Translational arrest and mRNA decay are independent activities of alphaherpesvirus virion host shutoff proteins.

The herpes simplex virus 1 (HSV1) virion host shutoff (vhs) protein is an endoribonuclease that regulates the translational environment of the infected cell, by inducing the degradation of host mRNA via cellular exonuclease activity. To further understand the relationship between translational shutoff and mRNA decay, we have used ectopic expression to compare HSV1 vhs (vhsH) to its homologues from four other alphaherpesviruses - varicella zoster virus (vhsV), bovine herpesvirus 1 (vhsB), equine herpesvirus 1 (vhsE) and Marek's disease virus (vhsM). Only vhsH, vhsB and vhsE induced degradation of a reporter luciferase mRNA, with poly(A)+ in situ hybridization indicating a global depletion of cytoplasmic poly(A)+ RNA and a concomitant increase in nuclear poly(A)+ RNA and the polyA tail binding protein PABPC1 in cells expressing these variants. By contrast, vhsV and vhsM failed to induce reporter mRNA decay and poly(A)+ depletion, but rather, induced cytoplasmic G3BP1 and poly(A)+ mRNA- containing granules and phosphorylation of the stress response proteins eIF2α and protein kinase R. Intriguingly, regardless of their apparent endoribonuclease activity, all vhs homologues induced an equivalent general blockade to translation as measured by single-cell puromycin incorporation. Taken together, these data suggest that the activities of translational arrest and mRNA decay induced by vhs are separable and we propose that they represent sequential steps of the vhs host interaction pathway.

Peste des petits ruminants virus infection induces endoplasmic reticulum stress and apoptosis via IRE1-XBP1 and IRE1-JNK signaling pathways.

BACKGROUND: Peste des petits ruminants (PPR) is a contagious and fatal disease of sheep and goats. PPR virus (PPRV) infection induces endoplasmic reticulum (ER) stress-mediated unfolded protein response (UPR). The activation of UPR signaling pathways and their impact on apoptosis and virus replication remains controversial. OBJECTIVES: To investigate the role of PPRV-induced ER stress and the IRE1-XBP1 and IRE1-JNK pathways and their impact on apoptosis and virus replication. METHODS: The cell viability and virus replication were assessed by 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay, immunofluorescence assay, and Western blot. The expression of ER stress biomarker GRP78, IRE1, and its downstream molecules, PPRV-N protein, and apoptosis-related proteins was detected by Western blot and quantitative reverse transcription-polymerase chain reaction, respectively. 4-Phenylbutyric acid (4-PBA) and STF-083010 were respectively used to inhibit ER stress and IRE1 signaling pathway. RESULTS: The expression of GRP78, IRE1α, p-IRE1α, XBP1s, JNK, p-JNK, caspase-3, caspase-9, Bax and PPRV-N were significantly up-regulated in PPRV-infected cells, the expression of Bcl-2 was significantly down-regulated. Due to 4-PBA treatment, the expression of GRP78, p-IRE1α, XBP1s, p-JNK, caspase-3, caspase-9, Bax, and PPRV-N were significantly down-regulated, the expression of Bcl-2 was significantly up-regulated. Moreover, in PPRV-infected cells, the expression of p-IRE1α, p-JNK, Bax, and PPRV-N was significantly decreased, and the expression of Bcl-2 was increased in the presence of STF-083010. CONCLUSIONS: PPRV infection induces ER stress and IRE1 activation, resulting in apoptosis and enhancement of virus replication through IRE1-XBP1s and IRE1-JNK pathways.

IRE1α recognizes a structural motif in cholera toxin to activate an unfolded protein response.

IRE1α is an endoplasmic reticulum (ER) sensor that recognizes misfolded proteins to induce the unfolded protein response (UPR). We studied cholera toxin (CTx), which invades the ER and activates IRE1α in host cells, to understand how unfolded proteins are recognized. Proximity labeling colocalized the enzymatic and metastable A1 segment of CTx (CTxA1) with IRE1α in live cells, where we also found that CTx-induced IRE1α activation enhanced toxicity. In vitro, CTxA1 bound the IRE1α lumenal domain (IRE1αLD), but global unfolding was not required. Rather, the IRE1αLD recognized a seven-residue motif within an edge ß-strand of CTxA1 that must locally unfold for binding. Binding mapped to a pocket on IRE1αLD normally occupied by a segment of the IRE1α C-terminal flexible loop implicated in IRE1α oligomerization. Mutation of the CTxA1 recognition motif blocked CTx-induced IRE1α activation in live cells, thus linking the binding event with IRE1α signal transduction and induction of the UPR.

