Bombyx mori UFL1 facilitates BmNPV proliferation by regulating host cell apoptosis through PERK.

Ubiquitin-fold modifier 1 (UFM1) is attached to protein substrates through the sequential activity of an E1 (UBA5)-E2 (UFC1)-E3 (UFL1) cascade. UFL1 is the E3 ligase for UFMylation in vertebrates. However, there have been no studies on UFL1 in silkworm to date. In this study, we identified a UFL1 ortholog in Bombyx mori genome. Spatio-temporal expression profiles showed that BmUFL1 expression was high in the midgut, epidermis, and testis and in the pupa-adult stage. BmUFL1 knockdown inhibited B. mori nucleopolyhedrovirus (BmNPV) proliferation, while BmUFL1 overexpression promoted BmNPV proliferation. Mechanically, protein kinase RNA-like endoplasmic reticulum kinase (PERK) signaling and cell apoptosis are involved in BmUFL1-regulated BmNPV proliferation. Overall, these results suggest that BmUFL1 facilitates BmNPV proliferation in silkworm.

[Research Advances in the Association Between Alzheimer's Disease and Double-Stranded RNA-Dependent Protein Kinase].

Alzheimer's disease (AD) is a severe threat to human health and one of the three major causes of human death.Double-stranded RNA-dependent protein kinase (PKR) is an interferon-induced protein kinase involved in innate immunity.In the occurrence and development of AD,PKR is upregulated and continuously activated.On the one hand,the activation of PKR triggers an integrated stress response in brain cells.On the other hand,it indirectly upregulates the expression of ß-site amyloid precursor protein cleaving enzyme 1 and facilitates the accumulation of amyloid-ß protein (Aß),which could activate PKR activator to further activate PKR,thus forming a sustained accumulation cycle of Aß.In addition,PKR can promote Tau phosphorylation,thereby reducing microtubule stability in nerve cells.Inflammation in brain tissue,neurotoxicity resulted from Aß accumulation,and disruption of microtubule stability led to the progression of AD and the declines of memory and cognitive function.Therefore,PKR is a key molecule in the development and progression of AD.Effective PKR detection can aid in the diagnosis and prediction of AD progression and provide opportunities for clinical treatment.The inhibitors targeting PKR are expected to control the activity of PKR,thereby controlling the progression of AD.Therefore,PKR could be a target for the development of therapeutic drugs for AD.

Qing Hua Chang Yin ameliorates chronic colitis in mice by inhibiting PERK-ATF4-CHOP pathway of ER stress and the NF-κB signalling pathway.

CONTEXT: Ulcerative colitis has been clinically treated with Qing Hua Chang Yin (QHCY), a traditional Chinese medicine formula. However, its precise mechanisms in mitigating chronic colitis are largely uncharted. OBJECTIVE: To elucidate the therapeutic efficiency of QHCY on chronic colitis and explore its underlying molecular mechanisms. MATERIALS AND METHODS: A total ion chromatogram fingerprint of QHCY was analysed. Chronic colitis was induced in male C57BL/6 mice using 2% dextran sodium sulphate (DSS) over 49 days. Mice were divided into control, DSS, DSS + QHCY (0.8, 1.6 and 3.2 g/kg/d dose, respectively) and DSS + mesalazine (0.2 g/kg/d) groups (n = 6). Mice were intragastrically administered QHCY or mesalazine for 49 days. The changes of disease activity index (DAI), colon length, colon histomorphology and serum pro-inflammatory factors in mice were observed. RNA sequencing was utilized to identify the differentially expressed transcripts (DETs) in colonic tissues and the associated signalling pathways. The expression of endoplasmic reticulum (ER) stress-related protein and NF-κB signalling pathway-related proteins in colonic tissues was detected by immunohistochemistry staining. RESULTS: Forty-seven compounds were identified in QHCY. Compared with the DSS group, QHCY significantly improved symptoms of chronic colitis like DAI increase, weight loss, colon shortening and histological damage. It notably reduced serum levels of IL-6, IL-1ß and TNF-α. QHCY suppressed the activation of PERK-ATF4-CHOP pathway of ER stress and NF-κB signalling pathways in colonic tissues. DISCUSSION AND CONCLUSIONS: The findings in this study provide novel insights into the potential of QHCY in treating chronic colitis patients.

