Overexpression Of X-box Binding Protein 1 Research Articles

X-Box binding protein 1 downregulates SIRT6 to promote injury in pancreatic ductal epithelial cells.

Acute pancreatitis (AP) stands as a frequent cause for clinical emergency hospital admissions. The X-box binding protein 1 (XBP1) was found to be implicated in pancreatic acinar cell apoptosis. The objective is to unveil the potential mechanisms governed by XBP1 and SIRT6 in the context of AP. Caerulein-treated human pancreatic duct epithelial (HPDE) cells to establish an in vitro research model. The levels and regulatory role of SIRT6 in the treated cells were evaluated, including its effects on inflammatory responses, oxidative stress, apoptosis, and endoplasmic reticulum stress. The relationship between XBP1 and SIRT6 was explored by luciferase and ChIP experiments. Furthermore, the effect of XBP1 overexpression on the regulatory function of SIRT6 on cells was evaluated. Caerulein promoted the decrease of SIRT6 and the increase of XBP1 in HPDE cells. Overexpression of SIRT6 slowed down the secretion of inflammatory factors, oxidative stress, apoptosis level, and endoplasmic reticulum stress in HPDE cells. However, XBP1 negatively regulated SIRT6, and XBP1 overexpression partially reversed the regulation of SIRT6 on the above aspects. Our study illuminates the role of XBP1 in downregulating SIRT6 in HPDE cells, thereby promoting cellular injury. Inhibiting XBP1 or augmenting SIRT6 levels holds promise in preserving cell function and represents a potential therapeutic avenue in the management of AP.

The unfolded protein response transcription factor XBP1s ameliorates Alzheimer’s disease by improving synaptic function and proteostasis

Alteration in the buffering capacity of the proteostasis network is an emerging feature of Alzheimer's disease (AD), highlighting the occurrence of endoplasmic reticulum (ER) stress. The unfolded protein response (UPR) is the main adaptive pathway to cope with protein folding stress at the ER. Inositol-requiring enzyme-1 (IRE1) operates asa central ER stress sensor, enabling the establishment ofadaptive and repair programs through the control of theexpression of the transcription factor X-box binding protein1 (XBP1). To artificially enforce the adaptive capacityof the UPR in the AD brain, we developed strategiesto express the active form of XBP1 in the brain. Overexpressionof XBP1 in the nervous system using transgenic mice reduced the load of amyloid deposits and preserved synaptic and cognitive function. Moreover, localdelivery of XBP1 into the hippocampus of an 5xFAD mice using adeno-associated vectors improved different ADfeatures. XBP1 expression corrected a large proportion of the proteomic alterations observed in the AD model, restoring the levels of several synaptic proteins and factorsinvolved in actin cytoskeleton regulation and axonal growth. Our results illustrate the therapeutic potential of targeting UPR-dependent gene expression programs as a strategy to ameliorate AD features and sustain synaptic function.

Boosting UPR transcriptional activator XBP1 accelerates acute wound healing.

Patients' suffering from large or deep wounds caused by traumatic and/or thermal injuries have significantly lower chances of recapitulating lost skin function through natural healing. We tested whether enhanced unfolded protein response (UPR) by expression of a UPR transcriptional activator, X-box-binding protein 1 (XBP1) can significantly promote wound repair through stimulating growth factor production and promoting angiogenesis. In mouse models of a second-degree thermal wound, a full-thickness traumatic wound, and a full-thickness diabetic wound, the topical gene transfer of the activated form of XBP1 (spliced XBP1, XBP1s) can significantly enhance re-epithelialization and increase angiogenesis, leading to rapid, nearly complete wound closure with intact regenerated epidermis and dermis. Overexpression of XBP1s stimulated the transcription of growth factors in fibroblasts critical to proliferation and remodeling during wound repair, including platelet-derived growth factor BB, basic fibroblast growth factor, and transforming growth factor beta 3. Meanwhile, the overexpression of XBP1s boosted the migration and tube formation of dermal microvascular endothelial cells in vitro. Our functional and mechanistic investigations of XBP1-mediated regulation of wound healing processes provide novel insights into the previously undermined physiological role of the UPR in skin injuries. The finding opens an avenue to developing potential XBP1-based therapeutic strategies in clinical wound care protocols.