Inhibition of signaling protein ERN1 increases the sensitivity of serine synthesis gene expressions to glucose and glutamine deprivations in U87MG glioblastoma cells.

Objective. Glucose and glutamine supply as well as serine synthesis and endoplasmic reticulum (ER) stress are important factors of glioblastoma growth. Previous studies showed that the knockdown of ERN1 (ER to nucleus signaling 1) suppressed glioblastoma cell proliferation and modified the sensitivity of numerous gene expressions to nutrient deprivations. The present study is aimed to investigate the impact of glucose and glutamine deprivations on the expression of serine synthesis genes in U87MG glioblastoma cells in relation to ERN1 knockdown with the intent to reveal the role of ERN1 signaling pathway on the ER stress-dependent regulation of these gene expressions. Clarification of the regulatory mechanisms of serine synthesis is a great significance for glioblastoma therapy. Methods. The control U87MG glioblastoma cells (transfected by empty vector) and ERN1 knockdown cells (transfected by dominant-negative ERN1) were exposed under glucose and glutamine deprivation conditions for 16 h. RNA was extracted from cells and reverse transcribed. The expression level of PHGDH (phosphoglycerate dehydrogenase), PSAT1 (phosphoserine amino-transferase 1), PSPH (phosphoserine phosphatase), ATF4 (activating transcription factor 4), and SHMT1 (serine hydroxymethyltransferase 1) genes was studied by real-time qPCR and normalized to ACTB. Results. It was found that the expression level of genes responsible for serine synthesis such as PHGDH, PSAT1, PSPH, and transcription factor ATF4 was up-regulated in U87MG glioblastoma cells under glucose and glutamine deprivations. Furthermore, inhibition of ERN1 significantly enhances the impact of glucose and especially glutamine deprivations on these gene expressions. At the same time, the expression of the SHMT1 gene, which is responsible for serine conversion to glycine, was down-regulated in both nutrient deprivation conditions with more significant changes in ERN1 knockdown glioblastoma cells. Conclusion. Taken together, the results of present study indicate that the expression of genes responsible for serine synthesis is sensitive to glucose and glutamine deprivations in gene-specific manner and that suppression of ERN1 signaling significantly modifies the impact of both glucose and glutamine deprivations on PHGDH, PSAT1, PSPH, ATF4, and SHMT1 gene expressions and reflects the ERN1-mediated genome reprograming introduced by nutrient deprivation condition.

SARS-CoV-2 nsp15 endoribonuclease antagonizes dsRNA-induced antiviral signaling.

Severe acute respiratory syndrome coronavirus (SARS-CoV)-2 has caused millions of deaths since its emergence in 2019. Innate immune antagonism by lethal CoVs such as SARS-CoV-2 is crucial for optimal replication and pathogenesis. The conserved nonstructural protein 15 (nsp15) endoribonuclease (EndoU) limits activation of double-stranded (ds)RNA-induced pathways, including interferon (IFN) signaling, protein kinase R (PKR), and oligoadenylate synthetase/ribonuclease L (OAS/RNase L) during diverse CoV infections including murine coronavirus and Middle East respiratory syndrome (MERS)-CoV. To determine how nsp15 functions during SARS-CoV-2 infection, we constructed a recombinant SARS-CoV-2 (nsp15mut) expressing catalytically inactivated nsp15, which we show promoted increased dsRNA accumulation. Infection with SARS-CoV-2 nsp15mut led to increased activation of the IFN signaling and PKR pathways in lung-derived epithelial cell lines and primary nasal epithelial air-liquid interface (ALI) cultures as well as significant attenuation of replication in ALI cultures compared to wild-type virus. This replication defect was rescued when IFN signaling was inhibited with the Janus activated kinase (JAK) inhibitor ruxolitinib. Finally, to assess nsp15 function in the context of minimal (MERS-CoV) or moderate (SARS-CoV-2) innate immune induction, we compared infections with SARS-CoV-2 nsp15mut and previously described MERS-CoV nsp15 mutants. Inactivation of nsp15 had a more dramatic impact on MERS-CoV replication than SARS-CoV-2 in both Calu3 cells and nasal ALI cultures suggesting that SARS-CoV-2 can better tolerate innate immune responses. Taken together, SARS-CoV-2 nsp15 is a potent inhibitor of dsRNA-induced innate immune response and its antagonism of IFN signaling is necessary for optimal viral replication in primary nasal ALI cultures.