Induction of the PERK-eIF2α-ATF4 Pathway in M1 Macrophages under Endoplasmic Reticulum Stress.

Translation inhibition can activate two cell death pathways. The first pathway is activated by translational aberrations, the second by endoplasmic reticulum (ER) stress. In this work, the effect of ribosome-inactivating protein type II (RIP-II) viscumin on M1 macrophages derived from the THP-1 cell line was investigated. The number of modified ribosomes was evaluated by real-time PCR. Transcriptome analysis revealed that viscumin induces the ER stress activated by the PERK sensor.

Differential effects of montelukast and zafirlukast on MDA­MB­231 triple­negative breast cancer cells: Cell cycle regulation, apoptosis, autophagy, DNA damage and endoplasmic reticulum stress.

Montelukast and zafirlukast, cysteinyl leukotriene receptor antagonists (LTRAs), trigger apoptosis and inhibit cell proliferation of triple­negative breast cancer MDA­MB­231 cells. By contrast, only zafirlukast induces G0/G1 cell cycle arrest. The present study compared the effects of these drugs on proteins regulating cell proliferation, apoptosis, autophagy, and endoplasmic reticulum (ER) and oxidative stress using reverse transcription­quantitative PCR, western blotting and flow cytometry. The expression of proliferating markers, Ki­67 and proliferating cell nuclear antigen, was decreased by both drugs. Zafirlukast, but not montelukast, decreased the expression of cyclin D1 and CDK4, disrupting progression from G1 to S phase. Zafirlukast also increased the expression of p27, a cell cycle inhibitor. Both drugs decreased the expression of anti­apoptotic protein Bcl­2 and ERK1/2 phosphorylation, and increased levels of the autophagy marker LC3­II and DNA damage markers, including cleaved PARP­1, phosphorylated (p)­ATM and p­histone H2AX. The number of caspase 3/7­positive cells was greater in montelukast­treated cells compared with zafirlukast­treated cells. Montelukast induced higher levels of the ER stress marker CHOP compared with zafirlukast. Montelukast activated PERK, activating transcription factor 6 (ATF6) and inositol­requiring enzyme type 1 (IRE1) pathways, while zafirlukast only stimulated ATF6 and IRE1 pathways. GSK2606414, a PERK inhibitor, decreased apoptosis mediated by montelukast, but did not affect zafirlukast­induced cell death. The knockdown of CHOP by small interfering RNA reduced apoptosis triggered by montelukast and zafirlukast. In conclusion, the effects on cell cycle regulator proteins may contribute to cell cycle arrest caused by zafirlukast. The greater apoptotic effects of montelukast may be caused by the higher levels of activated caspase enzymes and the activation of three pathways of ER stress: PERK, ATF6, and IRE1.

Dysregulated RNA editing of EIF2AK2 in polycystic ovary syndrome: clinical relevance and functional implications.