Exosomal miR-205-5p derived from periodontal ligament stem cells attenuates the inflammation of chronic periodontitis via targeting XBP1.

Chronic periodontitis (CP) is an inflammatory periodontal disease with high incidence and complex pathology. This research is aimed to investigate the function of exosomal miR-205-5p (Exo-miR-205-5p) in CP and the underlying molecular mechanisms. Exo-miR-205-5p was isolated from miR-205-5p mimics-transfected periodontal ligament stem cells (PDLSCs), and subsequently cocultured with lipopolysaccharide (LPS)-induced cells or injected into LPS-treated rats. The mRNA expression of inflammatory factors and Th17/Treg-related factors were measured by quantitative real-time PCR. The contents of inflammatory factors and the percentages of Th17/Treg cells were measured by enzyme-linked immunosorbent assayand flow cytometry, respectively. Besides, the target relation between miR-205-5p and X-box binding protein 1 (XBP1) was explored. MiR-205-5p was downregulated in LPS-induced PDLSCs and corresponding exosomes. Exo-miR-205-5p inhibited inflammatory cell infiltration, decreased the production of TNF-α, IL-1β, and IL-6, and decreased the percentage of Th17 cells in LPS-treated rats. In addition, XBP1 was a target of miR-205-5p. Overexpression of XBP1 weakened the effects of Exo-miR-205-5p on inhibiting inflammation and regulating Treg/Th17 balance in LPS-induced cells. Exo-miR-205-5p derived from PDLSCs relieves the inflammation and balances the Th17/Treg cells in CP through targeting XBP1.

X-box binding protein 1: A new metabolic mediator and drug target of metformin?

Accumulating evidence has demonstrated that metformin improved hypertriglyceridemia. The present study aim to investigate the molecular mechanism by which metformin improves hypertriglyceridemia via regulation of diacylglycerol O-acyltransferase 2 (DGAT2) and X-box binding protein 1 (XBP1) in the liver and whether AMP-activated protein kinase (AMPK) is involved. Mice were fed a high-fat diet (HFD) or high-fat diet with metformin for 5weeks to evaluate the effect of metformin on triglyceride (TG) levels and expression of DGAT2 and XBP1 in the liver. In vitro HepG2 cells or XBP1 knockout AML12 hepatocytes were stimulated with metformin, palmitic acid or small interfering RNA inducing XBP1 knockdown, or dominant-negative mutant AMPK plasmid. Metformin treatment reduced hepatic TG levels in the liver of HFD-fed mice. Expression of nuclear and cytoplasmic XBP1 protein and its downstream target gene DGAT2 decreased in the liver of HFD-fed mice and HepG2 cells after metformin treatment. AMPK inactivation or overexpression of XBP1 attenuates this effect. Our preliminary results demonstrate that metformin activates AMPK to reduce TG synthesis by inhibiting the XBP1-mediated DGAT2 pathway, at least in part, suggesting that XBP1 is a new metabolic mediator for metformin treatment of hypertriglyceridemia and associated metabolic disease.

LBODP078 Role Of Ire1a-xbp1 Pathway In Rodent Functional Pituitary Tumor Cells.

