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Chlamydia infection activates ER stress pathways that are dependent and independent of TLR4 signalling. (A) XBP-1 splicing in mDC following infection with C. trachomatis (CT) MOI ¼ 20 for 4 h in the presence of 4m8C (30 mM) or GSK PERK inhibitor D3 (PERKi) (1 mM) n ¼ 4 independent donors **p ¼ <0.01. Data represented as ±SEM. (B) XBP-1 splicing in mDC following infection with C. trachomatis (CT) MOI ¼ 20 in the presence of a TLR4 blocking antibody (aTLR4) (10 mg/ml) n ¼ 4 independent donors *p ¼ <0.05. (C) XBP-1 splicing in mDC following LPS stimulation (1 mg/ml) for 4 h in the presence of 4m8C (30 mM) or GSK PERK inhibitor D3 (PERKi) (1 mM) n ¼ 4 independent donors **p ¼ <0.01. Data represented as ±SEM. (D) XBP-1 splicing in mDC following LPS stimulation (1 mg/ml) for 4 h in the presence of a TLR4 blocking antibody (aTLR4) (10 mg/ml) n ¼ 6 independent donors **p ¼ <0.01. (E) CHOP mRNA expression in mDC following infection with C. trachomatis (CT) MOI ¼ 20 for 4 h in the presence of 4m8C (30 mM) or GSK PERK inhibitor D3 (PERKi) (1 mM) n ¼ 4 independent donors. Data represented as ±SEM. (F) CHOP mRNA expression in mDC following infection with C. trachomatis (CT) MOI ¼ 20 for 24 h in the presence of a TLR4 blocking antibody (aTLR4) (10 mg/ml). Data represented as ±SEM from 1 experiment performed in triplicate wells ***p ¼ <0.001. (G) CHOP mRNA expression in wild type ( gcn2 þ/þ ) or GCN2 knock out ( gcn2 À/À ) MEFs following infection with C. trachomatis or C. muridarum (CM) MOI ¼ 10 for 24 h. Data represented as ±SEM from 1 experiment performed in triplicate wells ***p ¼ <0.001.
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Protein kinase RNA activated (PKR) is a crucial mediator of anti-viral responses but is reported to be activated by multiple non-viral stimuli. However, mechanisms underlying PKR activation, particularly in response to bacterial infection, remain poorly understood. We have investigated mechanisms of PKR activation in human primary monocyte-derived...
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Context 1
... have previously reported that C. trachomatis infection of mDC induces activation of the Integrated Stress Response (ISR) resulting in CHOP expression that enhances inflamma- tory responses [12]. However, there are no published data investigating IRE1a activation in response to Chlamydia infection. We therefore investigated XBP-1 splicing as an in- dicator of IRE1a activation in response to C. trachomatis infection (Fig. 5A). Infection induced robust XBP-1 splicing that was inhibited by 4m8c but not by GSK PERK inhibitor D3 demonstrating that C. trachomatis infection was causing activation of IRE1a RNAse activity. Furthermore, we found that Chlamydia infection-induced IRE1a activation was dependent on TLR4 signalling as XBP-1 splicing was reduced in the presence of a TLR4 blocking antibody, and similar re- sults were obtained with LPS as a control (Fig. 5C and D). We also confirmed that C. trachomatis infection induced CHOP expression in mDC, indicating activation of the ISR (Fig. 5E). Surprisingly, CHOP expression was independent of IRE1a and PERK activation as 4m8c and GSK PERK inhibitor D3 had no effect on CHOP mRNA expression. Furthermore, Chlamydia infection-induced CHOP expression was independent of TLR4 signalling (Fig. 5F) as blocking TLR4 signalling with the TLR4 blocking antibody, resulted in increased CHOP expression in response to C. trachomatis suggesting that in- duction of the ISR occurs independently of LPS and TLR4 and is therefore distinct to the activation of IRE1a and PKR. Mammalian cells also express two additional eIF2a kinases, namely GCN2 and HRI, which are activated in response to amino acid or heme depletion respectively [27]. Chlamydiae sp have been termed 'energy parasites' as they utilise host cell metabolites such as amino acids [28]. Given that we have provided evidence that CHOP induction was independent of PERK and TLR4 induced PKR activation, we tested the hy- pothesis that CHOP induction in response to Chlamydia infection occurs through activation of the amino acid respon- sive eIF2a kinase GCN2. To do this, we infected wild type ( gcn2 þ/þ ) or GCN2 knock out ( gcn2 À/À ) MEFs with C. tra- chomatis or the murine pathogen Chlamydia muridarum (that induces a more potent CHOP response than C. trachomatis in MEFs) and examined CHOP expression (Fig. 5G). Interest- ingly, induction of CHOP expression by C. trachomatis or C. muridarum infection was entirely GCN2 dependent indicating that although infection resulted in PKR activation, GCN2 is the likely eIF2a kinase responsible for the induction of the ISR and is independent of IRE1a, PKR, PERK and TLR4 ...