BACKGROUND: Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder affecting women of reproductive ages. Our previous study has implicated a possible link between RNA editing and PCOS, yet the actual role of RNA editing, its association with clinical features, and the underlying mechanisms remain unclear. METHODS: Ten RNA-Seq datasets containing 269 samples of multiple tissue types, including granulosa cells, T helper cells, placenta, oocyte, endometrial stromal cells, endometrium, and adipose tissues, were retrieved from public databases. Peripheral blood samples were collected from twelve PCOS and ten controls and subjected to RNA-Seq. Transcriptome-wide RNA-Seq data analysis was conducted to identify differential RNA editing (DRE) between PCOS and controls. The functional significance of DRE was evaluated by luciferase reporter assays and overexpression in human HEK293T cells. Dehydroepiandrosterone and lipopolysaccharide were used to stimulate human KGN granulosa cells to evaluate gene expression. RESULTS: RNA editing dysregulations across multiple tissues were found to be associated with PCOS in public datasets. Peripheral blood transcriptome analysis revealed 798 DRE events associated with PCOS. Through weighted gene co-expression network analysis, our results revealed a set of hub DRE events in PCOS blood. A DRE event in the eukaryotic translation initiation factor 2-alpha kinase 2 (EIF2AK2:chr2:37,100,559) was associated with PCOS clinical features such as luteinizing hormone (LH) and the ratio of LH over follicle-stimulating hormone. Luciferase assays, overexpression, and knockout of RNA editing enzyme adenosine deaminase RNA specific (ADAR) showed that the ADAR-mediated editing cis-regulated EIF2AK2 expression. EIAF2AK2 showed a higher expression after dehydroepiandrosterone and lipopolysaccharide stimulation, triggering changes in the downstrean MAPK pathway. CONCLUSIONS: Our study presented the first evidence of cross-tissue RNA editing dysregulation in PCOS and its clinical associations. The dysregulation of RNA editing mediated by ADAR and the disrupted target EIF2AK2 may contribute to PCOS development via the MPAK pathway, underlining such epigenetic mechanisms in the disease.

Dihydropyrazine induces endoplasmic reticulum stress and inhibits autophagy in HepG2 human hepatoma cells.

Dihydropyrazines (DHPs) are formed by non-enzymatic glycation reactions in vivo and in food. We recently reported that 3-hydro-2,2,5,6-tetramethylpyrazine (DHP-3), which is a methyl-substituted DHP, caused severe oxidative stress and cytotoxicity. However, the molecular mechanisms underlying the cytotoxic pathways of the DHP response remain elusive. Because oxidative stress induces endoplasmic reticulum (ER) stress and autophagy, we investigated the ability of DHP-3 to modulate the ER stress and autophagy pathways. DHP-3 activated the ER stress pathway by increasing inositol-requiring enzyme 1 (IRE1) and PKR-like ER kinase (PERK) phosphorylation and transcription factor 6 (ATF6) expression. Moreover, DHP-3 increased the expression of activating transcription factor 4 (ATF4) and C/EBP homologous protein (CHOP), which are downstream targets of PERK. In addition, DHP-3 inhibited the autophagy pathway by increasing the accumulation of microtubule-associated protein 1 light chain 3 alpha-phosphatidylethanolamine conjugate (LC3-II) and p62/sequestosome 1 (p62), while decreasing autophagic flux. Taken together, these results indicate that DHP-3 activates the ER stress pathway and inhibits the autophagy pathway, suggesting that the resulting removal of damaged organelles is inadequate.

Polystyrene Nanoplastics Induce Lipid Metabolism Disorder by Activating the PERK-ATF4 Signaling Pathway in Mice.

In recent years, the study of microplastics (MPs) and nanoplastics (NPs) and their effects on human health has gained significant attention. The impacts of NPs on lipid metabolism and the specific mechanisms involved remain poorly understood. To address this, we utilized high-throughput sequencing and molecular biology techniques to investigate how endoplasmic reticulum (ER) stress might affect hepatic lipid metabolism in the presence of polystyrene nanoplastics (PS-NPs). Our findings suggest that PS-NPs activate the PERK-ATF4 signaling pathway, which in turn upregulates the expression of genes related to lipid synthesis via the ATF4-PPARγ/SREBP-1 pathway. This activation leads to an abnormal accumulation of lipid droplets in the liver. 4-PBA, a known ER stress inhibitor, was found to mitigate the PS-NPs-induced lipid metabolism disorder. These results demonstrate the hepatotoxic effects of PS-NPs and clarify the mechanisms of abnormal lipid metabolism induced by PS-NPs.