Abstract Introduction Pituitary tumors (PTs) occur in almost 17% of the population and they represent about 10% of all intracranial tumors. At present, many patients with pituitary tumors suffer from symptoms and complications caused by abnormal hormone production and/or mass effects due to limited treatment options. Therefore, the development of novel therapies is an urgent issue. The inositol requiring enzyme 1a (IRE1a)-X-box binding protein 1 (XBP1) pathway is an important component for the endoplasmic reticulum (ER) stress response. This pathway is activated by immature protein overload in the ER and elimination or repair of such protein can reduce ER stress. This pathway has been reported to regulate cancer cell growth and secretory protein production. The detection of potential XBP1 binding sites in the promoter region of rat GH and mouse POMC similar to those of human genes suggests the involvement of the IRE1a-XBP1 pathway in the production of these hormones and pituitary tumor cell growth. In this study, we aimed to clarify the role of the IRE1a-XBP1 pathway in functional pituitary tumor cells. Methods and Results We conducted in vitro experiments by using rat GH-producing pituitary GH4 cells and mouse ACTH-producing AtT-20 cells as the models of GH- and ACTH-producing pituitary adenomas, respectively. Cell proliferation was measured by WST-8 assay, gene expression levels by qPCR, and protein expression by western blotting, respectively. In GH4 cells, STF08310 (40µM), an IRE1a inhibitor, reduced levels of GH and XBP1 spliced form (XBP1s) mRNA and GH protein with suppressed cell proliferation. In AtT-20 cells, STF08310 (40µM) also suppressed protein expression of proopiomelanocortin (POMC) and cell proliferation. Gene suppression of IRE1a by siRNA and over-expression of XBP1s showed marked reduction and increase of POMC protein, respectively. Conclusion Our findings suggest that the IRE1a-XBP1s pathway would be a possible therapeutic target that has both anti-tumoral and hormone control effects against functional pituitary tumor. Presentation: No date and time listed

XBP1 Regulates the Transcription of HIF-1a in BALB/c Mice with Chronic Rhinosinusitis without Polyps.

X-box binding protein 1 (XBP1) is a transcription factor that recognizes the CRE-like element in enhancers of human T-cell leukemia virus and MHC class II gene and induces their transcription. This study was performed to characterize the function of XBP1, which was identified to be a differentially expressed gene via GEO database, in chronic rhinosinusitis (CRS) without nasal polyps (CRSsNP). XBP1 expression was significantly elevated in both CRSsNP patients and mice who were accompanied with mucosal thickening, goblet cell hyperplasia and chemosis, glandular hyperplasia, and dense infiltration of inflammatory cells. Silencing of XBP1 suppressed the development of CRSsNP in mice. Mechanistically, knockdown of XBP1 downregulated the expression of hypoxia-inducible factor 1-alpha (HIF-1a), and overexpression of XBP1 led to the opposite result. Silencing of HIF-1a inhibited β-catenin expression and impaired the Wnt/β-catenin pathway. Further overexpression of HIF-1a in XBP1-silenced CRSsNP mice exacerbated pathological changes in mouse nasal mucosal tissues, promoted inflammation, and activated the Wnt/β-catenin pathway. Taken together, overexpression of XBP1 may be associated with increased expression of HIF-1a and possibly contribute to the Wnt/β-catenin pathway activation and the development of CRSsNP.

057 Targeting keratinocytes to potentiate skin immunization

Skin is a unique target for vaccination. Targeting keratinocytes to produce proinflammatory mediators to improve immunity is rational approach for vaccine design. Keratinocytes genetically engineered to overexpress the stress response related transcription factor x-box binding protein 1 (XBP1) increased production of proinflammatory mediators and co-delivered antigen. Keratinocyte-specific overexpression of XBP1 through cutaneous genetic immunization was transient and induced a pro-immunogenic skin microenvironment, including enhanced expression of proinflammatory mediators and co-delivered antigen, and increased skin-infiltration and stimulatory function of dendritic cells.