Context 2
... have previously reported that C. trachomatis infection of mDC induces activation of the Integrated Stress Response (ISR) resulting in CHOP expression that enhances inflamma- tory responses [12]. However, there are no published data investigating IRE1a activation in response to Chlamydia infection. We therefore investigated XBP-1 splicing as an in- dicator of IRE1a activation in response to C. trachomatis infection (Fig. 5A). Infection induced robust XBP-1 splicing that was inhibited by 4m8c but not by GSK PERK inhibitor D3 demonstrating that C. trachomatis infection was causing activation of IRE1a RNAse activity. Furthermore, we found that Chlamydia infection-induced IRE1a activation was dependent on TLR4 signalling as XBP-1 splicing was reduced in the presence of a TLR4 blocking antibody, and similar re- sults were obtained with LPS as a control (Fig. 5C and D). We also confirmed that C. trachomatis infection induced CHOP expression in mDC, indicating activation of the ISR (Fig. 5E). Surprisingly, CHOP expression was independent of IRE1a and PERK activation as 4m8c and GSK PERK inhibitor D3 had no effect on CHOP mRNA expression. Furthermore, Chlamydia infection-induced CHOP expression was independent of TLR4 signalling (Fig. 5F) as blocking TLR4 signalling with the TLR4 blocking antibody, resulted in increased CHOP expression in response to C. trachomatis suggesting that in- duction of the ISR occurs independently of LPS and TLR4 and is therefore distinct to the activation of IRE1a and PKR. Mammalian cells also express two additional eIF2a kinases, namely GCN2 and HRI, which are activated in response to amino acid or heme depletion respectively [27]. Chlamydiae sp have been termed 'energy parasites' as they utilise host cell metabolites such as amino acids [28]. Given that we have provided evidence that CHOP induction was independent of PERK and TLR4 induced PKR activation, we tested the hy- pothesis that CHOP induction in response to Chlamydia infection occurs through activation of the amino acid respon- sive eIF2a kinase GCN2. To do this, we infected wild type ( gcn2 þ/þ ) or GCN2 knock out ( gcn2 À/À ) MEFs with C. tra- chomatis or the murine pathogen Chlamydia muridarum (that induces a more potent CHOP response than C. trachomatis in MEFs) and examined CHOP expression (Fig. 5G). Interest- ingly, induction of CHOP expression by C. trachomatis or C. muridarum infection was entirely GCN2 dependent indicating that although infection resulted in PKR activation, GCN2 is the likely eIF2a kinase responsible for the induction of the ISR and is independent of IRE1a, PKR, PERK and TLR4 ...