Identification and Validation of PKR as a Direct Target for the Novel Sulfonamide-Substituted Tetrahydroquinoline Nonselective Inhibitor of the NLRP3 Inflammasome.

The NLRP3 inflammasome is a critical component of the innate immune system. The persistent abnormal activation of the NLRP3 inflammasome is implicated in numerous human diseases. Herein, sulfonamide-substituted tetrahydroquinoline derivative S-9 was identified as the most promising NLRP3 inhibitor, without obvious cytotoxicity. In vitro, S-9 inhibited the priming and activation stages of the NLRP3 inflammasome. Incidentally, we also observed that S-9 had inhibitory effects on the NLRC4 and AIM2 inflammasomes. To elucidate the multiple anti-inflammatory activities of S-9, photoaffinity probe P-2, which contained a photoaffinity label and a functional handle, was developed for target identification by chemical proteomics. We identified PKR as a novel target of S-9 in addition to NLRP3 by target fishing. Furthermore, S-9 exhibited a significant anti-neuroinflammatory effect in vivo. In summary, our findings show that S-9 is a promising lead compound targeting both PKR and NLRP3 that could emerge as a molecular tool for treating inflammasome-related diseases.

Knockdown of EIF2AK2-OAS1 axis reduces ATP production inducing AMPK phosphorylation to inhibit the malignant behavior of gastric cancer cells.

Energy metabolism has always been a hot topic in cancer progression and targeted therapy, and exploring the role of genes in energy metabolic pathways in cancer cells has become key to address this issue. Eukaryotic translation initiation factor 2α kinase 2 (EIF2AK2) plays regulatory roles in cancer and disorders of energy metabolism. Indeed, the role of EIF2AK2 in energy metabolism has been underestimated. The aim of this study is to reveal the expression specificity of EIF2AK2 in gastric cancer (GC) progression and metastasis, and to demonstrate the role of EIF2AK2 in energy metabolism, cytoskeleton, proliferation, death and metastasis pathways in GC cells. Mechanistically, EIF2AK2 overexpression promoted cytoskeleton remodeling and ATP production, mediated cell proliferation and metastasis, upregulated OAS1 expression, decreases p-AMPK expression and inhibited apoptosis in GC cells. Conversely, knockdown of EIF2AK2 resulted in the opposite effect. However, overexpression of OAS1 mediated the upregulation of mitochondrial membrane potential and promoted ATP production and NAD+/NADH ratio, but knockdown of OAS1 inhibited the above effects. In addition, knockdown of OAS1 had no effect on EIF2AK2 expression, but inhibited AMPK and upregulated p-AMPK expression. In conclusion, our study identified EIF2AK2 and OAS1 as previously undescribed regulators of energy metabolism in GC cells. We hypothesized that EIF2AK2-OAS1 axis may regulate energy metabolism and inhibit cellular malignant behavior in cancer cells by affecting ATP production to induce AMPK phosphorylation, suggesting EIF2AK2 as a potential therapeutic target for cancer cell progression.

p20BAP31 promotes cell apoptosis via interaction with GRP78 and activating the PERK pathway in colorectal cancer.

Colorectal cancer (CRC) is the second most deadly cancer worldwide. Although various treatments for CRC have made progress, they have limitations. Therefore, the search for new effective molecular targets is important for the treatment of CRC. p20BAP31 induces apoptosis through diverse pathways and exhibits greater sensitivity in CRC. Therefore, a comprehensive exploration of the molecular functions of p20BAP31 is important for its application in anti-tumor therapy. In this study, we showed that exogenous p20BAP31 was still located in the ER and significantly activated the unfolded protein response (UPR) through the PERK pathway. The activation of the PERK pathway is prominent in p20BAP31-induced reactive oxygen species (ROS) accumulation and apoptosis. We found, for the first time, that p20BAP31 leads to ER stress and markedly attenuates tumor cell growth in vivo. Importantly, mechanistic investigations indicated that p20BAP31 competitively binds to GRP78 from PERK and causes hyperactivation of the UPR. Furthermore, p20BAP31 upregulates the expression of GRP78 by promoting HSF1 nuclear translocation and enhancing its binding to the GRP78 promoter. These findings reveal p20BAP31 as a regulator of ER stress and a potential target for tumor therapy, and elucidate the underlying mechanism by which p20BAP31 mediates signal transduction between ER and mitochondria.

