrCP toward its target β-catenin, resulting in degradation of β-catenin by the 26 S proteasome. However, the action of Ras/NORE1A/β-TrCP
is substrate-specific because IκB, another substrate of SCFβ-TrCP, is not sensitive to NORE1A-promoted degradation. We identify a completely new signaling mechanism for Ras that allows for
the specific regulation of SCFβ-TrCP targets. We show that the NORE1A levels in a cell may dictate the effects of Ras on the Wnt/β-catenin pathway. Moreover,
because NORE1A expression is frequently impaired in tumors, we provide an explanation for the observation that β-TrCP can
act as a tumor suppressor or an oncogene in different cell systems.
... Furthermore, it has been shown that Ras is activated at the cellular poles during cytokinesis in Dictyostelium (14,31,39,40) and that K-Ras is shown to have an important role in cytokinesis and genome stability in mammalian cells (32,41,42). The findings in Dictyostelium RasG potentially reflect the circumstance of tumorigenesis driven by hyper-activation of K-Ras: the expression of mutated K-Ras not only activates downstream signaling but also induces aneuploidy and promotes tumorigenesis (33,34,43,44). ...
... Recently, three types of ubiquitin-mediated Ras regulatory pathways have been discovered: 1) ubiquitination of Ras isoforms at Lys-117 or Lys-147 leads to RasGEF-independent Ras activation (14,39,40,45); 2) ubiquitination of H-Ras by the E3-ubiquitin ligase RabEX5 results in H-Ras endomembrane translocation (41,42); 3) ubiquitination of phosphorylated Ras isoforms by TrCP, another E3 ligase, results in Ras proteasome-mediated degradation (43,44). So far, K-Ras-specific degradation has not been described in any system. ...
Mammalian cells encode three closely related Ras proteins, H-Ras, N-Ras, and K-Ras. Oncogenic K-Ras mutations frequently occur
in human cancers, which lead to dysregulated cell proliferation and genomic instability. However, mechanistic role of the
Ras isoform regulation have remained largely unknown. Furthermore, the dynamics and function of negative regulation of GTP-loaded
K-Ras have not been fully investigated. Here, we demonstrate RasG, the Dictyostelium orthologue of K-Ras, is targeted for degradation by polyubiquitination. Both ubiquitination and degradation of RasG were
strictly associated with RasG activity. High resolution tandem mass spectrometry (LC-MS/MS) analysis indicated that RasG ubiquitination
occurs at C-terminal lysines equivalent to lysines found in human K-Ras but not in H-Ras and N-Ras homologues. Substitution
of these lysine residues with arginines (4KR-RasG) diminished RasG ubiquitination and increased RasG protein stability. Cells
expressing 4KR-RasG failed to undergo proper cytokinesis and resulted in multinucleated cells. Ectopically expressed human
K-Ras undergoes polyubiquitin-mediated degradation in Dictyostelium, whereas human H-Ras and a Dictyostelium H-Ras homologue (RasC) are refractory to ubiquitination. Our results indicate the existence of GTP-loaded K-Ras orthologue-specific
degradation system in Dictyostelium, and further identification of the responsible E3-ligase may provide a novel therapeutic approach against K-Ras-mutated cancers.
... P phosphorylation. Literature: (1) Torre et al. (2011); (2) Hwang et al. (2012); (3) Faivre and Lange (2007); (4) Musgrove (2004); (5) Civenni et al. (2003); (6) Zeller et al. (2012); (7) Guturi et al. (2012); (8) Halleskog and Schulte (2013); (9) Kawasaki et al. (2009); (10) Kawasaki et al. (2010); (11) Park et al. (2006); (12) Jeon et al. (2007); (13) Kim et al. (2009b); (14) Jeong et al. (2012). b Crosstalk from RAS/MAPK to WNT/b-catenin. ...
... Wild-type b-catenin was indispensable for this negative regulation of RAS signaling and cellular proliferation by Axin ( Jeon et al. 2007). A similar model of Axin and bTRCP-dependent HA-RAS degradation is proposed by Kim et al. (2009b). This is in line with the observations from the same group that the b-TRCP-dependent polyubiquitinylation and subsequent degradation of the RAS protein is contributed by negative regulators of WNT/bcatenin signaling, thus resulting in RAS stabilization after mutational activation of b-catenin ( Jeong et al. 2012). ...
Signaling through the WNT/β-catenin and the RAS (rat sarcoma)/MAPK (mitogen-activated protein kinase) pathways plays a key role in the regulation of various physiological cellular processes including proliferation, differentiation, and cell death. Aberrant mutational activation of these signaling pathways is closely linked to the development of cancer in many organs, in humans as well as in laboratory animals. Over the past years, more and more evidence for a close linkage of the two oncogenic signaling cascades has accumulated. Using different experimental approaches, model systems, and experimental conditions, a variety of molecular mechanisms have been identified by which signal transduction through WNT/β-catenin and RAS interact, either in a synergistic or an antagonistic manner. Mechanisms of interaction comprise an upstream crosstalk at the level of pathway-activating ligands and their receptors, interrelations of cytosolic kinases involved in either pathways, as well as interaction in the nucleus related to the joint regulation of target gene transcription. Here, we present a comprehensive review of the current knowledge on the interaction of RAS/MAPK- and WNT/β-catenin-driven signal transduction in mammalian cells.
... Several other studies identified the regulatory role of the RAS/ERK axis by crosstalk with the Wnt/βcatenin pathway, which is connected directly with the cell's proliferation and transformation (53,54) through activation of Wnt3a to the Raf-1-MEK-ERK axis, revealing direct interaction of the two pathways. The regulatory effect of Wnt/β-catenin signaling on the MEK-ERK pathway in the proliferation and transformation of cells was further approved by modulating the Wnt/β-catenin axis components, such as GSK3β, Axin, and APC (54,55). Wnt/β-catenin controls the stability regulation of RAS through GSK3β kinase that mediated phosphorylation of RAS, suggesting the established crosstalk between the Wnt/β-catenin and RAS-ERK pathways (56). ...
... The "hot-spot" mutations lead to Ras retain in GTP-bound state, and directly activate several downstream signaling pathways including the MAPK and PI3K pathways, and is involved in the progression of human cancers (43). Additionally, emerging evidence suggests that posttranslational modification regulated the stabilization of Ras proteins and associated with tumorigenesis (11,44,45). Although microRNA-mediated regulation of RAS family genes has well been studied (46), posttranscriptional modification and regulation of the RAS genes remain poorly understood. ...
Overexpression of Ras, in addition to the oncogenic mutations, occurs in various human cancers. However, the mechanisms for epitranscriptic regulation of RAS in tumorigenesis remain unclear. Here, we report that the widespread N6-methyladenosine (m6A) modification of HRAS, but not KRAS and NRAS, is higher in cancer tissues compared with the adjacent tissues, which results in the increased expression of H-Ras protein, thus promoting cancer cell proliferation and metastasis. Mechanistically, three m6A modification sites of HRAS 3' UTR, which is regulated by FTO and bound by YTHDF1, but not YTHDF2 nor YTHDF3, promote its protein expression by the enhanced translational elongation. In addition, targeting HRAS m6A modification decreases cancer proliferation and metastasis. Clinically, up-regulated H-Ras expression correlates with down-regulated FTO and up-regulated YTHDF1 expression in various cancers. Collectively, our study reveals a linking between specific m6A modification sites of HRAS and tumor progression, which provides a new strategy to target oncogenic Ras signaling.