Notch-activated mesenchymal stromal/stem cells enhance the protective effect against acetaminophen-induced acute liver injury by activating AMPK/SIRT1 pathway

BackgroundNotch signaling plays important roles in regulating innate immunity. However, little is known about the role of Notch in mesenchymal stromal/stem cell (MSC)-mediated immunomodulation during liver inflammatory response.MethodsNotch activation in human umbilical cord-derived MSCs was performed by a tissue culture plate coated with Notch ligand, recombinant human Jagged1 (JAG1). Mice were given intravenous injection of Notch-activated MSCs after acetaminophen (APAP)-induced acute liver injury. Liver tissues were collected and analyzed by histology and immunohistochemistry.ResultsMSC administration reduced APAP-induced hepatocellular damage, as manifested by decreased serum ALT levels, intrahepatic macrophage/neutrophil infiltration, hepatocellular apoptosis and proinflammatory mediators. The anti-inflammatory activity and therapeutic effects of MSCs were greatly enhanced by Notch activation via its ligand JAG1. However, Notch2 disruption in MSCs markedly diminished the protective effect of MSCs against APAP-induced acute liver injury, even in the presence of JAG1 pretreatment. Strikingly, Notch-activated MSCs promoted AMP-activated protein kinase (AMPKα) phosphorylation, increased the sirtuins 1 (SIRT1) deacetylase expression, but downregulated spliced X-box-binding protein 1 (XBP1s) expression and consequently reduced NLR family pyrin domain-containing 3 (NLRP3) inflammasome activation. Furthermore, SIRT1 disruption or XBP1s overexpression in macrophages exacerbated APAP-triggered liver inflammation and augmented NLRP3/caspase-1 activity in MSC-administrated mice. Mechanistic studies further demonstrated that JAG1-pretreated MSCs activated Notch2/COX2/PGE2 signaling, which in turn induced macrophage AMPK/SIRT1 activation, leading to XBP1s deacetylation and inhibition of NLRP3 activity.ConclusionsActivation of Notch2 is required for the ability of MSCs to reduce the severity of APAP-induced liver damage in mice. Our findings underscore a novel molecular insights into MSCs-mediated immunomodulation by activating Notch2/COX2/AMPK/SIRT1 pathway and thus provide a new strategy for the treatment of liver inflammatory diseases.

MicroRNA-194-5p Attenuates Doxorubicin-Induced Cardiomyocyte Apoptosis and Endoplasmic Reticulum Stress by Targeting P21-Activated Kinase 2.

ObjectiveMany studies have reported that microRNAs (miRs) are involved in the regulation of doxorubicin (DOX)-induced cardiotoxicity. MiR-194-5p has been reported significantly upregulated in patients with myocardial infarction; however, its role in myocardial diseases is still unclear. Various stimuluses can trigger the endoplasmic reticulum (ER) stress and it may activate the apoptosis signals eventually. This study aims to explore the regulatory role of miR-194-5p in DOX-induced ER stress and cardiomyocyte apoptosis.MethodsH9c2 was treated with 2 μM DOX to induce apoptosis, which is to stimulate the DOX-induced cardiotoxicity model. The expression of miR-194-5p was detected by quantitative real-time PCR (qRT-PCR); the interaction between miR-194-5p and P21-activated kinase 2 (PAK2) was tested by dual luciferase reporter assay; terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay and caspase-3/7 activity were used to assess apoptosis; trypan blue staining was applied to measure cell death; Western blotting was performed to detect protein expressions; and ER-related factors splicing X-box binding protein 1 (XBP1s) was detected by polyacrylamide gel electrophoresis and immunofluorescence to verify the activation of ER stress.ResultsMiR-194-5p was upregulated in cardiomyocytes and mouse heart tissue with DOX treatment, while the protein level of PAK2 was downregulated. PAK2 was predicted as the target of miR-194-5p; hence, dual luciferase reporter assay indicated that miR-194-5p directly interacted with PAK2 and inhibited its expression. TUNEL assay, caspase-3/7 activity test, and trypan blue stain results showed that either inhibition of miR-194-5p or overexpression of PAK2 reduced DOX-induced cardiomyocyte apoptosis. Silencing of miR-194-5p also improved DOX-induced cardiac dysfunction. In addition, DOX could induce ER stress in H9c2, which led to XBP1 and caspase-12 activation. The expression level of XBP1s with DOX treatment increased first then decreased. Overexpression of XBP1s suppressed DOX-induced caspase-3/7 activity elevation as well as the expression of cleaved caspase-12, which protected cardiomyocyte from apoptosis. Additionally, the activation of XBP1s was regulated by miR-194-5p and PAK2.ConclusionOur findings revealed that silencing miR-194-5p could alleviate DOX-induced cardiotoxicity via PAK2 and XBP1s in vitro and in vivo. Thus, the novel miR-194-5p/PAK2/XBP1s axis might be the potential prevention/treatment targets for cancer patients receiving DOX treatment.