Deciphering ER stress-unfolded protein response relationship by visualizing unfolded proteins in the ER.

Despite the consensus that accumulation of unfolded proteins in the endoplasmic reticulum (ER) lumen, i.e. ER stress, activates the unfolded protein response (UPR), studies under physiological and pathophysiological conditions suggest that ER stress may not always trigger the UPR, and the UPR can be activated in an ER stress-independent way. To better understand how the UPR is regulated and its relationship with ER stress requires direct detection of unfolded proteins in the ER, a method that is still lacking. Here, we report a strategy of visualizing unfolded protein accumulation in the ER lumen in living cells by employing an engineered ER stress sensor, PERK, which forms fluorescence puncta upon unfolded protein binding, in a fast and reversible way. Our reporter enables us to clarify the involvement of unfolded proteins in UPR activation under several physiological conditions and suggests that persistent unfolded protein accumulation in the ER despite UPR attenuation predicts cell death.

The impact of single-stranded RNAs on the dimerization of double-stranded RNA-dependent protein kinase PKR.

The RNA-binding protein PKR serves as a crucial antiviral innate immune factor that globally suppresses translation by sensing viral double-stranded RNA (dsRNA) and by phosphorylating the translation initiation factor eIF2α. Recent findings have unveiled that single-stranded RNAs (ssRNAs), including in vitro transcribed (IVT) mRNA, can also bind to and activate PKR. However, the precise mechanism underlying PKR activation by ssRNAs, remains incompletely understood. Here, we developed a NanoLuc Binary Technology (NanoBiT)-based in vitro PKR dimerization assay to assess the impact of ssRNAs on PKR dimerization. Our findings demonstrate that, akin to double-stranded polyinosinic:polycytidylic acid (polyIC), an encephalomyocarditis virus (EMCV) RNA, as well as NanoLuc luciferase (Nluc) mRNA, can induce PKR dimerization. Conversely, homopolymeric RNA lacking secondary structure fails to promote PKR dimerization, underscoring the significance of secondary structure in this process. Furthermore, adenovirus VA RNA 1, another ssRNA, impedes PKR dimerization by competing with Nluc mRNA. Additionally, we observed structured ssRNAs capable of forming G-quadruplexes induce PKR dimerization. Collectively, our results indicate that ssRNAs have the ability to either induce or inhibit PKR dimerization, thus representing potential targets for the development of antiviral and anti-inflammatory agents.

LC3A-mediated autophagy elicits PERK-eIF2α-ATF4 axis activation and mitochondrial dysfunction: Exposing vulnerability in aggresome-positive cancer cells.