... For example, SMURF2 governs the chromatin organization, dynamics, and genome integrity by controlling the proteasomal degradation or the protein stability of its substrates including RNF20 or DNA topoisomerase IIa 41,43 , which in turn regulate tumorigenesis and tumor progression. Moreover, SMURF2 regulates the stability of pro-oncogenic transcription factors such as KLF5, YY1, and ID1 [44][45][46] , in addition to regulating Wnt/β-catenin oncogenic signaling and KRAS oncoproteins [47][48][49] . Although we show here that SMURF2 Thr249 phosphorylation plays a crucial role in stemness and tumorigenicity by modulating TGF-β signaling through the ubiquitin-proteasome-dependent degradation of TGFBR proteins in GSCs, it should be emphasized that additional molecular mechanisms might be involved in the control of tumorigenicity in GSCs by SMURF2 Thr249 phosphorylation. ...
Glioma stem cells (GSCs) contribute to the pathogenesis of glioblastoma, the most malignant form of glioma. The implication and underlying mechanisms of SMAD specific E3 ubiquitin protein ligase 2 (SMURF2) on the GSC phenotypes remain unknown. We previously demonstrated that SMURF2 phosphorylation at Thr ²⁴⁹ (SMURF2 Thr249 ) activates its E3 ubiquitin ligase activity. Here, we demonstrate that SMURF2 Thr249 phosphorylation plays an essential role in maintaining GSC stemness and tumorigenicity. SMURF2 silencing augmented the self-renewal potential and tumorigenicity of patient-derived GSCs. The SMURF2 Thr249 phosphorylation level was low in human glioblastoma pathology specimens. Introduction of the SMURF2 T249A mutant resulted in increased stemness and tumorigenicity of GSCs, recapitulating the SMURF2 silencing. Moreover, the inactivation of SMURF2 Thr249 phosphorylation increases TGF-β receptor (TGFBR) protein stability. Indeed, TGFBR1 knockdown markedly counteracted the GSC phenotypes by SMURF2 T249A mutant. These findings highlight the importance of SMURF2 Thr249 phosphorylation in maintaining GSC phenotypes, thereby demonstrating a potential target for GSC-directed therapy.
... Previous studies demonstrated that increased RAS stability could drive oncogenic transformation. Higher stability of RAS proteins in colorectal cancer could be due to decreased expression of the E3 ubiquitin ligase NEDD4 (38) or dysregulation of b-TrCPmediated RAS degradation in APC-mutated tumors (39,40). Another proteostatic regulator of RAS proteins is the leucine zipper-like transcriptional regulator 1 (LZTR1)/CUL3 ubiquitin-ligase complex that controls localization and expression levels of RAS proteins (19,41). ...
Meningiomas are the most common benign brain tumors. Mutations of the E3 ubiquitin ligase TRAF7 occur in 25% of meningiomas and commonly cooccur with mutations in KLF4, yet the functional link between TRAF7 and KLF4 mutations remains unclear. By generating an in vitro meningioma model derived from primary meningeal cells, we elucidated the cooperative interactions that promote meningioma development. By integrating TRAF7-driven ubiquitinome and proteome alterations in meningeal cells and the TRAF7 interactome, we identified TRAF7 as a proteostatic regulator of RAS-related small GTPases. Meningioma-associated TRAF7 mutations disrupted either its catalytic activity or its interaction with RAS GTPases. TRAF7 loss in meningeal cells altered actin dynamics and promoted anchorage-independent growth by inducing CDC42 and RAS signaling. TRAF deficiency–driven activation of the RAS/MAPK pathway promoted KLF4-dependent transcription that led to upregulation of the tumor-suppressive Semaphorin pathway, a negative regulator of small GTPases. KLF4 loss of function disrupted this negative feedback loop and enhanced mutant TRAF7-mediated cell transformation. Overall, this study provides new mechanistic insights into meningioma development, which could lead to novel treatment strategies.
Significance
The intricate molecular cross-talk between the ubiquitin ligase TRAF7 and the transcription factor KLF4 provides a first step toward the identification of new therapies for patients with meningioma.
... Overexpression of RAS protein can also affect the occurrence of cancer, including hepatocellular carcinomas (HCC) and is associated with poor prognosis in patients [60]. Stabilization of RAS proteins participates in the activation of downstream signaling pathways associated with tumorigenesis [61][62][63][64]. Jeong et al used purified glutathione-S-transferase (GST)-fused HRAS protein (GST-HRAS) to determined HRAS-binding partner proteins in tissues of HCC tumors expressing high level of RAS compared with normal liver tissues [16]. ...
WD40 repeat (WDR) domain is one of the most abundant protein interaction domains in the human proteome. More than 360 protein interaction domains have been annotated until now. WDR domains frequently mediate interaction with peptide regions of important interaction partners during a variety of biological processes. Proteins with WDR domain which is typically a seven-bladed β propeller, are continuously being discovered. They represent a large class of proteins that are likely to be central proteins. WDR domain-containing protein 76 (WDR76) is a member of WDR domain-containing protein. It is still poorly understood with possible involvement in DNA damage repair, apoptosis, cell cycle progression, and gene expression regulation. With further study on WDR76, knowledge of its basic functions and its role in different pathophysiological processes are increasing. Diverse interactions between WDR76 and its partners are complex and made a study on WDR76 challenging. In the present review, we summarized the process of understanding WDR76, its physiological functions, the close relationship with human diseases, and further potential opportunity for target therapy.
... TrCP1 and β-TrCP2 axis is important to activate DNA-damage response and repair pathway to maintain the genome integrity. have shown that β-TrCP1 can function as a tumor suppressor while β-TrCP2can function as an oncogene(17)(18)(36)(37)(38)(39)(40)(41)(42). It is also reported that tumor suppressors play important role in DNA damage response(43-45) and β-TrCP is known to be involved in DNA damage(16,(21)(22)(23). ...
F-box proteins β-TrCP1 and β-TrCP2 are paralogs present in the human genome. They control several cellular processes including cell cycle and DNA damage signaling. Moreover, it is reported that they facilitate DNA damage-induced accumulation of p53 by directing proteasomal degradation of MDM2, a protein that promotes p53 degradation. However, the individual roles of β-TrCP1 and β-TrCP2 in the genotoxic stress-induced activation of cell cycle checkpoints and DNA damage repair remains largely unknown. Here, using biochemical, molecular biology, flow cytometric, and immunofluorescence techniques, we show that β-TrCP1 and β-TrCP2 communicate during genotoxic stress. We found that expression levels of β-TrCP1 are significantly increased while levels of β-TrCP2 are markedly decreased upon induction of genotoxic stress. Further, our results revealed that DNA damage-induced activation of ATM kinase plays an important role in maintaining the reciprocal expression levels of β-TrCP1 and β-TrCP2 via the phosphorylation of β-TrCP1 at Ser158. Phosphorylated β-TrCP1 potently promotes the proteasomal degradation of β-TrCP2 and MDM2, resulting in the activation of p53. Additionally, β-TrCP1 impedes MDM2 accumulation via abrogation of its lysine 63-linked polyubiquitination by β-TrCP2. Thus, β-TrCP1 helps to arrest cells at the G2/M phase of the cell cycle and promotes DNA repair upon DNA damage through attenuation of β-TrCP2. Collectively, our findings elucidate an intriguing post-translational regulatory mechanism of these two paralogs under genotoxic stress and revealed β-TrCP1 as a key player in maintaining the genome integrity through the attenuation of β-TrCP2 levels in response to genotoxic stress.