XBP1s promotes the development of lung adenocarcinoma via the p‑JNK MAPK pathway.

Spliced X-box binding protein 1 (XBP1s) has been reported to participate in the pathogenesis of numerous types of cancer; however, whether XBP1s plays a role in lung cancer remains to be elucidated. In the present study, bioinformatics analysis was performed to determine the mRNA expression level of XBP1 in lung cancer and adjacent normal tissues. Gene Ontology terms, pathway enrichment and Pearson's correlation analysis were performed to investigate the possible mechanism involved. Western blot and reverse transcription-quantitative PCR were performed to quantify the protein and mRNA expression level of target proteins, respectively. Small interfering RNA or overexpression plasmid were used to knockdown or overexpress the expression level of XBP1s. EdU staining, colony formation, Cell Counting Kit-8, Transwell and wound healing assays, and flow cytometry were performed to detect the proliferation, colony forming ability, cell viability, migration and invasion ability, and the apoptosis rate. The results showed that the mRNA and protein expression level of XBP1 was higher in tumor tissues compared with that in adjacent normal tissues using data from the TIMER2.0, ONCOMINE and UALCAN online databases. In addition, the mRNA expression level of XBP1 was also associated with clinical features, including age, smoking habit, individual cancer stage and nodal metastasis status. In the in vitro experiments, the mRNA and protein expression level of XBP1s was increased in the A549 cell line compared with that in the human bronchial epithelial (HBE), H1299, PC9 and H460 cell lines. Hypoxia further increased the protein expression level of XBP1s in the A549 cell line. Knockdown of XBP1s expression in the A549 cell line resulted in decreased proliferation, colony formation, cell viability, migration and invasion, and increased apoptosis. By contrast, overexpressing XBP1s in the HBE cell line led to the opposite results. To investigate the mechanism involved, proteins associated with XBP1 were analyzed using the LinkedOmics database. Pathway enrichment revealed the MAPK pathway to be the possible XBP1 downstream target. Furthermore, Pearson's correlation and western blot analyses verified that phosphorylated (p)-JNK rather than p-ERK or p-p38 was the downstream effector of XBP1s. Phosphorylation of JNK was decreased when XBP1s expression was knocked down in the A549 cell line under normoxic and hypoxic conditions. Inhibiting p-JNK with SP600125 reversed the increased prosurvival effects caused by XBP1s overexpression. The results from the present study suggest that XBP1s/p-JNK function as a prosurvival factors in the A549 cell line and could be a potential target for the treatment of lung adenocarcinoma.

Scutellarin ameliorates hepatic lipid accumulation by enhancing autophagy and suppressing IRE1α/XBP1 pathway.