Context 3
... have previously reported that C. trachomatis infection of mDC induces activation of the Integrated Stress Response (ISR) resulting in CHOP expression that enhances inflamma- tory responses [12]. However, there are no published data investigating IRE1a activation in response to Chlamydia infection. We therefore investigated XBP-1 splicing as an in- dicator of IRE1a activation in response to C. trachomatis infection (Fig. 5A). Infection induced robust XBP-1 splicing that was inhibited by 4m8c but not by GSK PERK inhibitor D3 demonstrating that C. trachomatis infection was causing activation of IRE1a RNAse activity. Furthermore, we found that Chlamydia infection-induced IRE1a activation was dependent on TLR4 signalling as XBP-1 splicing was reduced in the presence of a TLR4 blocking antibody, and similar re- sults were obtained with LPS as a control (Fig. 5C and D). We also confirmed that C. trachomatis infection induced CHOP expression in mDC, indicating activation of the ISR (Fig. 5E). Surprisingly, CHOP expression was independent of IRE1a and PERK activation as 4m8c and GSK PERK inhibitor D3 had no effect on CHOP mRNA expression. Furthermore, Chlamydia infection-induced CHOP expression was independent of TLR4 signalling (Fig. 5F) as blocking TLR4 signalling with the TLR4 blocking antibody, resulted in increased CHOP expression in response to C. trachomatis suggesting that in- duction of the ISR occurs independently of LPS and TLR4 and is therefore distinct to the activation of IRE1a and PKR. Mammalian cells also express two additional eIF2a kinases, namely GCN2 and HRI, which are activated in response to amino acid or heme depletion respectively [27]. Chlamydiae sp have been termed 'energy parasites' as they utilise host cell metabolites such as amino acids [28]. Given that we have provided evidence that CHOP induction was independent of PERK and TLR4 induced PKR activation, we tested the hy- pothesis that CHOP induction in response to Chlamydia infection occurs through activation of the amino acid respon- sive eIF2a kinase GCN2. To do this, we infected wild type ( gcn2 þ/þ ) or GCN2 knock out ( gcn2 À/À ) MEFs with C. tra- chomatis or the murine pathogen Chlamydia muridarum (that induces a more potent CHOP response than C. trachomatis in MEFs) and examined CHOP expression (Fig. 5G). Interest- ingly, induction of CHOP expression by C. trachomatis or C. muridarum infection was entirely GCN2 dependent indicating that although infection resulted in PKR activation, GCN2 is the likely eIF2a kinase responsible for the induction of the ISR and is independent of IRE1a, PKR, PERK and TLR4 ...
Context 4
... have previously reported that C. trachomatis infection of mDC induces activation of the Integrated Stress Response (ISR) resulting in CHOP expression that enhances inflamma- tory responses [12]. However, there are no published data investigating IRE1a activation in response to Chlamydia infection. We therefore investigated XBP-1 splicing as an in- dicator of IRE1a activation in response to C. trachomatis infection (Fig. 5A). Infection induced robust XBP-1 splicing that was inhibited by 4m8c but not by GSK PERK inhibitor D3 demonstrating that C. trachomatis infection was causing activation of IRE1a RNAse activity. Furthermore, we found that Chlamydia infection-induced IRE1a activation was dependent on TLR4 signalling as XBP-1 splicing was reduced in the presence of a TLR4 blocking antibody, and similar re- sults were obtained with LPS as a control (Fig. 5C and D). We also confirmed that C. trachomatis infection induced CHOP expression in mDC, indicating activation of the ISR (Fig. 5E). Surprisingly, CHOP expression was independent of IRE1a and PERK activation as 4m8c and GSK PERK inhibitor D3 had no effect on CHOP mRNA expression. Furthermore, Chlamydia infection-induced CHOP expression was independent of TLR4 signalling (Fig. 5F) as blocking TLR4 signalling with the TLR4 blocking antibody, resulted in increased CHOP expression in response to C. trachomatis suggesting that in- duction of the ISR occurs independently of LPS and TLR4 and is therefore distinct to the activation of IRE1a and PKR. Mammalian cells also express two additional eIF2a kinases, namely GCN2 and HRI, which are activated in response to amino acid or heme depletion respectively [27]. Chlamydiae sp have been termed 'energy parasites' as they utilise host cell metabolites such as amino acids [28]. Given that we have provided evidence that CHOP induction was independent of PERK and TLR4 induced PKR activation, we tested the hy- pothesis that CHOP induction in response to Chlamydia infection occurs through activation of the amino acid respon- sive eIF2a kinase GCN2. To do this, we infected wild type ( gcn2 þ/þ ) or GCN2 knock out ( gcn2 À/À ) MEFs with C. tra- chomatis or the murine pathogen Chlamydia muridarum (that induces a more potent CHOP response than C. trachomatis in MEFs) and examined CHOP expression (Fig. 5G). Interest- ingly, induction of CHOP expression by C. trachomatis or C. muridarum infection was entirely GCN2 dependent indicating that although infection resulted in PKR activation, GCN2 is the likely eIF2a kinase responsible for the induction of the ISR and is independent of IRE1a, PKR, PERK and TLR4 ...