... Smurf2 and UbcH5 as a critical E3 and E2, respectively, are important in maintaining K-Ras protein stability, and targeting such a complex was supposed to be a unique strategy to degrade mutant K-Ras G12/V or C to kill cancer cells [51]. However, knock-down Smurf2 can accumulate the F-box protein βTrCP1 which mediates poly-ubiquitination and proteasome-dependent degradation of Ras [51,77]. H-and N-Ras, but not K-Ras, are subjected to mono-and K63-linked di-ubiquitination, and stabilize their associations with the endosome, resulting in a change in the signaling output [49], while K-Ras shows only minor or transient association with the endosome [78]. ...
Abstract The ubiquitin system, known as a common feature in eukaryotes, participates in multiple cellular processes, such as signal transduction, cell-cycle progression, receptor trafficking and endocytosis, and even the immune response. In lung cancer, evidence has revealed that aberrant events in ubiquitin-mediated processes can cause a variety of pathological outcomes including tumorigenesis and metastasis. Likewise, ubiquitination on the core components contributing to the activity of cell signaling controls bio-signal turnover and cell final destination. Given this, inhibitors targeting the ubiquitin system have been developed for lung cancer therapies and have shown great prospects for clinical application. However, the exact biological effects and physiological role of the drugs used in lung cancer therapies are still not clearly elucidated, which might seriously impede the progress of treatment. In this work, we summarize current research advances in cell signal regulation processes mediated through the ubiquitin system during the development of lung cancer, with the hope of improving the therapeutic effects by means of aiming at efficient targets.
... In this manner, Smurf2 may also contribute to the activity of the proto-oncoprotein β-catenin. Intriguingly, GSK3β also phosphorylates KRAS, leading KRAS degradation by β-TrCP [86]. Inhibition of β-TrCP mediated degradation might facilitate KRAS induced colorectal cancer transformation [79,87]. ...
Smad ubiquitination regulatory factor 1 (Smurf1) and Smurf2 are HECT-type E3 ubiquitin ligases, and both Smurfs were initially identified to regulate Smad protein stability in the TGF-β/BMP signaling pathway. In recent years, Smurfs have exhibited E3 ligase-dependent and -independent activities in various kinds of cells. Smurfs act as either potent tumor promoters or tumor suppressors in different tumors by regulating biological processes, including metastasis, apoptosis, cell cycle, senescence and genomic stability. The regulation of Smurfs activity and expression has therefore emerged as a hot spot in tumor biology research. Further, the Smurf1- or Smurf2-deficient mice provide more in vivo clues for the functional study of Smurfs in tumorigenesis and development. In this review, we summarize these milestone findings and, in turn, reveal new avenues for the prevention and treatment of cancer by regulating Smurfs.
... In addition, and more classically, polyubiquitination can alter RAS function through degradation. Proteasomal degradation of HRAS has been reported to occur in response to activation by the Wnt/b-catenin signaling pathway (85)(86)(87). This polyubiquitindependent degradation occurs upon recruitment of β-TrCP-E3 ligase after HRAS phosphorylation by glycogen synthase kinase 3β. ...
Many sensory and chemical signal inputs are transmitted by intracellular GTP-binding (G) proteins. G proteins make up two major subfamilies: "large" G proteins comprising three subunits and "small" G proteins, such as the proto-oncogene product RAS, which contains a single subunit. Members of both subfamilies are regulated by post-translational modifications, including lipidation, proteolysis and carboxyl methylation. Emerging studies have shown that these proteins are also modified by ubiquitination. Much of our current understanding of this post-translational modification comes from investigations of the large G protein α subunit from yeast (Gpa1) and the three RAS isotypes in humans, NRAS, KRAS and HRAS. Gα undergoes both mono- and polyubiquitination, and these modifications have distinct consequences for determining the sites and mechanisms of its degradation. Genetic and biochemical reconstitution studies have revealed the enzymes and binding partners required for addition and removal of ubiquitin, as well as the delivery and destruction of both the mono- and polyubiquitinated forms of the G protein. Complementary studies of RAS have identified multiple ubiquitination sites, each having distinct consequences for binding to regulatory proteins, shuttling to and from the plasma membrane and degradation. Here, we review what is currently known about these two well-studied examples, Gpa1 and the human RAS proteins, which have revealed additional mechanisms of signal regulation and dysregulation relevant to human physiology. We also compare and contrast the effects of G protein ubiquitination with other post-translational modifications of these proteins.
... LZRT1 functions as a substrate receptor in the cullin 3 ubiquitin ligase complex involved in the ubiquitination and functional down modulation of HRAS. 33,74,75 Interestingly, the SCF-b-TrCP E3 ligase complex has also been implicated in the ubiquitination and proteasomal degradation of HRAS, 76 supporting an unanticipated functional link between FBXW11 and RAS signaling modulation warranting further exploration. ...
The identification of genetic variants implicated in human developmental disorders has been revolutionized by second-generation sequencing combined with international pooling of cases. Here, we describe seven individuals who have diverse yet overlapping developmental anomalies, and who all have de novo missense FBXW11 variants identified by whole exome or whole genome sequencing and not reported in the gnomAD database. Their phenotypes include striking neurodevelopmental, digital, jaw, and eye anomalies, and in one individual, features resembling Noonan syndrome, a condition caused by dysregulated RAS signaling. FBXW11 encodes an F-box protein, part of the Skp1-cullin-F-box (SCF) ubiquitin ligase complex, involved in ubiquitination and proteasomal degradation and thus fundamental to many protein regulatory processes. FBXW11 targets include β-catenin and GLI transcription factors, key mediators of Wnt and Hh signaling, respectively, critical to digital, neurological, and eye development. Structural analyses indicate affected residues cluster at the surface of the loops of the substrate-binding domain of FBXW11, and the variants are predicted to destabilize the protein and/or its interactions. In situ hybridization studies on human and zebrafish embryonic tissues demonstrate FBXW11 is expressed in the developing eye, brain, mandibular processes, and limb buds or pectoral fins. Knockdown of the zebrafish FBXW11 orthologs fbxw11a and fbxw11b resulted in embryos with smaller, misshapen, and underdeveloped eyes and abnormal jaw and pectoral fin development. Our findings support the role of FBXW11 in multiple developmental processes, including those involving the brain, eye, digits, and jaw.
... One of the key events in the crosstalk between the two pathways is the stabilization of RAS, especially oncogenic mutant KRAS, as well as β-catenin, caused by APC loss. RAS, as well as β-catenin, is degraded via glycogen synthase kinase 3 beta (GSK3β)-mediated phosphorylation and subsequent poly-ubiquitination-dependent proteasomal degradation; however, the inactivation of GSK3β by the loss of APC stabilizes these proteins 9,10 . ...
A recently identified small molecule shows promise for tackling resistance to a leading colorectal cancer drug. Three proteins that are over-expressed in colorectal cancer are epidermal growth factor receptor (EGFR), RAS and β-catenin. These proteins and their interconnected signaling pathways are therefore important therapeutic targets. EGFR is the target of the drug cetuximab, but many patients are resistant to this drug attributed to mutations in a gene that influences the signaling pathways of the three key proteins. Kang-Yell Choi at Yonsei University in Seoul, South Korea, and co-workers trialed a novel molecular drug on human colorectal cancer tissues and on mice. They confirmed that the new drug leads to reduced EGFR levels by degrading RAS and β-catenin and therefore suppresses the growth of colorectal cancer cells in samples with or without the resistant mutations.
... Interestingly, GSK3β phosphorylates and primes RAS proteins for SCF β−TrCP -mediated degradation (73). Inhibition of this degradation pathway by aberrant Wnt/β-catenin signaling may contribute to Ras-induced transformation in colorectal tumorigenesis (68). ...