Nonalcoholic fatty liver disease is the most prevalent liver disease characterized by excessive lipid accumulation in hepatocytes. Endoplasmic reticulum (ER) stress and autophagy play an important role in lipid accumulation. In this study, scutellarin (Scu) was examined in palmitic acid-treated HepG2 cells and C57/BL6 mice fed a high-fat diet (HFD). Scu reduced intracellular lipid content and inhibited sterol regulatory element binding protein-1c (SREBP-1c)-mediated lipid synthesis and fatty acid translocase-mediated lipid uptake in HepG2 cells. Additionally, Scu restored impaired autophagy and inhibited excessive activation of ER stress in vivo and in vitro. Moreover, Scu upregulated forkhead box O transcription factor 1-mediated autophagy by inhibiting inositol-requiring enzyme 1α (IRE1α)/X-box-binding protein 1 (XBP1) branch activation, while XBP1s overexpression exacerbated the lipid accumulation and impaired autophagy in HepG2 cells and also weakened the positive effects of Scu. Furthermore, Scu attenuated ER stress by activating autophagy, ultimately downregulating SREBP-1c-mediated lipid synthesis, and autophagy inhibitors offset these beneficial effects. Scu inhibited the crosstalk between autophagy and ER stress and downregulated saturated fatty acid-induced lipid accumulation in hepatocytes. These findings demonstrate that Scu ameliorates hepatic lipid accumulation by enhancing autophagy and suppressing ER stress via the IRE1α/XBP1 pathway.

X-box-binding protein 1 is required for pancreatic development in Xenopus laevis.

X-box-binding protein 1 (XBP1) is a protein containing the basic leucine zipper structure. It belongs to the cAMP-response element binding protein (CREB)/activating transcription factor transcription factor family. As the main transcription factor, spliced XBP1 (XBP1s) participates in many physiological and pathological processes and plays an important role in embryonic development. Previous studies showed that XBP1-knockout mice died because of pancreatic exocrine function deficiency, indicating that XBP1 plays an important role in pancreatic development. However, the exact role of XBP1 in pancreatic development remains unclear. This study aimed to investigate the role of XBP1 in the pancreatic development of Xenopus laevis embryos. Whole-mount in situ hybridization and quantitative real-time PCR results revealed that the expression levels of pancreatic progenitor marker genes pdx1, p48, ngn3, and sox9 were downregulated in XBP1s morpholino oligonucleotide (MO)-injected embryos. The expression levels of pancreatic exocrine and endocrine marker genes insulin and amylase were also downregulated. Through the overexpression of XBP1s, the phenotype and gene expressions were opposite to those in XBP1s MO-injected embryos. Luciferase and chromatin immunoprecipitation assays showed that XBP1s could bind to the XBP1-binding site in the foxa2 promoter. These results revealed that XBP1 is required in the pancreatic development of Xenopus laevis and might function by regulating foxa2.

Selenoprotein S regulates adipogenesis through IRE1α-XBP1 pathway.

The induction of endoplasmic reticulum (ER) stress is associated with adipogenesis, during which the inositol-requiring enzyme 1 alpha (IRE1α)-X-box-binding protein 1 (XBP1) pathway is involved. Selenoprotein S (SelS), which is an ER resident selenoprotein, is involved in ER homeostasis regulation; however, little is known about the role of SelS in regulating adipogenesis. In vivo studies showed that SelS protein levels in white adipose tissue were increased in obese subjects and high-fat diet (HFD)-fed mice. Moreover, we identified that SelS protein levels increased in the early phase of adipogenesis and then decreased in the late phase during adipogenesis. Overexpression of SelS promoted adipogenesis. Conversely, knockdown (KD) of SelS resulted in the inhibition of adipogenesis, which was related to increasing cell death, decreased mitotic clonal expansion, and cell cycle G1 arrest. In vivo studies also showed that ER stress markers (p-IRE1α/IRE1α, XBP1s, and Grp78) were significantly increased with upregulating of SelS expression in subcutaneous and visceral adipose tissues in the obese subjects and HFD-fed mice. Furthermore, in SelS KD cells, the levels of Grp78 were increased and the levels of p-IRE1α/IRE1α were unchanged , but mRNA levels of spliced XBP1 (XBP1s) produced by IRE1α-mediated splicing were decreased, suggesting a role of SelS in the modulation of IRE1α-XBP1 pathway. Moreover, inhibition of adipogenesis by SelS suppression can be rescued by overexpression of XBP1s. Thus, SelS appears to function as a novel regulator of adipogenesis through the IRE1α-XBP1 signaling pathway.