Context 5
... have previously reported that C. trachomatis infection of mDC induces activation of the Integrated Stress Response (ISR) resulting in CHOP expression that enhances inflamma- tory responses [12]. However, there are no published data investigating IRE1a activation in response to Chlamydia infection. We therefore investigated XBP-1 splicing as an in- dicator of IRE1a activation in response to C. trachomatis infection (Fig. 5A). Infection induced robust XBP-1 splicing that was inhibited by 4m8c but not by GSK PERK inhibitor D3 demonstrating that C. trachomatis infection was causing activation of IRE1a RNAse activity. Furthermore, we found that Chlamydia infection-induced IRE1a activation was dependent on TLR4 signalling as XBP-1 splicing was reduced in the presence of a TLR4 blocking antibody, and similar re- sults were obtained with LPS as a control (Fig. 5C and D). We also confirmed that C. trachomatis infection induced CHOP expression in mDC, indicating activation of the ISR (Fig. 5E). Surprisingly, CHOP expression was independent of IRE1a and PERK activation as 4m8c and GSK PERK inhibitor D3 had no effect on CHOP mRNA expression. Furthermore, Chlamydia infection-induced CHOP expression was independent of TLR4 signalling (Fig. 5F) as blocking TLR4 signalling with the TLR4 blocking antibody, resulted in increased CHOP expression in response to C. trachomatis suggesting that in- duction of the ISR occurs independently of LPS and TLR4 and is therefore distinct to the activation of IRE1a and PKR. Mammalian cells also express two additional eIF2a kinases, namely GCN2 and HRI, which are activated in response to amino acid or heme depletion respectively [27]. Chlamydiae sp have been termed 'energy parasites' as they utilise host cell metabolites such as amino acids [28]. Given that we have provided evidence that CHOP induction was independent of PERK and TLR4 induced PKR activation, we tested the hy- pothesis that CHOP induction in response to Chlamydia infection occurs through activation of the amino acid respon- sive eIF2a kinase GCN2. To do this, we infected wild type ( gcn2 þ/þ ) or GCN2 knock out ( gcn2 À/À ) MEFs with C. tra- chomatis or the murine pathogen Chlamydia muridarum (that induces a more potent CHOP response than C. trachomatis in MEFs) and examined CHOP expression (Fig. 5G). Interest- ingly, induction of CHOP expression by C. trachomatis or C. muridarum infection was entirely GCN2 dependent indicating that although infection resulted in PKR activation, GCN2 is the likely eIF2a kinase responsible for the induction of the ISR and is independent of IRE1a, PKR, PERK and TLR4 ...
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Citations
... O'Connell et al. [81] identified that TLR2 and MyD88 were specifically recruited to the inclusion body membrane during productive infection with chlamydia and that TLR2 was actively involved in intracellular signaling. Webster et al. [83] discovered that Chlamydia trachomatis infection also resulted in TLR4-dependent and MyD88 signaling-dependent protein kinase RNA activation (PKR) was also efficiently activated. ...
The female reproductive tract consists of the vagina, cervix, uterus, and fallopian tubes. In particular, the lower region of the reproductive tract, which contains the vagina and cervix, is often attacked by various pathogenic microorganisms such as bacteria, fungi, and viruses. The immune response of the female lower genital tract is the first line of defense against pathogenic microorganisms. The toll-like receptors (TLRs), a critical pattern recognition receptor, are essential for fighting infections in the female lower genital tract. Here we give an overview of the current research on TLR expression in the female lower genital tract and review the role of TLRs and their signaling pathways in the identification of numerous pathogens in female lower genital tract infections. Our review will contribute to a deeper understanding of the connection between TLRs and the pathological mechanisms of female lower reproductive tract infections, serving as a reference for both fundamental research and preventative strategies for these diseases.
... HMGB1, an endogenous TLR4 ligand, can trigger in ammatory responses by activating TLR4 [2]. When stimulated by HMGB1, TLR4 further stimulates double-stranded RNA-activated protein kinase R (PKR) [24,25], which controls the secretion of in ammatory factors (TNF-, IL-1, and IL-6) in ALI [26]. To our knowledge, a rise in HMGB1 has not been observed in pulmonary IR. ...
Background Pulmonary ischemia/reperfusion (I/R) injury derived from hemorrhagic shock-resuscitation is a significant cause of death in trauma patients. However, effective preventive interventions in early trauma resuscitation are lacking.