The unfolded protein response pathways (UPR), autophagy, and compartmentalization of misfolded proteins into inclusion bodies are critical components of the protein quality control network. Among inclusion bodies, aggresomes are particularly intriguing due to their association with cellular survival, drug resistance, and aggresive cancer behavior. Aggresomes are molecular condensates formed when collapsed vimentin cages encircle misfolded proteins before final removal by autophagy. Yet significant gaps persist in the mechanisms governing aggresome formation and elimination in cancer cells. Understanding these mechanisms is crucial, especially considering the involvement of LC3A, a member of the MAP1LC3 family, which plays a unique role in autophagy regulation and has been reported to be epigenetically silenced in many cancers. Herein, we utilized the tetracycline-inducible expression of LC3A to investigate its role in choroid plexus carcinoma cells, which inherently exhibit the presence of aggresomes. Live cell imaging was employed to demonstrate the effect of LC3A expression on aggresome-positive cells, while SILAC-based proteomics identified LC3A-induced protein and pathway alterations. Our findings demonstrated that extended expression of LC3A is associated with cellular senescence. However, the obstruction of lysosomal degradation in this context has a deleterious effect on cellular viability. In response to LC3A-induced autophagy, we observed significant alterations in mitochondrial morphology, reflected by mitochondrial dysfunction and increased ROS production. Furthermore, LC3A expression elicited the activation of the PERK-eIF2α-ATF4 axis of the UPR, underscoring a significant change in the protein quality control network. In conclusion, our results elucidate that LC3A-mediated autophagy alters the protein quality control network, exposing a vulnerability in aggresome-positive cancer cells.

p20BAP31 Induces Autophagy in Colorectal Cancer Cells by Promoting PERK-Mediated ER Stress.

B-cell receptor-associated protein 31 (BAP31) is an endoplasmic reticulum (ER) membrane protein involved in apoptosis and autophagy by communication with ER and mitochondria. BAP31 is cleaved by caspase-8 and generates a proapoptotic fragment, p20BAP31, which has shown to induce ER stress and apoptosis through multiple pathways. In this study, we found that p20BAP31 significantly increased the agglomeration of LC3 puncta, suggesting the occurrence of autophagy. Therefore, it is meaningful to explore the mechanism of p20BAP31-induced autophagy, and further analyze the relationships among p20BAP31-induced autophagy, ER stress and apoptosis. The data showed that p20BAP31 induced autophagy by inhibition of the PI3K/AKT/mTOR signaling in colorectal cells. ER stress inhibitor 4-PBA and PERK siRNA alleviated p20BAP31-induced autophagy; in turn, autophagy inhibitors 3-MA and CQ did not affect p20BAP31-induced ER stress, suggesting that p20BAP31-induced ER stress is the upstream of autophagy. We also discovered that ROS inhibitor NAC inhibited p20BAP31-induced autophagy. Furthermore, inhibition of autophagy by CQ suppressed p20BAP31-induced apoptosis and ameliorated cell proliferation. Importantly, p20BAP31 markedly reduced the tumor size in vivo, and significantly enhanced the autophagy levels in the tumor tissues. Collectively, p20BAP31 initiates autophagy by inhibiting the PI3K/AKT/mTOR signaling and activating the PERK-mediated ROS accumulation, further promotes p20BAP31-induced apoptosis and ultimately results in cell death. This study comprehensively reveals the potential mechanism of p20BAP31-induced cell death, which may provide new strategies for antitumor therapy.

Suppression of PERK/eIF2α/CHOP pathway enhances oridonin-induced apoptosis by inhibiting autophagy in Small-Cell lung cancer cells.

Chinese herbs have been used to treat small-cell lung cancer (SCLC) due to their low toxicity and significant efficacy. This study focused on oridonin, a natural compound extracted from Rabdosia rubescens, and aimed to investigate its potential antitumor activity on SCLC and to evaluate the synergistic effect of combining oridonin with other small molecules. In this study, oridonin exhibited a dual effect. At lower concentrations, it suppressed the cell viability of SCLC cells (H1688 and H446). At high concentrations, oridonin induced SCLC cell apoptosis, damaged HBE cells in vitro and compromised the function of the liver and heart in vivo. The lower concentration of oridonin induced autophagy by enhancing the expression of p62 and the LC3B-II/LC3B-I ratio. This phenomenon might be associated with the activation of the protein kinase RNA-like ER kinase (PERK)/eukaryotic initiation factor 2 alpha (eIF2α)/growth arrest and DNA damage-inducible gene 153 (CHOP/GAD153) pathway. Therefore, the combined effect of oridonin with GSK2606414 or 3- methyladenine increased apoptosis in SCLC cells and reduced tumor growth. A similar phenomenon was observed after oridonin was combined with p62 or CHOP RNA interference treatment. Simultaneously, the combination of oridonin and GSK2606414 exhibited therapeutic efficacy without manifesting adverse effects. Our findings suggest that oridonin at lower concentrations can induce autophagy by activating the PERK/eIF2α/CHOP signaling pathway. The inhibition of the PERK/eIF2α/CHOP pathway could enhance oridonin therapeutic responses by triggering apoptosis. The novel therapeutic approach of combining oridonin with a PERK inhibitor is promising as a strategy for the treatment of SCLC.