Beneficial effect of ER stress preconditioning in protection against FFA-induced adipocyte inflammation via XBP1 in 3T3-L1 adipocytes

Adipose tissue inflammation is closely associated with the development of obesity and insulin resistance. Free fatty acids (FFAs) are a major inducer of obesity-related insulin resistance. Previously, we reported that endoplasmic reticulum (ER) stress potentially mediated retinal inflammation in diabetic retinopathy. The unfolded protein response (UPR) protects cells against damage induced by oxidative stress. X-box binding protein 1 (XBP1) plays a major role in protecting cells by modulating the UPR. However, the link between ER stress and adipocyte inflammation has been poorly investigated. In the present study, we found that pretreatment of 3T3-L1 adipocytes with a low dose of ER stress inducer tunicamycin inhibited FFA-induced upregulated expression of inflammatory cytokines. In addition, FFAs induced phosphorylation of the p65 subunit of NF-κB was largely inhibited by pretreatment with tunicamycin in 3T3-L1 adipocytes. Knockdown of XBP1 by siRNA markedly mitigated the protective effects of preconditioning against inflammation. Conversely, overexpression of XBP1 alleviated FFA-induced phosphorylation of IκB-α, IKKα/β, and NF-κB, which was accompanied by decreased inflammatory cytokine expression. Collectively, these results imply a beneficial role of ER stress preconditioning in protecting against FFA-induced 3T3-L1 adipocyte inflammation, which is likely mediated through inhibition of the IKK/NF-κB pathway via XBP1.

Deacetylation of XBP1s by sirtuin 6 confers resistance to ER stress-induced hepatic steatosis

The active spliced form of X-box-binding protein 1 (XBP1s) is a key modulator of ER stress, but the functional role of its post-translational modification remains unclear. Here, we demonstrate that XBP1s is a deacetylation target of Sirt6 and that its deacetylation protects against ER stress-induced hepatic steatosis. Specifically, the abundance of acetylated XBP1s and concordant hepatic steatosis were increased in hepatocyte-specific Sirt6 knockout and obese mice but were decreased by genetic overexpression and pharmacological activation of Sirt6. Mechanistically, we identified that Sirt6 deacetylated a transactivation domain of XBP1s at Lys257 and Lys297 and promoted XBP1s protein degradation through the ubiquitin-proteasome system. Overexpression of XBP1s, but not its deacetylation mutant 2KR (K257/297R), in mice increased lipid accumulation in the liver. Importantly, in liver tissues obtained from patients with nonalcoholic fatty liver disease (NAFLD), the extent of XBP1s acetylation correlated positively with the NAFLD activity score but negatively with the Sirt6 level. Collectively, we present direct evidence supporting the importance of XBP1 acetylation in ER stress-induced hepatic steatosis.

Spliced X-box Binding Protein 1 Stimulates Adaptive Growth Through Activation of mTOR

The unfolded protein response plays versatile roles in physiology and pathophysiology. Its connection to cell growth, however, remains elusive. Here, we sought to define the role of unfolded protein response in the regulation of cardiomyocyte growth in the heart. We used both gain- and loss-of-function approaches to genetically manipulate XBP1s (spliced X-box binding protein 1), the most conserved signaling branch of the unfolded protein response, in the heart. In addition, primary cardiomyocyte culture was used to address the role of XBP1s in cell growth in a cell-autonomous manner. We found that XBP1s expression is reduced in both human and rodent cardiac tissues under heart failure. Furthermore, deficiency of XBP1s leads to decompensation and exacerbation of heart failure progression under pressure overload. On the other hand, cardiac-restricted overexpression of XBP1s prevents the development of cardiac dysfunction. Mechanistically, we found that XBP1s stimulates adaptive cardiac growth through activation of the mechanistic target of rapamycin signaling, which is mediated via FKBP11 (FK506-binding protein 11), a novel transcriptional target of XBP1s. Moreover, silencing of FKBP11 significantly diminishes XBP1s-induced mechanistic target of rapamycin activation and adaptive cell growth. Our results reveal a critical role of the XBP1s-FKBP11-mechanistic target of rapamycin axis in coupling of the unfolded protein response and cardiac cell growth regulation.