Aims To investigate the critical role of the HMGB1-TLR4 pathway in the early inflammatory response after ischemia-reperfusion lung injury, and its specific regulatory mechanisms and the types of downstream cytokines regulated by this pathway.
Methods In this study, to determine the molecular mechanism underlying pulmonary I/R injury and identify effective therapeutic targets to reduce the incidence and mortality, the expression of the HMGB1-TLR4 pathway in a pulmonary I/R injury model, its correlation with downstream inflammatory factors, and the effects of HMGB1-neutralizing antibodies on inflammation were evaluated.
Results IL-6 and TNF-α levels in the three mouse models showed a rapid increase， IL-1β, IL-6, and TNF-α were upregulated in alveolar macrophages after LPS stimulation，TNF-α and HMGB1 were upregulated in TLR4+/+ cells and peaked at 48 h but was not upregulated in TLR4-/- cells. Western blot assays revealed that in TLR4+/+ cells, TLR4 was upregulated after stimulation by LPS and was rapidly downregulated after treatment with the HMGB1-neutralizing antibody. In contrast, TLR4-/- cells did not respond to LPS stimulation, and the HMGB1-neutralizing antibody did not significantly alter the TLR4 concentration.
Conclusions HMGB1-TLR4 pathway plays an important role in the regulation of inflammation in pulmonary I/R injury. Furthermore, HMGB1 upregulated downstream inflammatory factors via TLR4. HMGB1-neutralizing antibodies had a protective effect against lung injury by downregulating the inflammatory response.
... PKR has also been researched to be closely linked with the UPR [49]. Specifically, a study has shown the RNase activity of IRE1α can directly activate PKR downstream of TLR4 signaling [50]. The unique properties of PKR range from an immune response against viral infection to a survival regulator [51][52][53], but most importantly it has been linked to the phosphorylation of cofilin-1 (Ser3) [54]. ...
... The current research also highlights the underlying mechanisms by which PSP induces a shift in the phosphorylation states of PKR and consequently cofilin-1. It has been postulated in the literature research that both PKR and IRE1α can overlap with each other [50]. Inhibitory experiments in our dataset and validated with two pharmaceutical blockers in THP1 cells have confirmed these studies. ...
... Among them, NF-κB signaling and production of type I IFNs have been researched to gain insights into their respective roles such as survivability and immune response [49,91,92]. Interestingly, a study has also shown that IRE1α directly activates PKR in response to Chlamydia trachomatis infection in a TLR4-dependent manner [50]. Having said this, our previously published data stand out by demonstrating the anti-viral effects of PSP through TLR4 and NF-κB. ...
Actin depolymerization factor (ADF) cofilin-1 is a key cytoskeleton component that serves to lessen cortical actin. HIV-1 manipulates cofilin-1 regulation as a pre- and post-entry requisite. Disruption of ADF signaling is associated with denial of entry. The unfolded protein response (UPR) marker Inositol-Requiring Enzyme-1α (IRE1α) and interferon-induced protein (IFN-IP) double-stranded RNA- activated protein kinase (PKR) are reported to overlap with actin components. In our published findings, Coriolus versicolor bioactive extract polysaccharide peptide (PSP) has demonstrated anti-HIV replicative properties in THP1 monocytic cells. However, its involvement towards viral infectivity has not been elucidated before. In the present study, we examined the roles of PKR and IRE1α in cofilin-1 phosphorylation and its HIV-1 restrictive roles in THP1. HIV-1 p24 antigen was measured through infected supernatant to determine PSP’s restrictive potential. Quantitative proteomics was performed to analyze cytoskeletal and UPR regulators. PKR, IRE1α, and cofilin-1 biomarkers were measured through immunoblots. Validation of key proteome markers was done through RT-qPCR. PKR/IRE1α inhibitors were used to validate viral entry and cofilin-1 phosphorylation through Western blots. Our findings show that PSP treatment before infection leads to an overall lower infectivity. Additionally, PKR and IRE1α show to be key regulators in cofilin-1 phosphorylation and viral restriction.
... ATF4, CHOP, and ATF5 are also central players in the initiation of the mitochondrial UPR (UPR mt ) (10,13). Intriguingly, a study has shown that IRE1 contributes to PKR activation in response to infection by Chlamydia trachomatis, suggesting crosstalk exists between UPR ER and ISR pathways (14). Whether IRE1 has an impact on the activation of PKR or any other eif2α kinases during ER stress conditions is unknown. ...