Protein ubiquitination is an evolutionary conserved highly-orchestrated enzymatic cascade essential for normal cellular functions and homeostasis maintenance. This pathway relies on a defined set of cellular enzymes, among them, substrate-specific E3 ubiquitin ligases (E3s). These ligases are the most critical players, as they define the spatiotemporal nature of ubiquitination and confer specificity to this cascade. Smurf1 and Smurf2 (Smurfs) are the C2-WW-HECT-domain E3 ubiquitin ligases, which recently emerged as important determinants of pivotal cellular processes. These processes include cell proliferation and differentiation, chromatin organization and dynamics, DNA damage response and genomic integrity maintenance, gene expression, cell stemness, migration, and invasion. All these processes are intimately connected and profoundly altered in cancer. Initially, Smurf proteins were identified as negative regulators of the bone morphogenetic protein (BMP) and the transforming growth factor beta (TGF-β) signaling pathways. However, recent studies have extended the scope of Smurfs' biological functions beyond the BMP/TGF-β signaling regulation. Here, we provide a critical literature overview and updates on the regulatory roles of Smurfs in molecular and cell biology, with an emphasis on cancer. We also highlight the studies demonstrating the impact of Smurf proteins on tumor cell sensitivity to anticancer therapies. Further in-depth analyses of Smurfs' biological functions and influences on molecular pathways could provide novel therapeutic targets and paradigms for cancer diagnosis and treatment.
... Most previous approaches to control the activity of oncogenic mutant Ras have involved searches for small molecules that bind directly to Ras, farnesyltransferase inhibitors, and inducers of synthetic lethality [9][10][11][12][13][14] . As an alternative approach controlling Ras activity, on the basis of our studies illustrated the cross-talk between the Wnt/ β-catenin and Ras-ERK pathways 23,25,34,35 , we identified and characterized small molecules degrading Ras 20, 21 , especially oncogenic mutant K-Ras. These compounds and derivatives, KYA1797K and KY1022, inhibited the growth of CRC cells via degradation of both β-catenin and Ras 20,21 . ...
Although the development of drugs that control Ras is an emerging topic in cancer therapy, no clinically applicable drug is currently available. We have previously utilized knowledge of the Wnt/β-catenin signaling-dependent mechanism of Ras protein stability regulation to identify small molecules that inhibit the proliferation and transformation of various colorectal cancer (CRC) cells via degradation of both β-catenin and Ras. Due to the absence of Ras degradation in cells expressing a nondegradable mutant form of β-catenin and the need to determine an alternative mechanism of Ras degradation, we designed a cell-based system to screen compounds that degrade Ras independent of the Wnt/β-catenin signaling pathway. A cell-based high-content screening (HCS) system that monitors the levels of EGFP-K-RasG12V was established using HCT-116 cells harboring a nondegradable mutant CTNNB1 (ΔS45). Through HCS of a chemical library composed of 10,000 compounds and subsequent characterization of hits, we identified several compounds that degrade Ras without affecting the β-catenin levels. KY7749, one of the most effective compounds, inhibited the proliferation and transformation of CRC cells, especially KRAS-mutant cells that are resistant to the EGFR monoclonal antibody cetuximab. Small molecules that degrade Ras independent of β-catenin may able to be used in treatments for cancers caused by aberrant EGFR and Ras.
... The pJFH1/GFP 60 harboring a GFP-coding gene in the context of genotype 2a fulllength HCV (JFH1) cDNA clone was provided by Dr. Xulin Chen (Institute for Virus Research, Chinese Academic of Sciences, Wuhan, China). The pCS4-3xHA-Ub plasmid 61 , which expresses a HA-tagged ubiquitin, was provided by Prof. Jong-Bok Yoon (Yonsei University, Seoul, Korea). Plasmids expressing hTLR4, MD2, and CD14 62 individually were provided by Prof. Seung Hyun Han (Dental Research Institute, School of Dentistry, Seoul National University, Seoul, Korea). ...
The recognition of pathogen-derived ligands by pattern recognition receptors activates the innate immune response, but the potential interaction of quorum-sensing (QS) signaling molecules with host anti-viral defenses remains largely unknown. Here we show that the Vibrio vulnificus QS molecule cyclo(Phe-Pro) (cFP) inhibits interferon (IFN)-β production by interfering with retinoic-acid-inducible gene-I (RIG-I) activation. Binding of cFP to the RIG-I 2CARD domain induces a conformational change in RIG-I, preventing the TRIM25-mediated ubiquitination to abrogate IFN production. cFP enhances susceptibility to hepatitis C virus (HCV), as well as Sendai and influenza viruses, each known to be sensed by RIG-I but did not affect the melanoma-differentiation-associated gene 5 (MDA5)-recognition of norovirus. Our results reveal an inter-kingdom network between bacteria, viruses and host that dysregulates host innate responses via a microbial quorum-sensing molecule modulating the response to viral infection.
... 38 The Raf-1-MEK-ERK pathway regulation via the Wnt/βcatenin signaling and its roles in the proliferation and transformation of cells were further confirmed by the experiments modulating of the components of the Wnt/β-catenin signaling, such as Axin, APC, and GSK3β, and so on. [35][36][37]39 The highlight in the cross-talk between the Wnt/β-catenin and RAS-ERK pathways is the stability regulation of RAS by the Wnt/β-catenin signaling involving the GSK3β kinase mediated phosphorylaton of RAS. 40 In a resting status of Wnt/β-catenin signaling, HRas protein was subjected to the phosphorylations at the threonine (Thr)-144 and Thr-148 by GSK3β, and subsequently the β-transducin repeatcontaining protein (β-TrCP) E3 linker is recruited to the phosphorylated Ras and degraded by polyubiqutinationdependent proteasomal degradation (Fig. 4a). ...
Aberrant activation of the Wnt/β-catenin and RAS-extracellular signal-regulated kinase (ERK) pathways play important roles in the tumorigenesis of many different types of cancer, most notably colorectal cancer (CRC). Genes for these two pathways, such as adenomatous polyposis coli (APC) and KRAS are frequently mutated in human CRC, and involved in the initiation and progression of the tumorigenesis, respectively. Moreover, recent studies revealed interaction of APC and KRAS mutations in the various stages of colorectal tumorigenesis and even in metastasis accompanying activation of the cancer stem cells (CSCs). A key event in the synergistic cooperation between Wnt/β-catenin and RAS-ERK pathways is a stabilization of both β-catenin and RAS especially mutant KRAS by APC loss, and pathological significance of this was indicated by correlation of increased β-catenin and RAS levels in human CRC where APC mutations occur as high as 90% of CRC patients. Together with the notion of the protein activity reduction by lowering its level, inhibition of both β-catenin and RAS especially by degradation could be a new ideal strategy for development of anti-cancer drugs for CRC. In this review, we will discuss interaction between the Wnt/β-catenin and RAS-ERK pathways in the colorectal tumorigenesis by providing the mechanism of RAS stabilization by aberrant activation of Wnt/β-catenin. We will also discuss our small molecular anti-cancer approach controlling CRC by induction of specific degradations of both β-catenin and RAS via targeting Wnt/β-catenin pathway especially for the KYA1797K, a small molecule specifically binding at the regulator of G-protein signaling (RGS)-domain of Axin.
... HRAS has been shown to be polyubiquitinated when co-overexpressed with FLAG-Ub (Jeong et al. 2012). The E3 ligase was identified as β-TrCP (Kim et al. 2009). Zeng et al. reported interactions of H,N and KRAS4B with the E3 ligase Nedd4-1 and showed polyubiquitination of KRAS4B on Lys5 using HA-Ub. ...