Gα12 and endoplasmic reticulum stress-mediated pyroptosis in a single cycle of dextran sulfate-induced mouse colitis.

Inflammatory bowel disease (IBD) pathogenesis involves complex inflammatory events and cell death. Although IBD involves mainly necrosis in the digestive tract, pyroptosis has also been recognized. Nonetheless, the underlying basis is elusive. Gα12/13 overexpression may affect endoplasmic reticulum (ER) stress. This study examined how Gα12/13 and ER stress affect pyroptosis using dextran sulfate sodium (DSS)-induced colitis models. Gα12/13 levels were increased in the distal and proximal colons of mice exposed to a single cycle of DSS, as accompanied by increases of IRE1α, ATF6, and p-PERK. Moreover, Il-6, Il-1ß, Ym1, and Arg1 mRNA levels were increased with caspase-1 and IL-1ß activation, supportive of pyroptosis. In the distal colon, RIPK1/3 levels were enhanced to a greater degree, confirming necroptosis. By contrast, the mice subjected to three cycles of DSS treatments showed decreases of Gα12/13, as accompanied by IRE1α and ATF6 suppression, but increases of RIPK1/3 and c-Cas3. AZ2 treatment, which inhibited Gα12, has an anti-pyroptotic effect against a single cycle of colitis. These results show that a single cycle of DSS-induced colitis may cause ER stress-induced pyroptosis as mediated by Gα12 overexpression in addition to necroptosis, but three cycles model induces only necroptosis, and that AZ2 may have an anti-pyroptotic effect.

HSP47 Increases the Expression of Type I Collagen in Fibroblasts through IRE1α Activation, XBP1 Splicing, and Nuclear Translocation of ß-Catenin.

Heat shock protein 47 (HSP47), also known as SERPINH1, functions as a collagen-specific molecular chaperone protein essential for the formation and stabilization of the collagen triple helix. Here, we delved into the regulatory pathways governed by HSP47, shedding light on collagen homeostasis. Our investigation revealed a significant reduction in HSP47 mRNA levels in the skin tissue of older mice as compared to their younger counterparts. The augmented expression of HSP47 employing lentivirus infection in fibroblasts resulted in an increased secretion of type I collagen. Intriguingly, the elevated expression of HSP47 in fibroblasts correlated with increased protein and mRNA levels of type I collagen. The exposure of fibroblasts to IRE1α RNase inhibitors resulted in the reduced manifestation of HSP47-induced type I collagen secretion and expression. Notably, HSP47-overexpressing fibroblasts exhibited increased XBP1 mRNA splicing. The overexpression of HSP47 or spliced XBP1 facilitated the nuclear translocation of ß-catenin and transactivated a reporter harboring TCF binding sites on the promoter. Furthermore, the overexpression of HSP47 or spliced XBP1 or the augmentation of nuclear ß-catenin through Wnt3a induced the expression of type I collagen. Our findings substantiate that HSP47 enhances type I collagen expression and secretion in fibroblasts by orchestrating a mechanism that involves an increase in nuclear ß-catenin through IRE1α activation and XBP1 splicing. This study therefore presents potential avenues for an anti-skin-aging strategy targeting HSP47-mediated processes.

The role of the 5' sensing function of ribonuclease E in cyanobacteria.

RNA degradation is critical for synchronising gene expression with changing conditions in prokaryotic and eukaryotic organisms. In bacteria, the preference of the central ribonucleases RNase E, RNase J and RNase Y for 5'-monophosphorylated RNAs is considered important for RNA degradation. For RNase E, the underlying mechanism is termed 5' sensing, contrasting to the alternative 'direct entry' mode, which is independent of monophosphorylated 5' ends. Cyanobacteria, such as Synechocystis sp. PCC 6803 (Synechocystis), encode RNase E and RNase J homologues. Here, we constructed a Synechocystis strain lacking the 5' sensing function of RNase E and mapped on a transcriptome-wide level 283 5'-sensing-dependent cleavage sites. These included so far unknown targets such as mRNAs encoding proteins related to energy metabolism and carbon fixation. The 5' sensing function of cyanobacterial RNase E is important for the maturation of rRNA and several tRNAs, including tRNAGluUUC. This tRNA activates glutamate for tetrapyrrole biosynthesis in plant chloroplasts and in most prokaryotes. Furthermore, we found that increased RNase activities lead to a higher copy number of the major Synechocystis plasmids pSYSA and pSYSM. These results provide a first step towards understanding the importance of the different target mechanisms of RNase E outside Escherichia coli.