The unfolded protein response (UPR) is an elaborate signaling network that evolved to maintain proteostasis in the endoplasmic reticulum (ER) and mitochondria (mt). These organelles are functionally and physically associated and consequently, their stress responses are often intertwined. It is unclear how these two adaptive stress responses are coordinated during ER stress. The inositol-requiring enzyme-1 (IRE1), a central ER stress sensor and proximal regulator of the UPRER, harbors dual kinase and endoribonuclease (RNase) activities. IRE1 RNase activity initiates the transcriptional layer of the UPRER, but IRE1’s kinase substrate(s) and their functions are largely unknown. Here, we discovered that sphingosine 1-phosphate (S1P) lyase (SPL), the enzyme that degrades S1P, is a substrate for the mammalian IRE1 kinase. Our data show that IRE1-dependent SPL phosphorylation inhibits SPL’s enzymatic activity, resulting in increased intracellular S1P levels. S1P has previously been shown to induce the activation of mitochondrial UPR (UPRmt) in nematodes. We determined that IRE1 kinase-dependent S1P induction during ER stress potentiates UPRmt signaling in mammalian cells. Phosphorylation of eukaryotic translation initiation factor 2α (eif2α) is recognized as a critical molecular event for UPRmt activation in mammalian cells. Our data further demonstrate that inhibition of the IRE1-SPL axis abrogates the activation of two eif2α kinases, namely double-stranded RNA-activated protein kinase (PKR) and PKR–like ER kinase (PERK) upon ER stress. These findings show that the IRE1-SPL axis plays a central role in coordinating the adaptive responses of both organelles to ER stress in mammalian cells.
... This promotes the release of glycerol and fatty acids and the production of ATP, which in turn is exploited by the pathogen for its survival (37). C. pneumoniae has been shown to induce the UPR by activating IRE1a RNase via the TLR4 pathway (38). The upregulation of Bip is not only an indicator of ERS in mammalian cells, but also of UPR activation (18). ...
Chlamydia psittaci is an important pathogen that causes chronic and atypical pneumonia in humans. Autophagy and the unfolded protein response (UPR) are important mechanisms for regulating the growth of infectious parasitic pathogens in living cells. Here, we explored whether C. psittaci infection induced autophagy via the UPR and the effect of these cellular responses on the survival and replication of C. psittaci in human bronchial epithelial cells (HBEs). Not only were the numbers of autophagosomes and the expression of LC3-II and Beclin1 increased following C. psittaci infection of HBEs, but also the expression of p62 (also called sequestosome-1) was downregulated. Moreover, after C. psittaci infection, the UPR and UPR sensors PERK/eIF2α and IRE1α/XBP1 were activated, but not the ATF6 pathway. When either Bip siRNA was used to block normal initiation of the UPR, or activation of the PERK and IER1α pathways was blocked with specific inhibitors GSK2606414 and 4μ8C, the level of autophagy caused by C. psittaci infection was significantly inhibited. Furthermore, blocking activation of the UPR and associated pathways significantly reduced the number of C. psittaci inclusions. Our research suggests that the UPR, via the PERK and IRE1α, but not ATF6 signaling pathways, regulates HBE-cell autophagy induced by C. psittaci infection and the replication of C. psittaci.
... (Pillich et al., 2012), Chlamydia spp. (Webster et al., 2016), Aspergillus fumigatus (Lee et al., 2016), and a number of viruses (Jheng et al., 2014;Li et al., 2015). As a matter of course, Mycobacterium, a well-known intracellular pathogen, can also induce ER stress during infection. ...