The three human RAS genes encode four proteins that play central roles in oncogenesis by acting as binary molecular switches that regulate signaling pathways for growth and differentiation. Each is subject to a set of posttranslational modifications (PTMs) that modify their activity or are required for membrane targeting. The enzymes that catalyze the various PTMs are potential targets for anti-RAS drug discovery. The PTMs of RAS proteins are the focus of this review. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.
... In light of this, though attempts to effectively pharmacologically inhibit K-RAS have been unsuccessful, recent data suggests that targeted inhibition of ubiquitin modifying enzymes may lead to downregulation of K-RAS function. The F-box protein β-transducin repeat-containing protein (β-TrCP) mediates polyubiquitination of all RAS isoforms, leading to proteasome-dependent degradation of RAS [148]. Interestingly, this process is partly mediated by Wnt/β-catenin signalling pathway whereby Axin and Adenomatous Polyposis Coli (APC) enhance the binding of the WD40 domain of β-TrCP with H-RAS leading to downregulation of the transforming capability of H-RAS. ...
... The F-box protein β-transducin repeats-containing protein (β-TrCP) promotes poly-ubiquitination and proteasome-mediated degradation of all Ras isoforms and inhibits the Ras transformation ability in cells [99]. In addition, Wnt signaling decreases β-TrCP-induced polyubiquitination of Ras, thereby enhancing Ras activities [100]. ...
Ras proteins are molecular switches cycling between the GTP-bound state and the GDP-bound state to transduce signals from outside of the cell to the interior. They all have the CAAX motifs at the C-terminus. Post-translational modifications of the CAAX motifs determine the subcellular localization and correct biological function of Ras proteins. Here we have described how prenylation of the CAAX motifs affects membrane association of Ras and how this modification can be targeted to block Ras-transduced signaling. In addition, other post-translational modifications of Ras, such as palmitoylation, phosphorylation, acetylation, ubiquitination, and S-nitrosylation, are discussed. Because oncogenic Ras mutants are drivers for tumorigenesis, understanding how post-translational modifications of Ras mutants influence their activities will help us design therapeutic drugs to treat patients with cancer.
... AXIN1 has been reported to inhibit Ras/ERK signaling, although the effect appeared to be dependent on β-catenin so it is not clear if AXIN plays a direct role in the modulation of ERK signaling56 . A second study linking AXIN1 to Ras regulation found that both AXIN1 and APC can promote Ras degradation via the same E3-ligase that targets β-catenin57 . AXIN1 has also been show to interact with MEKK1 and MEKK4 and to activate JNK when overexpressed58; 59 . ...
The axis inhibition proteins 1 and 2 (AXIN1, AXIN2) are negative regulators of Wnt/beta-catenin signaling. The Wnt pathway has a key role in cell fate and development and is dysregulated in 90% of colorectal cancers (CRCs). In 2011, we reported a family with a CRC/oligodontia (lack of several teeth) phenotype. Affected individuals carry a heterozygous, germline mutation in AXIN2. This mutation introduces a premature stop codon and segregates with the oligodontia/CRC phenotype. This family is the second case of an AXIN2 mutation associated with oligodontia/CRC.
A premature stop codon suggests that mutant transcripts might undergo nonsense-mediated degradation, but transcripts from both wild-type and mutant allele were detected in the proband???s lymphocytes, predicting that the mutant allele can encode a truncated protein, trAXIN2. The trAXIN2 protein is more stable than wid-type, displayed a reduced function in Wnt/beta-catenin target gene inhibition, retained interactions with beta-catenin, AXIN1 and AXIN2, and inhibited colony formation in a Wnt-dependent CRC cell line similar to AXIN2. This protein appears to be hypomorphic, which could produce reduced function in vivo, but with massive over-expression in vitro, increased protein stability may mask a reduced functionality. A knock-in mouse model has been generated to further explore the significance of this allele in cancer and development.
Sequence alterations in AXIN1 and AXIN2 have been reported in a subset of many cancers. Whether these mutations have an active role in cancer is unclear. In a mouse model of CRC, loss of one or both alleles of Axin2 did not alter survival time, but may alter the distribution of tumors in the colon. These studies are ongoing. The AXIN proteins were classically thought of as tumor suppressors, but some studies suggest that high levels of AXIN2 might actually promote tumor phenotypes such as invasion and metastasis. Moreover, stabilization of AXIN proteins has been proposed as a therapeutic strategy for cancer; therefore, it is important to better understand the role of AXIN proteins in normal cells and cancer cells. This work has begun to clarify how AXIN2 functions at a biochemical level, and how AXIN2 mutations or changes in gene dosage impact tumor development.
... 最 好 的 例 子 莫 过 于 小 三 磷 酸 鸟 苷 (guanosine triphosphate, GTP)酶 Kirsten 鼠肉瘤病毒癌 基因(Kirsten rat sarcoma viral oncogene, K-RAS), 在 缺乏受体刺激的情况下, 对其单泛素化修饰即可激活 K-RAS. 在以往的研究中, 研究者发现 RAS 被 Rabex-5 连 接 酶 泛 素 化 修 饰 可 以 调 控 其 内 体 的 定 位[60] , 而被-TrCP 连接酶泛素化则介导其蛋白酶体 降 解[61] , 然 而 近 期 研 究却 发 现 泛 素本 身 可 以 影响 K-RAS 对 GTP 酶 激 活 蛋 白 (GTPase-activating proteins, GAPs)的反应从而增加 GTP-RAS 的量, 进而 增加下游通路的激活[62,63] . 另一方面, 研究者又发现 RAS 的 泛 素 化 可 以 增 强 其 与 磷 脂 酰 肌 醇 3-激 酶(phosphatidylinositol 3-kinase, PI3K)的相互作用, 从 而强化 PI3K-蛋白激酶 B(protein kinase B, AKT)信号 通路. ...
... A DNA fragment encoding the human Sur8 protein was inserted into a pGEX-4T-1 vector using XhoI and BamHI restriction sites. Using this plasmid, recombinant human Sur8 protein was expressed in Escherichia coli and purified using glutathione sepharose 4B (Sigma) as previously described [48]. Recombinant His-tagged p110α (His-p110α) (Upstate Biotechnology, 3 μg) was incubated with GST or GST-Sur8 (3 μg) at 4°C for 1 hour, and then pulled down using glutathione agarose. ...
Sur8 (also known as Shoc2) is a Ras-Raf scaffold protein that modulates signaling through extracellular signal-regulated kinase (ERK) pathway. Although Sur8 has been shown to be a scaffold protein of the Ras-ERK pathway, its interaction with other signaling pathways and its involvement in tumor malignancy has not been reported. We identified that Sur8 interacts with the p110α subunit of phosphatidylinositol 3-kinase (PI3K), as well as with Ras and Raf, and these interactions are increased in an epidermal growth factor (EGF)- and oncogenic Ras-dependent manner. Sur8 regulates cell migration and invasion via activation of Rac and matrix metalloproteinases (MMPs). Interestingly, using inhibitors of MEK and PI3K we found Sur8 mediates these cellular behaviors predominantly through PI3K pathway. We further found that human metastatic melanoma tissues had higher Sur8 content followed by activations of Akt, ERK, and Rac. Lentivirus-mediated Sur8-knockdown attenuated metastatic potential of highly invasive B16-F10 melanoma cells indicating the role of Sur8 in melanoma metastasis. This is the first report to identify the role of scaffold protein Sur8 in regulating cell motility, invasion, and metastasis through activation of both ERK and PI3K pathways.