Seasonal changes in hepatic lipid metabolism and apoptosis in Chinese soft-shelled turtle (Pelodiscus sinensis).

Chinese soft-shelled turtle (Pelodiscus sinensis) hibernates without eating and drinking when the ambient temperature is very low. To better understand the characteristics of energy utilization during hibernation, the turtles in the physiological phases of summer active (SA), Pre-Hibernation (Pre-H), Mid-Hibernation (Mid-H) and early arousal (EA) were sampled. The results showed that the levels of serum triglyceride and hepatic lipid droplet were markedly increased in Pre-H and decreased in Mid-H compared with that in SA, indicating that P. sinensis experiences lipid accumulation in Pre-H and lipid is the predominant energy reserve during hibernation. The mRNA expression levels of genes (FABP and CPT-2) involved in lipolysis and lipid oxidation were up-regulated in Mid-H, while the genes related to lipid synthesis (FAS, ACSL-1, ACC, elovl5, and SCD1) were inhibited in Mid-H. Meanwhile, the mRNA expression levels of endoplasmic reticulum stress marker gene Bip and key genes (ATF4, ATF6, and IRE1α) involving the unfolded protein response were significantly increased in Mid-H and EA. Also, the expression levels of genes (ASK1, JNK1, and Bax) associated with cell apoptosis increased in Mid-H and EA, however, the expression of Bcl2 was inhibited in Mid-H. Therefore, hibernation can cause endoplasmic reticulum stress and apoptosis. The findings will provide a theoretical framework for an animal's cold adaptation and offer insights into preventing and managing metabolic syndrome.

Prenatal inflammation exposure accelerates lung cancer tumorigenesis in offspring mouse: possible links to IRE1α/XBP1-mediated M2-like polarization of TAMs and PD-L1 up-expression.

BACKGROUND: Prenatal inflammation exposure (PIE) can increase the disease susceptibility in offspring such as lung cancer. Our purpose was to investigate the mechanisms of PIE on lung cancer. METHODS: Prenatal BALB/c mice were exposed to lipopolysaccharide (LPS), and then, their offspring were intraperitoneally instilled with urethane to establish the two-stage lung cancer carcinogenesis model. At the 48 weeks of age, the offspring mice were killed and lung tissues were collected for HE, immunohistochemistry, immunofluorescence, and Luminex MAGPIX®-based assays. CD11b + F4/80 + tumor-associated macrophages (TAMs) were sorted out from lung tumor tissues by cell sorting technique. Flow cytometry was employed to evaluate the extent of M2-like polarization of TAMs and PD-L1 expression. RESULTS: The offspring of PIE mice revealed more lung lesion changes, including atypical hyperplasia and intrapulmonary metastases. The number of lung nodules, lung organ index, and PCNA, MMP-9 and Vimentin positive cells in lung tissue of PIE group were higher than those of Control group. The increases of mRNA encoding M2 macrophage markers and cytokines in offspring of prenatal LPS-treated mice confirmed the induced effect of PIE on macrophage polarization. Additionally, PIE treatment increased the percentage of CD163 + CD206 + cells in the sorted TAMs. Importantly, endoplasmic reticulum (ER) stress-markers like GRP78/BIP and CHOP, p-IRE1α and XBP1s, and PD-L1 were up-regulated in TAMs from PIE group. Besides, we also observed that IRE1α inhibitor (KIRA6) reversed the M2-like TAMs polarization and metastasis induced by PIE. CONCLUSIONS: IRE1α/XBP1-mediated M2-like TAMs polarization releases the pro-tumorigenic cytokines and PD-L1 expression, which may be the regulatory mechanism of accelerating lung cancer in offspring of mice undergoing PIE.

Unraveling the Role of RNase L Knockout in Alleviating Immune Response Activation in Mice Bone Marrow after Irradiation.