The normal operation of the endoplasmic reticulum (ER) is critical for cells and organisms. However, ER stress, caused by imbalanced protein folding, occurs frequently, which perturbs the function of the ER and even results in cell apoptosis eventually. Many insults can induce ER stress; pathogen infection is one of them. Most of the genes involved in ER stress have been reported to be upregulated in Mycobacterium tuberculosis (Mtb) granulomas of humans and mice, implicating that infection with Mtb can induce ER stress. However, little is known about the molecular mechanism of Mtb induction of ER stress. Here, we reveal that Mycobacterium protein CDP-diglyceride hydrolase of Mycobacteriumn (CdhM) could target the ER and cause abnormal ER morphology and cell death. RNA-seq analysis suggests that most of the ER stress-involved genes were modulated by CdhM. Further assessed by biochemical experiments, the transcription and protein levels of ER stress markers BiP and CHOP, as well as the levels of XBP1 splicing and eIF2α phosphorylation, were significantly increased by CdhM, confirming that CdhM could induce ER stress alone or during infection. A single conserved amino acid mutant of CdhM, including L44A, G96A, H150A, and W175A, was incapable of inducing ER stress, which indicates that induction of ER stress by CdhM is specific and functional. Furthermore, CdhM-induced ER stress could also promote apoptosis of macrophages during Mtb infection. Overexpression of CdhM conferred a significant benefit for Mtb replication by releasing Mtb into extracellular during infection of macrophage in vitro, as presented in CFU assays. Overall, our study identified a novel Mtb effector protein CdhM which may promote Mtb dissemination and proliferation by induction of ER stress and apoptosis and provided new insight into the physiological significance of induction of ER stress in tuberculosis (TB) granulomas.
... A microtubule stabilizing factor with UPR-inducing properties is secreted by Brucella [145]; it is responsible for restructuration of the ER, which becomes condensed and fragmented. Whereas Chlamydia infection induces transient BiP upregulation and eIF2α phosphorylation but no ATF6 cleavage, it triggers IRE1 activation and XBP1 splicing and induces CHOP [146,147]. UPR enables infected cells to sense invasion by pathogens and during infection, providing greater cytokine responses when threats impact cell function than when there is only a PRRs activation. The balance between the PRR stimulation and the degree of ER stress sways the cell to either UPR or microbial stress response. ...
In cystic fibrosis (CF), p.Phe508del is the most frequent mutation in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene. The p.Phe508del-CFTR protein is retained in the ER and rapidly degraded. This retention likely triggers an atypical Unfolded Protein Response (UPR) involving ATF6, which reduces the expression of p.Phe508del-CFTR. There are still some debates on the role of the UPR in CF: could it be triggered by the accumulation of misfolded CFTR proteins in the endoplasmic reticulum as was proposed for the most common CFTR mutation p.Phe508del? Or, is it the consequence of inflammation and infection that occur in the disease? In this review, we summarize recent findings on UPR in CF and show how infection, inflammation and UPR act together in CF. We propose to rethink their respective role in CF and to consider them as a whole.
... In 2001, Horng and colleagues observed that PKR is phosphorylated following treatment with LPS, a TLR4 agonist (133). Since then, there have been numerous reports of PKR activation occurring in a TLR4-dependent manner (87,93,(134)(135)(136)(137)(138). TLR4 signaling has also been shown to increase EIF2AK2 mRNA and protein levels (135). ...
... As such, we speculate that bacteria can trigger downstream PKR phosphorylation and expression by activating TLR4 signaling ( Figure 2). Indeed, a link between TLR4 signaling and PKR activity has been reported during infection with bacteria such as S. Typhimurium, Y. pseudotuberculosis, B. anthracis, and C. trachomatis (87,93). Importantly, both heattreated and gamma-irradiated attenuated C. trachomatis can activate PKR to the same extent as live bacteria, indicating that intracellular bacterial replication or secretion of heat-labile bacterial products are not responsible for PKR activation (93). ...
... Indeed, a link between TLR4 signaling and PKR activity has been reported during infection with bacteria such as S. Typhimurium, Y. pseudotuberculosis, B. anthracis, and C. trachomatis (87,93). Importantly, both heattreated and gamma-irradiated attenuated C. trachomatis can activate PKR to the same extent as live bacteria, indicating that intracellular bacterial replication or secretion of heat-labile bacterial products are not responsible for PKR activation (93). In contrast, inhibition of TLR4 prevented PKR activation (93). ...