... Animals were allowed to acclimate for three days before exposed to normobaric hypoxia (10% O 2 ). 2,15,16 Then mice were randomized to receive weekly rmWnt5a injections at 75 ng per mouse 17,18 in a final volume of 150 mL or equal volume of phosphate buffered saline (PBS) via the tail vein from the first day to the end of the two or four weeks after exposed to hypoxia. ...
Hypoxic pulmonary hypertension (HPH), which is characterized by pulmonary arteriolar remodeling and right ventricular hypertrophy, is still a life-threatening disease with the current treatment strategies. The underlying molecular mechanisms of HPH remain unclear. Our previously published study showed that Wnt5a, one of the ligands in the Wnt family, was critically involved in the inhibition of hypoxia-induced pulmonary arterial smooth muscle cell proliferation by downregulation of β-catenin/cyclin D1 in vitro. In this study, we investigated the possible functions and mechanisms of Wnt5a in HPH in vivo. Recombinant mouse Wnt5a (rmWnt5a) or phosphate buffered saline (PBS) was administered to male C57/BL6 mice weekly from the first day to the end of the two or four weeks after exposed to hypoxia (10% O2). Hypoxia-induced pulmonary hypertension was associated with a marked increase in β-catenin/cyclin D1 expression in lungs. Right ventricular systolic pressure and right ventricular hypertrophy index were reduced in animals treated with rmWnt5a compared with PBS. Histology showed less pulmonary vascular remodeling and right ventricular hypertrophy in the group treated with rmWnt5a than with PBS. Treatment with rmWnt5a resulted in a concomitant reduction in β-catenin/cyclin D1 levels in lungs. These data demonstrate that Wnt5a exerts its beneficial effects on HPH by regulating pulmonary vascular remodeling and right ventricular hypertrophy in a manner that is associated with reduction in β-catenin/cyclin D1 signaling. A therapy targeting the β-catenin/cyclin D1 signaling pathway might be a potential strategy for HPH treatment.
... 24 More recently, H-Ras was found to be polyubiquitylated and degraded via proteasome by Wnt/bcatenin signaling. 26 How MAT2B affects Ras protein stability will require further examination. ...
Methionine adenosyltransferase 2B (MAT2B) encodes for variant proteins V1 and V2 that interact with GIT1 to increase ERK activity and growth in human liver and colon cancer cells. MAT2B or GIT1 overexpression activates MEK. This study explores the mechanism for MEK activation. We examined protein-protein interactions by co-immunoprecipitation and verified by confocal microscopy and pull-down assay using recombinant or in vitro translated proteins. Results were confirmed in an orthotopic liver cancer model. We found that MAT2B and GIT1-mediated MEK1/2 activation was not mediated by PAK1 or Src in HepG2 or RKO cells. Instead, MAT2B and GIT1 interact with B-Raf and c-Raf and enhance recruitment of Raf proteins to MEK1/2. MAT2B-GIT1 activates c-Raf, which is the key mediator for MEK/12 activation, because this still occurred in RKO cells that express constitutively active B-Raf mutant. The mechanism lies with the ability of MAT2B-GIT1 to activate Ras and promote B-Raf/c-Raf heterodimerization. Interestingly, MAT2B but not GIT1 can directly interact with Ras, which increases protein stability. Finally, increased Ras-Raf-MEK signaling occurred in phenotypically more aggressive liver cancers overexpressing MAT2B variants and GIT1. In conclusion, interaction between MAT2B and GIT1 serves as a scaffold and facilitates signaling in multiple steps of the Ras/Raf/MEK/ERK pathway, further emphasizing the importance of MAT2B/GIT1 interaction in cancer growth.
Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.
... A good example is the oncogenic GTPase K-Ras, which is activated by monoubiquitination even in the absence of receptor stimulation. It was previously known that ubiquitination of Ras by Rabex-5 ligase regulates its endosomal localization 25 and that Ras ubiquitination mediated by β-TrCP ligase leads to its proteasomal degradation 26 . It was recently proposed, on the basis of structural and mutational studies, that Ub also impairs the response of K-Ras to GTPase-activating proteins (GAPs) and thus increases the amount of GTP-bound (activated) Ras as well as its binding to downstream effectors 27,28 . ...
Ubiquitination is crucial for a plethora of physiological processes, including cell survival and differentiation and innate and adaptive immunity. In recent years, considerable progress has been made in the understanding of the molecular action of ubiquitin in signaling pathways and how alterations in the ubiquitin system lead to the development of distinct human diseases. Here we describe the role of ubiquitination in the onset and progression of cancer, metabolic syndromes, neurodegenerative diseases, autoimmunity, inflammatory disorders, infection and muscle dystrophies. Moreover, we indicate how current knowledge could be exploited for the development of new clinical therapies.
... While some findings suggest that increased levels of AXIN proteins inhibit colon cancer cells that are dependent on β-catenin signaling, a recent mouse study suggests the opposite: that high levels of AXIN2 actually promote cancer cell invasion and perhaps metastasis [69]. Additionally, the AXIN proteins have been reported to be involved in multiple non-canonical signaling pathways, including the Ras/ERK [70,71] and SAPK/JNK pathways [72], so it will also be valuable to assess if any of the reported mutations in AXIN1/2 could promote tumorigenesis via a nonβ-catenin dependent mechanism. Without clear evidence of the pathogenicity of the mutations identified to date or strong evidence from animal models, the importance of AXIN1 and AXIN2 as tumor suppressor genes or oncogenes in cancer and the potential role of the variants in non-Wnt pathway functions remain unresolved issues of keen interest. ...
Mutations in the APC (adenomatous polyposis coli) gene, which encodes a multi-functional protein with a well-defined role in the canonical Wnt pathway, underlie familial adenomatous polypsosis, a rare, inherited form of colorectal cancer (CRC) and contribute to the majority of sporadic CRCs. However, not all sporadic and familial CRCs can be explained by mutations in APC or other genes with well-established roles in CRC. The AXIN1 and AXIN2 proteins function in the canonical Wnt pathway, and AXIN1/2 alterations have been proposed as key defects in some cancers. Here, we review AXIN1 and AXIN2 sequence alterations reported in gastrointestinal cancers, with the goal of vetting the evidence that some of the variants may have key functional roles in cancer development.
... Recent studies showed that mono-or diubiquitination restricted activities of H-Ras and N-Ras (Colicelli, 2010;Xu et al., 2010;Yan et al., 2009Yan et al., , 2010, whereas mono-ubiquitination of K-Ras promoted its activity (Sasaki et al., 2011). It has been reported that H-Ras can also be targeted for polyubiquitination and degradation by b-TrCP (Kim et al., 2009), and aberrant activation of Wnt/b-catenin signaling promotes intestinal tumorigenesis by stabilizing H-Ras (Jeong et al., 2012). However, it remains unclear whether it has to be distinct ubiquitin ligases for different Ras proteins, or one same ubiquitin ligase can target all three forms of Ras, and what roles of Ras ubiquitination would exert in tumorigenesis (Pfleger, 2011). ...
RAS genes are among the most frequently mutated proto-oncogenes in cancer. However, how Ras stability is regulated remains largely unknown. Here, we report a regulatory loop involving the E3 ligase Nedd4-1, Ras, and PTEN. We found that Ras signaling stimulates the expression of Nedd4-1, which in turn acts as an E3 ubiquitin ligase that regulates Ras levels. Importantly, Ras activation, either by oncogenic mutations or by epidermal growth factor (EGF) signaling, prevents Nedd4-1-mediated Ras ubiquitination. This leads to Ras-induced Nedd4-1 overexpression, and subsequent degradation of the tumor suppressor PTEN in both human cancer samples and cancer cells. Our study thus unravels the molecular mechanisms underlying the interplay of Ras, Nedd4-1, and PTEN and suggests a basis for the high prevalence of Ras-activating mutations and EGF hypersignaling in cancer.