Ionizing radiation (IR) induces severe hematopoietic injury by causing DNA and RNA damage as well as activating the immune responses, necessitating the development of effective therapeutic strategies. Ribonuclease L (RNase L) as an innate immune response pathway is triggered by exogenous and endogenous abnormal dsRNA under viral infection and dyshomeostasis, thereby activating the immune responses. Thus, we investigated the effect of RNase L on irradiation-induced bone marrow damage using RNase L knockout (RNase L-/-) mice. Phenotypic analysis revealed that RNase L knockout mitigates irradiation-induced injury in the bone marrow. Further investigation into the mechanism of RNase L by RNA-seq, qRT-PCR, and CBA analysis demonstrated that RNase L deficiency counteracts the upregulation of genes related to immune responses induced by irradiation, including cytokines and interferon-stimulated genes. Moreover, RNase L deficiency inhibits the increased levels of immunoglobulins in serum induced by irradiation. These findings indicate that RNase L plays a role in the immune response induced by irradiation in the bone marrow. This study further enhances our understanding of the biological functions of RNase L in the immune response induced by irradiation and offers a novel approach for managing irradiation-induced bone marrow injury through the regulation of RNase L activation.

Lack of the Histone Deacetylase SIRT1 Leads to Protection against Endoplasmic Reticulum Stress through the Upregulation of Heat Shock Proteins.

Histone deacetylase SIRT1 represses gene expression through the deacetylation of histones and transcription factors and is involved in the protective cell response to stress and aging. However, upon endoplasmic reticulum (ER) stress, SIRT1 impairs the IRE1α branch of the unfolded protein response (UPR) through the inhibition of the transcriptional activity of XBP-1 and SIRT1 deficiency is beneficial under these conditions. We hypothesized that SIRT1 deficiency may unlock the blockade of transcription factors unrelated to the UPR promoting the synthesis of chaperones and improving the stability of immature proteins or triggering the clearance of unfolded proteins. SIRT1+/+ and SIRT1-/- fibroblasts were exposed to the ER stress inducer tunicamycin and cell survival and expression of heat shock proteins were analyzed 24 h after the treatment. We observed that SIRT1 loss significantly reduced cell sensitivity to ER stress and showed that SIRT1-/- but not SIRT1+/+ cells constitutively expressed high levels of phospho-STAT3 and heat shock proteins. Hsp70 silencing in SIRT1-/- cells abolished the resistance to ER stress. Furthermore, accumulation of ubiquitinated proteins was lower in SIRT1-/- than in SIRT1+/+ cells. Our data showed that SIRT1 deficiency enabled chaperones upregulation and boosted the proteasome activity, two processes that are beneficial for coping with ER stress.

Coevolution of RNA and protein subunits in RNase P and RNase MRP, two RNA processing enzymes.

RNase P and RNase mitochondrial RNA processing (MRP) are ribonucleoproteins (RNPs) that consist of a catalytic RNA and a varying number of protein cofactors. RNase P is responsible for precursor tRNA maturation in all three domains of life, while RNase MRP, exclusive to eukaryotes, primarily functions in rRNA biogenesis. While eukaryotic RNase P is associated with more protein cofactors and has an RNA subunit with fewer auxiliary structural elements compared to its bacterial cousin, the double-anchor precursor tRNA recognition mechanism has remarkably been preserved during evolution. RNase MRP shares evolutionary and structural similarities with RNase P, preserving the catalytic core within the RNA moiety inherited from their common ancestor. By incorporating new protein cofactors and RNA elements, RNase MRP has established itself as a distinct RNP capable of processing ssRNA substrates. The structural information on RNase P and MRP helps build an evolutionary trajectory, depicting how emerging protein cofactors harmonize with the evolution of RNA to shape different functions for RNase P and MRP. Here, we outline the structural and functional relationship between RNase P and MRP to illustrate the coevolution of RNA and protein cofactors, a key driver for the extant, diverse RNP world.

Endoribonuclease DNE1 Promotes Ethylene Response by Modulating EBF1/2 mRNA Processing in Arabidopsis.