Acacetin inhibits activation of microglia to improve neuroinflammation after subarachnoid hemorrhage through the PERK signaling pathway mediated autophagy.

PURPOSE: To explore the effect of acacetin on subarachnoid hemorrhage (SAH) and its possible mechanism. METHODS: SAH model of rat was established, and intraperitoneally injected with three doses of acacetin. To verify the role of PERK pathway, we used the CCT020312 (PERK inhibitor) and Tunicamycin (activators of endoplasmic reticulum stress). The SAH score, neurological function score, brain edema content, and Evans blue (EB) exudate were evaluated. Western blot was used to determine the expression of inflammation-associated proteins and PERK pathway. The activation of microglia was also determined through Iba-1 detection. TEM and immunofluorescence staining of LC3B were performed to observe the autophagy degree of SAH rats after acacetin. Tunel/NeuN staining, HE and Nissl' staining were performed for neuronal damage. RESULTS: Acacetin increased the neurological function score, reduce brain water content, Evans blue exudation and SAH scores. The microglia in cerebral cortex were activated after SAH, while acacetin could inhibit its activation, and decreased the expression of TNF-α and IL-6 proteins. The pathological staining showed the severe neuronal damage and increased neuronal apoptosis after SAH, while acacetin could improve these pathological changes. We also visualized the alleviated autophagy after acacetin. The expression of Beclin1 and ATF4 proteins were increased, but acacetin could inhibit them. Acacetin also inactivated PERK pathway, which could improve the neuronal injury and neuroinflammation after SAH, inhibit the microglia activation and the overactivated autophagy through PERK pathway. CONCLUSION: Acacetin may alleviate neuroinflammation and neuronal damage through PERK pathway, thus having the protective effect on EBI after SAH.

Calreticulin regulates the expression of MMP14 and ADAR1 through EIF2AK2 signaling to promote the proliferation and progression of malignant melanoma cells.

It has been demonstrated that calreticulin (CALR) is expressed abnormally in various tumors and is involved in the occurrence and development of tumors. In this study, CALR and EIF2AK2 expression was measured in the clinical specimens of 39 patients with melanoma. Then, we constructed knockdown and overexpression cell models of CALR and EIF2AK2 and used wound healing and Transwell assays to observe cell migration and invasion. Apoptosis, EDU, and ROS assays were used to measure cell apoptosis and proliferation, as well as ROS levels. The effect of CALR on endoplasmic reticulum stress was detected using endoplasmic reticulum fluorescent probes. Western blotting was used to detect protein levels of CALR, EIF2AK2, ADAR1, and MMP14. The results indicated that CALR and EIF2AK2 expression levels were significantly higher in human melanoma tissues than in adjacent non-tumor tissue. In addition, we found a correlation between CALR and the expression of EIF2AK2 and MMP14, and the experimental results indicated that overexpression of CALR significantly upregulated the expression of EIF2AK2, MMP14, and ADAR1, while knockdown of CALR inhibited their expression. Notably, the knockdown of EIF2AK2 in the CALR overexpression group blocked the upregulation of MMP14 and ADAR1 expression by CALR, and the knockdown of both CALR and EIF2AK2 significantly inhibited MMP14 and ADAR1 expression. In conclusion, CALR and EIF2AK2 play a promoting role in melanoma progression, and knockdown of CALR and EIF2AK2 may be an effective anti-tumor target, and its mechanism may be through MMP14, ADAR1 signaling.