... It however remains to be discovered whether a similar regulatory mechanism exists for other Ras isoforms under other cellular contexts. It is also noteworthy that all Ras isoforms are subject to polyubiquitination mediated by the F-box protein b-TrCP (b-transducin repeat-containing protein), leading to proteasome-dependent degradation of Ras [61]. In conclusion, the above studies suggest that ubiquitination is an essential mechanism controlling Ras compartmentalisation and its signalling output. ...
Ubiquitination, the covalent attachment of ubiquitin to target proteins, has emerged as a ubiquitous post-translational modification (PTM) whose function extends far beyond its original role as a tag for protein degradation identified three decades ago. Although sharing parallel properties with phosphorylation, ubiquitination distinguishes itself in important ways. Nevertheless, the interplay and crosstalk between ubiquitination and phosphorylation events have become a recurrent theme in cell signalling regulation. Understanding how these two major PTMs intersect to regulate signal transduction is an important research question. In this review, we first discuss the involvement of ubiquitination in the regulation of the EGF-mediated ERK signalling pathway via the EGF receptor, highlighting the interplay between ubiquitination and phosphorylation in this cancer-implicated system and addressing open questions. The roles of ubiquitination in pathways crosstalking to EGFR/MAPK signalling will then be discussed. In the final part of the review, we demonstrate the rich and versatile dynamics of crosstalk between ubiquitination and phosphorylation by using quantitative modelling and analysis of network motifs commonly observed in cellular processes. We argue that given the overwhelming complexity arising from inter-connected PTMs, a quantitative framework based on systems biology and mathematical modelling is needed to efficiently understand their roles in cell signalling.
... In particular, H-Ras and N-Ras can be mono-and di-ubiquitinated by Rabex5, a Rab-specific GEF endowed with E3 ligase activity, and such modification promotes their internalization into endosomes and a concomitant attenuation of the Ras/ERK signaling [13]. Additionally, H-Ras can be polyubiquitinated by SCF(beta-TrCP)E3 ubiquitin ligase resulting in proteosomal degradation [14]. Mono-ubiquitination of K-Ras4A, preferentially at lysine 147, seems to promote GDP/GTP exchange, and thus protein activation and signal transduction. ...
Introduction:
Ras proteins are small GTPases molecular switches that cycle through two alternative conformational states, a GDP-bound inactive state and a GTP-bound active state. In the active state, Ras proteins interact with and modulate the activity of several downstream effectors regulating key cellular processes including proliferation, differentiation, survival, senescence, migration and metabolism. Activating mutations of RAS genes and of genes encoding Ras signaling members have a great incidence in proliferative disorders, such as cancer, immune and inflammatory diseases and developmental syndromes. Therefore, Ras and Ras signaling represent important clinical targets for the design and development of pharmaceutically active agents, including anticancer agents.
Areas covered:
The authors summarize methods available to down-regulate the Ras pathway and review recent patents covering Ras signaling modulators, as well as methods designed to kill specifically cancer cells bearing activated RAS oncogene.
Expert opinion:
Targeted therapy approach based on direct targeting of molecules specifically altered in Ras-dependent diseases is pursued with molecules that down-regulate expression or inhibit the biological function of mutant Ras or Ras signaling members. The low success rate in a clinical setting of molecules targeting activated members of the Ras pathway may require development of novel approaches, including combined and synthetic lethal therapies.
... The Rab subfamily of small GTPases consists of 57 members in A. thaliana, most of which are involved in vesicle trafficking (Vernoud et al. 2003). The interaction between APD2 and the Rab homologue E1b implies a possible link between E3 ligases and small GTPases in plants, which was previously reported only in animals (Wilkins et al. 2004;Moren et al. 2005;Jura et al. 2006;Schwamborn et al. 2007;Berthold et al. 2008;Kim et al. 2009;Kawabe et al. 2010). In conclusion, we provide evidence to show that the four APDs, APD1, 2, 3 and 4, play important but redundant roles in the PMII process during male gametogenesis. ...
Ubiquitination of proteins is one of the critical regulatory mechanisms in eukaryotes. In higher plants, protein ubiquitination plays an essential role in many biological processes, including hormone signaling, photomorphogenesis, and pathogen defense. However, the roles of protein ubiquitination in the reproductive process are not clear. In this study, we identified four plant-specific RING-finger genes designated Aberrant Pollen Development 1 (APD1) to APD4, as regulators of pollen mitosis II (PMII) in Arabidopsis thaliana (L.). The apd1 apd2 double mutant showed a significantly increased percentage of bicellular-like pollen at the mature pollen stage. Further downregulation of the APD3 and APD4 transcripts in apd1 apd2 by RNA interference (RNAi) resulted in more severe abnormal bicellular-like pollen phenotypes than in apd1 apd2, suggesting that cell division was defective in male gametogenesis. All of the four genes were expressed in multiple stages at different levels during male gametophyte development. Confocal analysis using green florescence fusion proteins (GFP) GFP-APD1 and GFP-APD2 showed that APDs are associated with intracellular membranes. Furthermore, APD2 had E2-dependent E3 ligase activity in vitro, and five APD2-interacting proteins were identified. Our results suggest that these four genes may be involved, redundantly, in regulating the PMII process during male gametogenesis.
... In both these pathways kinase GSK3 performs a pre-required phosphorylation step. Proto-oncogene H-Ras is also a target for βTrCP ubiquitination [98]. As a result, βTrCP has both promoting and inhibiting roles in EMT. ...
Epithelial to Mesenchymal transition (EMT) in cancer, a process permitting cancer cells to become mobile and metastatic, has a signaling hardwire forged from development. Multiple signaling pathways that regulate carcinogenesis enabling characteristics in neoplastic cells such as proliferation, resistance to apoptosis and angiogenesis are also the main players in EMT. These pathways, as almost all cellular processes, are in their turn regulated by ubiquitination and the Ubiquitin-Proteasome System (UPS). Ubiquitination is the covalent link of target proteins with the small protein ubiquitin and serves as a signal to target protein degradation by the proteasome or to other outcomes such as endocytosis, degradation by the lysosome or specification of cellular localization. This paper reviews signal transduction pathways regulating EMT and being regulated by ubiquitination.
... 16 -18). It is noteworthy that the proto-oncogenes H-Ras and N-Ras are ubiquitinated (18,19), and this ubiquitination markedly disrupts the balance between plasma and endomembrane localization of Ras, favoring the latter and disfavoring signal transduction toward ERK1/2 (18,20,21). We show here that the endogenous RalA and RalB GTPases are ubiquitinated, which contributes to selective membrane localization. ...
Ras GTPases signal by orchestrating a balance among several effector pathways, of which those driven by the GTPases RalA and
RalB are essential to Ras oncogenic functions. RalA and RalB share the same effectors but support different aspects of oncogenesis.
One example is the importance of active RalA in anchorage-independent growth and membrane raft trafficking. This study has
shown a new post-translational modification of Ral GTPases: nondegradative ubiquitination. RalA (but not RalB) ubiquitination
increases in anchorage-independent conditions in a caveolin-dependent manner and when lipid rafts are endocytosed. Forcing
RalA mono-ubiquitination (by expressing a protein fusion consisting of ubiquitin fused N-terminally to RalA) leads to RalA
enrichment at the plasma membrane and increases raft exposure. This study suggests the existence of an ubiquitination/de-ubiquitination
cycle superimposed on the GDP/GTP cycle of RalA, involved in the regulation of RalA activity as well as in membrane raft trafficking.