The gaseous phytohormone ethylene plays a crucial role in plant growth, development, and stress responses. In the ethylene signal transduction cascade, the F-box proteins EIN3-BINDING F-BOX 1 (EBF1) and EBF2 are identified as key negative regulators governing ethylene sensitivity. The translation and processing of EBF1/2 mRNAs are tightly controlled, and their 3' untranslated regions (UTRs) are critical in these regulations. However, despite their significance, the exact mechanisms modulating the processing of EBF1/2 mRNAs remain poorly understood. In this work, we identified the gene DCP1-ASSOCIATED NYN ENDORIBONUCLEASE 1 (DNE1), which encodes an endoribonuclease and is induced by ethylene treatment, as a positive regulator of ethylene response. The loss of function mutant dne1-2 showed mild ethylene insensitivity, highlighting the importance of DNE1 in ethylene signaling. We also found that DNE1 colocalizes with ETHYLENE INSENSITIVE 2 (EIN2), the core factor manipulating the translation of EBF1/2, and targets the P-body in response to ethylene. Further analysis revealed that DNE1 negatively regulates the abundance of EBF1/2 mRNAs by recognizing and cleaving their 3'UTRs, and it also represses their translation. Moreover, the dne1 mutant displays hypersensitivity to 1,4-dithiothreitol (DTT)-induced ER stress and oxidative stress, indicating the function of DNE1 in stress responses. This study sheds light on the essential role of DNE1 as a modulator of ethylene signaling through regulation of EBF1/2 mRNA processing. Our findings contribute to the understanding of the intricate regulatory process of ethylene signaling and provide insights into the significance of ribonuclease in stress responses.

MazEF Homologs in Symbiobacterium thermophilum Exhibit Cross-Neutralization with Non-Cognate MazEFs.

Toxin-antitoxin systems are preserved by nearly every prokaryote. The type II toxin MazF acts as a sequence-specific endoribonuclease, cleaving ribonucleotides at specific sequences that vary from three to seven bases, as has been reported in different host organisms to date. The present study characterized the MazEF module (MazEF-sth) conserved in the Symbiobacterium thermophilum IAM14863 strain, a Gram-negative syntrophic bacterium that can be supported by co-culture with multiple bacteria, including Bacillus subtilis. Based on a method combining massive parallel sequencing and the fluorometric assay, MazF-sth was determined to cleave ribonucleotides at the UACAUA motif, which is markedly similar to the motifs recognized by MazF from B. subtilis (MazF-bs), and by several MazFs from Gram-positive bacteria. MazF-sth, with mutations at conserved amino acid residues Arg29 and Thr52, lost most ribonuclease activity, indicating that these residues that are crucial for MazF-bs also play significant roles in MazF-sth catalysis. Further, cross-neutralization between MazF-sth and the non-cognate MazE-bs was discovered, and herein, the neutralization mechanism is discussed based on a protein-structure simulation via AlphaFold2 and multiple sequence alignment. The conflict between the high homology shared by these MazF amino acid sequences and the few genetic correlations among their host organisms may provide evidence of horizontal gene transfer.

Licochalcone A induces endoplasmic reticulum stress-mediated apoptosis of endometrial cancer cells via upregulation of GRP78 expression.

Licochalcone A (LicA), a natural compound extracted from licorice root, has been shown to exert a variety of anticancer activities. Whether LicA has such effects on endometrial cancer (EMC) is unclear. This study aims to investigate the antitumor effects of LicA on EMC. Our results show that LicA significantly reduced the viability and induced apoptosis of EMC cells and EMC-7 cells from EMC patients. LicA was also found to induce endoplasmic reticulum (ER) stress, leading to increased expression of ER-related proteins (GRP78/PERK/IRE1α/CHOP) in EMC cell lines. Suppression of GRP78 expression in human EMC cells treated with LicA significantly attenuated the effects of LicA, resulting in reduced ER-stress mediated cell apoptosis and decreased expression of ER- and apoptosis-related proteins. Our findings demonstrate that LicA induces apoptosis in EMC cells through the GRP78-mediated ER-stress pathway, emphasizing the potential of LicA as an anticancer therapy for EMC.

Endoplasmic reticulum stress responses and epigenetic alterations in arsenic carcinogenesis.