The global antimicrobial resistance crisis poses a significant threat to humankind in the coming decades. Challenges associated with the development of novel antibiotics underscore the urgent need to develop alternative treatment strategies to combat bacterial infections. Host-directed therapy is a promising new therapeutic strategy that aims to boost the host immune response to bacteria rather than target the pathogen itself, thereby circumventing the development of antibiotic resistance. However, host-directed therapy depends on the identification of druggable host targets or proteins with key functions in antibacterial defense. Protein Kinase R (PKR) is a well-characterized human kinase with established roles in cancer, metabolic disorders, neurodegeneration, and antiviral defense. However, its role in antibacterial defense has been surprisingly underappreciated. Although the canonical role of PKR is to inhibit protein translation during viral infection, this kinase senses and responds to multiple types of cellular stress by regulating cell-signaling pathways involved in inflammation, cell death, and autophagy – mechanisms that are all critical for a protective host response against bacterial pathogens. Indeed, there is accumulating evidence to demonstrate that PKR contributes significantly to the immune response to a variety of bacterial pathogens. Importantly, there are existing pharmacological modulators of PKR that are well-tolerated in animals, indicating that PKR is a feasible target for host-directed therapy. In this review, we provide an overview of immune cell functions regulated by PKR and summarize the current knowledge on the role and functions of PKR in bacterial infections. We also review the non-canonical activators of PKR and speculate on the potential mechanisms that trigger activation of PKR during bacterial infection. Finally, we provide an overview of existing pharmacological modulators of PKR that could be explored as novel treatment strategies for bacterial infections.
... Following this, the C. trachomatis protein pORF5 activated PERK, along with UPR-associated proteins, and P-eIF2α, suggesting that both the UPR and ISR are activated upon Chlamydia infection (Wen et al., 2020). Furthermore, it was also determined that ER stress induced by Chlamydia infection promoted PKR activation via the Toll-like receptor signaling, possible due to IRE1dependent host degradation activating PKR alongside PERK (Webster et al., 2016). ...
Stress granule (SG) formation is a host cell response to stress-induced translational repression. SGs assemble with RNA-binding proteins and translationally silent mRNA. SGs have been demonstrated to be both inhibitory to viruses, as well as being subverted for viral roles. In contrast, the function of SGs during non-viral microbial infections remains largely unexplored. A handful of microbial infections have been shown to result in host SG assembly. Nevertheless, a large body of evidence suggests SG formation in hosts is a widespread response to microbial infection. Diverse stresses caused by microbes and their products can activate the integrated stress response in order to inhibit translation initiation through phosphorylation of the eukaryotic translation initiation factor 2α (eIF2α). This translational response in other contexts results in SG assembly, suggesting that SG assembly can be a general phenomenon during microbial infection. This review explores evidence for host SG formation in response to bacterial, fungal, and protozoan infection and potential functions of SGs in the host and for adaptations of the pathogen.
... On the other side, C. trachomatis infects and develops inside white blood cells, such as macrophages (24,25) or dendritic cells (DCs) (26)(27)(28)(29). DCs are the most potent antigen-presenting cells of the immune system and tightly regulate their endocytic system to process and present antigens efficiently onto MHC molecules (30). ...
During cross-presentation, exogenous antigens (i.e. intracellular pathogens or tumor cells) are internalized and processed within the endocytic system and also by the proteasome in the cytosol. Then, antigenic peptides are associated with Major Histocompatibility Complex (MHC) class I molecules and these complexes transit to the plasma membrane in order to trigger cytotoxic immune responses through the activation of CD8+ T lymphocytes. Dendritic cells (DCs) are particularly adapted to achieve efficient antigen cross-presentation and their endocytic network displays important roles during this process, including a sophisticated MHC-I transport dependent on recycling compartments. In this study, we show that C. trachomatis, an obligate intracellular pathogen that exhibits multiple strategies to evade the immune system, is able to induce productive infections in the murine DC line JAWS-II. Our results show that when C. trachomatis infects these cells, the bacteria-containing vacuole strongly recruits host cell recycling vesicles, but no other endosomal compartments. Furthermore, we found that chlamydial infection causes significant alterations of MHC-I trafficking in JAWS-II DCs: reduced levels of MHC-I expression at the cell surface, disruption of the perinuclear MHC-I intracellular pool, and impairment of MHC-I endocytic recycling to the plasma membrane. We observed that all these modifications lead to a hampered cross-presentation ability of soluble and particulate antigens by JAWS-II DCs and primary bone marrow-derived DCs. In summary, our findings provide substantial evidence that C. trachomatis hijacks the DC endocytic recycling system, causing detrimental changes on MHC-I intracellular transport, which are relevant for competent antigen cross-presentation.