The immune response to RNA suppresses nucleic acid synthesis by limiting ribose 5-phosphate.

During infection viruses hijack host cell metabolism to promote their replication. Here, analysis of metabolite alterations in macrophages exposed to poly I:C recognises that the antiviral effector Protein Kinase RNA-activated (PKR) suppresses glucose breakdown within the pentose phosphate pathway (PPP). This pathway runs parallel to central glycolysis and is critical to producing NADPH and pentose precursors for nucleotides. Changes in metabolite levels between wild-type and PKR-ablated macrophages show that PKR controls the generation of ribose 5-phosphate, in a manner distinct from its established function in gene expression but dependent on its kinase activity. PKR phosphorylates and inhibits the Ribose 5-Phosphate Isomerase A (RPIA), thereby preventing interconversion of ribulose- to ribose 5-phosphate. This activity preserves redox control but decreases production of ribose 5-phosphate for nucleotide biosynthesis. Accordingly, the PKR-mediated immune response to RNA suppresses nucleic acid production. In line, pharmacological targeting of the PPP during infection decreases the replication of the Herpes simplex virus. These results identify an immune response-mediated control of host cell metabolism and suggest targeting the RPIA as a potential innovative antiviral treatment.

Role of Neurocellular Endoplasmic Reticulum Stress Response in Alzheimer's Disease and Related Dementias Risk.

Currently, more than 55 million people around the world suffer from dementia, and Alzheimer's Disease and Related Dementias (ADRD) accounts for nearly 60-70% of all those cases. The spread of Alzheimer's Disease (AD) pathology and progressive neurodegeneration in the hippocampus and cerebral cortex is strongly correlated with cognitive decline in AD patients; however, the molecular underpinning of ADRD's causality is still unclear. Studies of postmortem AD brains and animal models of AD suggest that elevated endoplasmic reticulum (ER) stress may have a role in ADRD pathology through altered neurocellular homeostasis in brain regions associated with learning and memory. To study the ER stress-associated neurocellular response and its effects on neurocellular homeostasis and neurogenesis, we modeled an ER stress challenge using thapsigargin (TG), a specific inhibitor of sarco/endoplasmic reticulum Ca2+ ATPase (SERCA), in the induced pluripotent stem cell (iPSC)-derived neural stem cells (NSCs) of two individuals from our Mexican American Family Study (MAFS). High-content screening and transcriptomic analysis of the control and ER stress-challenged NSCs showed that the NSCs' ER stress response resulted in a significant decline in NSC self-renewal and an increase in apoptosis and cellular oxidative stress. A total of 2300 genes were significantly (moderated t statistics FDR-corrected p-value ≤ 0.05 and fold change absolute ≥ 2.0) differentially expressed (DE). The pathway enrichment and gene network analysis of DE genes suggests that all three unfolded protein response (UPR) pathways, protein kinase RNA-like ER kinase (PERK), activating transcription factor-6 (ATF-6), and inositol-requiring enzyme-1 (IRE1), were significantly activated and cooperatively regulated the NSCs' transcriptional response to ER stress. Our results show that IRE1/X-box binding protein 1 (XBP1) mediated transcriptional regulation of the E2F transcription factor 1 (E2F1) gene, and its downstream targets have a dominant role in inducing G1/S-phase cell cycle arrest in ER stress-challenged NSCs. The ER stress-challenged NSCs also showed the activation of C/EBP homologous protein (CHOP)-mediated apoptosis and the dysregulation of synaptic plasticity and neurotransmitter homeostasis-associated genes. Overall, our results suggest that the ER stress-associated attenuation of NSC self-renewal, increased apoptosis, and dysregulated synaptic plasticity and neurotransmitter homeostasis plausibly play a role in the causation of ADRD.