Arsenic is a well-known human carcinogen whose environmental exposure via drinking water, food, and air impacts millions of people across the globe. Various mechanisms of arsenic carcinogenesis have been identified, ranging from damage caused by excessive production of free radicals and epigenetic alterations to the generation of cancer stem cells. A growing body of evidence supports the critical involvement of the endoplasmic stress-activated unfolded protein response (UPR) in promoting as well as suppressing cancer development/progression. Various in vitro and in vivo models have also demonstrated that arsenic induces the UPR via activation of the PERK, IRE1α, and ATF6 proteins. In this review, we discuss the mechanisms of arsenic-induced endoplasmic reticulum stress and the role of each UPR pathway in the various cancer types with a focus on the epigenetic regulation and function of the ATF6 protein. The importance of UPR in arsenic carcinogenesis and cancer stem cells is a relatively new area of research that requires additional investigations via various omics-based and computational tools. These approaches will provide interesting insights into the mechanisms of arsenic-induced cancers for prospective target identification and development of novel anti-cancer therapies.

IRE1 RNase controls CD95-mediated cell death.

Signalling by the Unfolded Protein Response (UPR) or by the Death Receptors (DR) are frequently activated towards pro-tumoral outputs in cancer. Herein, we demonstrate that the UPR sensor IRE1 controls the expression of the DR CD95/Fas, and its cell death-inducing ability. Both genetic and pharmacologic blunting of IRE1 activity increased CD95 expression and exacerbated CD95L-induced cell death in glioblastoma (GB) and Triple-Negative Breast Cancer (TNBC) cell lines. In accordance, CD95 mRNA was identified as a target of Regulated IRE1-Dependent Decay of RNA (RIDD). Whilst CD95 expression is elevated in TNBC and GB human tumours exhibiting low RIDD activity, it is surprisingly lower in XBP1s-low human tumour samples. We show that IRE1 RNase inhibition limited CD95 expression and reduced CD95-mediated hepatic toxicity in mice. In addition, overexpression of XBP1s increased CD95 expression and sensitized GB and TNBC cells to CD95L-induced cell death. Overall, these results demonstrate the tight IRE1-mediated control of CD95-dependent cell death in a dual manner through both RIDD and XBP1s, and they identify a novel link between IRE1 and CD95 signalling.

RACK1 and IRE1 participate in the translational quality control of amyloid precursor protein in Drosophila models of Alzheimer's disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by dysregulation of the expression and processing of the amyloid precursor protein (APP). Protein quality control systems are dedicated to remove faulty and deleterious proteins to maintain cellular protein homeostasis (proteostasis). Identidying mechanisms underlying APP protein regulation is crucial for understanding AD pathogenesis. However, the factors and associated molecular mechanisms regulating APP protein quality control remain poorly defined. In this study, we show that mutant APP with its mitochondrial-targeting sequence ablated exhibited predominant endoplasmic reticulum (ER) distribution and led to aberrant ER morphology, deficits in locomotor activity, and shortened lifespan. We searched for regulators that could counteract the toxicity caused by the ectopic expression of this mutant APP. Genetic removal of the ribosome-associated quality control (RQC) factor RACK1 resulted in reduced levels of ectopically expressed mutant APP. By contrast, gain of RACK1 function increased mutant APP level. Additionally, overexpression of the ER stress regulator (IRE1) resulted in reduced levels of ectopically expressed mutant APP. Mechanistically, the RQC related ATPase VCP/p97 and the E3 ubiquitin ligase Hrd1 were required for the reduction of mutant APP level by IRE1. These factors also regulated the expression and toxicity of ectopically expressed wild type APP, supporting their relevance to APP biology. Our results reveal functions of RACK1 and IRE1 in regulating the quality control of APP homeostasis and mitigating its pathogenic effects, with implications for the understanding and treatment of AD.

Two Main Cancer Biomarkers as Molecular Targets of Binase Antitumor Activity.

Cancer is frequently coupled with the disturbance of key signaling pathways. Aberrant activation of the mitogen-activated protein kinase (MAPK) signaling cascade, occurring in over 85% of cancers, is mainly caused by the genetic alterations of its main components-oncogenes EGFR and RAS, and plays a crucial role in cell fate. The importance of EGFR and RAS proteins in a variety of tumors suggests that they would be good therapeutic targets, but at present, no effective targeted therapy against these two oncogenes has been proven. Here, we show that ribonuclease from Bacillus pumilus (binase) inhibits MAPK signaling through direct interaction with EGFR and RAS proteins. This effect contributes to the antitumor potential of binase along with its enzymatic activity. Multitargeticity of binase prevents the development of drug resistance, which is considered a major obstacle to effective anticancer treatment.