... The selective degradation of small GTPases is best characterised for rho family proteins (described below), but it is likely that this control system will be widely used in other related pathways. For example, H-Ras has recently been shown to be poly-ubiquitylated by SCF-TrCP under the control of the Wnt signalling pathway [44]. ...
A variety of post-translational modifications such as phosphorylation, acetylation and ubiquitylation transduce cellular signals, which culminate in changes in gene transcription. In this article we examine the ways in which selective protein degradation provides an extra dimension to the regulation of such signalling cascades. We discuss (i) how both lysosomal and proteasomal systems are used to attenuate kinase and rho family GTPase signalling, thereby coupling activation with degradation, (ii) signal propagation contingent upon the selective degradation of inhibitory components, exemplified by the degradation of IκB to activate NF-κB signalling, and (iii) tonic suppression of signalling pathways by turnover of the transcription factors β-catenin and p53.
Mutation in RAS gene is one of the most common genetic alterations, which seems to be seen in one third of human cancers.
Ras, as a molecular switch, has been considered in wide variety of signaling pathways such as cell division and apoptosis. Ras
proteins have a binary function to transmit diferent extracellular messages into intracellular signaling network. It has been
proved that Ras proteins associate with diferent plasma membranes. Although all Ras isoforms have been found at plasma
membrane, H-Ras and N-Ras are located in Golgi, and K-Ras at ER and mitochondria outer membrane. There have been a
lot of eforts to inhibit Ras signaling that can be a pivotal approach to treat Ras-induced tumors. Efect of RalA and RalB on
the growth of embryonal tumors, at downstream region of Ras, has been studied in a number of studies, which showed that
inhibition of these signaling pathways can provide a strong therapeutic approach to cancer. Also, post translational modifca-
tions (PTMs) in proteins interfere extremely with cell signaling pathways in cells that can react to external signals. In this
review, the role of Ral signaling in cancer and PTM of Ras proteins has been reviewed.
Ras/MAPK (mitogen active protein kinase) signaling plays contradictory roles in adipocyte differentiation and is tightly regulated during adipogenesis. However, mechanisms regulating adipocyte differentiation involving Ras protein stability regulation are unknown. Here, we show that WD40 repeat protein 76 (WDR76), a novel Ras regulating E3 linker protein, controls 3T3-L1 adipocyte differentiation through HRas stability regulation. The roles of WDR76 in obesity and metabolic regulation were characterized using a high-fat diet (HFD)-induced obesity model using Wdr76−/− mice and liver-specific Wdr76 transgenic mice (Wdr76Li−TG). Wdr76−/− mice are resistant to HFD-induced obesity, insulin resistance and hyperlipidemia with an increment of HRas levels. In contrast, Wdr76Li-TG mice showed increased HFD-induced obesity, insulin resistance with reduced HRas levels. Our findings suggest that WDR76 controls HFD-induced obesity and hepatic steatosis via HRas destabilization. These data provide insights into the links between WDR76, HRas, and obesity.
The three RAS genes — HRAS, NRAS and KRAS — are collectively mutated in one-third of human cancers, where they act as prototypic oncogenes. Interestingly, there are rather distinct patterns to RAS mutations; the isoform mutated as well as the position and type of substitution vary between different cancers. As RAS genes are among the earliest, if not the first, genes mutated in a variety of cancers, understanding how these mutation patterns arise could inform on not only how cancer begins but also the factors influencing this event, which has implications for cancer prevention. To this end, we suggest that there is a narrow window or ‘sweet spot’ by which oncogenic RAS signalling can promote tumour initiation in normal cells. As a consequence, RAS mutation patterns in each normal cell are a product of the specific RAS isoform mutated, as well as the position of the mutation and type of substitution to achieve an ideal level of signalling.
F-box proteins, which are the substrate-recognition subunits of SKP1-cullin 1-F-box protein (SCF) E3 ligase complexes, have pivotal roles in multiple cellular processes through ubiquitylation and subsequent degradation of target proteins. Dysregulation of F-box protein-mediated proteolysis leads to human malignancies. Notably, inhibitors that target F-box proteins have shown promising therapeutic potential, urging us to review the current understanding of how F-box proteins contribute to tumorigenesis. As the physiological functions for many of the 69 putative F-box proteins remain elusive, additional genetic and mechanistic studies will help to define the role of each F-box protein in tumorigenesis, thereby paving the road for the rational design of F-box protein-targeted anticancer therapies.
While it is very clear that activation of β-catenin through the canonical Wnt pathway plays a role in a multitude of human
cancers, it has also been noted time and time again that activation of β-catenin is observed in tumors without any clear mutation
in any of the major components of the pathway or any increase in Wnt signaling. This suggests that other factors maybe capable
of inducing activation and downstream signaling via β-catenin. Indeed, multiple growth factor and developmental signaling
pathways have been found to transactivate β-catenin. It is also reasonable to consider the converse: that the Wnt pathway
is capable of transactivating other signaling pathways. In fact, multiple studies have outlined the capability of Wnt signaling
to activate other pathways in various types of cancer. Such association has led to the initiative to develop approaches to
therapy that are capable of targeting both the Wnt pathway and the pathways it crosstalks with simultaneously. The crosstalk
that occurs between Wnt and other signaling pathways will be explored in this chapter with a primary focus on how these pathways
interact in the development and progression of cancer.
KeywordsCrosstalk-Cancer-Signaling-Wnt-Notch-Hedgehog-Growth factor-Receptor tyrosine-kinase-Ras-PI3K-mTOR-Cox-2-Growth-Metastasis-Proliferation-Development-Suppression-Transactivation
Epithelial to Mesenchymal Transition (EMT) in cancer is a process that allows cancer cells to detach from neighboring cells, become mobile and metastasize and shares many signaling pathways with development. Several molecular mechanisms which regulate oncogenic properties in neoplastic cells such as proliferation, resistance to apoptosis and angiogenesis through transcription factors or other mediators are also regulators of EMT. These pathways and downstream transcription factors are, in their turn, regulated by ubiquitination and the Ubiquitin-Proteasome System (UPS). Ubiquitination, the covalent link of the small 76-amino acid protein ubiquitin to target proteins, serves as a signal for protein degradation by the proteasome or for other outcomes such as endocytosis, degradation by the lysosome or directing these proteins to specific cellular compartments. This review discusses aspects of the regulation of EMT by ubiquitination and the UPS and underlines its complexity focusing on transcription and transcription factors regulating EMT and are being regulated by ubiquitination.
The regulation of the small GTPases leading to their membrane localization has long been attributed to processing of their C-terminal CAAX box. As deregulation of many of these GTPases have been implicated in cancer and other disorders, prenylation and methylation of this CAAX box has been studied in depth as a possibility for drug targeting, but unfortunately, to date no drug has proved clinically beneficial. However, these GTPases also undergo other modifications that may be important for their regulation. Ubiquitination has long been demonstrated to regulate the fate of numerous cellular proteins and recently it has become apparent that many GTPases, along with their GAPs, GeFs and GDis, undergo ubiquitination leading to a variety of fates such as re-localization or degradation. in this review we focus on the recent literature demonstrating that the regulation of small GTPases by ubiquitination, either directly or indirectly, plays a considerable role in controlling their function and that targeting these modifications could be important for disease treatment.
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