XBP1 inhibits mesangial cell apoptosis in response to oxidative stress via the PTEN/AKT pathway in diabetic nephropathy.

Diabetic nephropathy (DN) is a complication of diabetes mellitus (DM) that frequently results in renal disease, and is characterized by a variety of symptoms, including albuminuria. It has been shown that apoptosis of glomerular mesangial cells (MCs) can aggravate albuminuria and contribute to the development of diabetic glomerulosclerosis. Hence, determination of the mechanisms leading to MC apoptosis may help us gain insights into the pathogenesis of DN. As our understanding of the role of high glucose (HG) in MC apoptosis remains elusive, we explored the interplay between X‐box binding protein 1 (XBP1) and MC apoptosis in this study. XBP1 was observed to be downregulated both in vivo and in vitro. Treatment of XBP1‐overexpressing cells with HG resulted in a decrease of reactive oxygen species (ROS) and a suppression of cell apoptosis, concomitant with decreases in cleaved caspase‐3 and Bax. Subsequent analyses demonstrated that XBP1 overexpression inhibited the expression of phosphatase and tensin homolog deleted on chromosome ten (PTEN) and enhanced the activation of AKT in MCs exposed to HG. In addition, XBP1‐induced injuries in MC were reversed by overexpression of PTEN, and XBP1 inhibited apoptosis, which was mediated by the activated PTEN/AKT signaling pathway. Thus, our data indicate that XBP1 can activate the PTEN/AKT signaling pathway, thereby alleviating oxidative stress caused by HG or MC apoptosis. These findings suggest that XBP1 may have potential in the development of treatment methods for DN.

Nitrosative stress drives heart failure with preserved ejection fraction.

Heart failure with preserved ejection fraction (HFpEF) is a common syndrome with high morbidity and mortality for which there are no evidence-based therapies. Here we report that concomitant metabolic and hypertensive stress in mice-elicited by a combination of high-fat diet and inhibition of constitutive nitric oxide synthase using Nω-nitro-L-arginine methyl ester (L-NAME)-recapitulates the numerous systemic and cardiovascular features of HFpEF in humans. Expression of one of the unfolded protein response effectors, the spliced form of X-box-binding protein 1 (XBP1s), was reduced in the myocardium of ourrodent model and in humans with HFpEF. Mechanistically, the decrease in XBP1s resulted from increased activity of inducible nitric oxide synthase (iNOS) and S-nitrosylation of theendonuclease inositol-requiring protein 1α (IRE1α), culminating in defective XBP1 splicing. Pharmacological or genetic suppression of iNOS, or cardiomyocyte-restricted overexpression of XBP1s, each ameliorated the HFpEF phenotype. We report that iNOS-driven dysregulation of the IRE1α-XBP1 pathway is a crucial mechanism of cardiomyocyte dysfunction in HFpEF.

The oncogenic role of GNL3 in the progression and metastasis of osteosarcoma.

BackgroundGNL3 has been reported to be up-regulated in cancers and function in tumor progression, whereas the role of GNL3 in the progression of osteosarcoma remains unclear.Materials and methodsIn this study, we blocked the expression of GNL3 by siRNA interference in osteosarcoma cell lines MG63 and U20S. CCK8, colony formation, wound-healing, Transwell, flow cytometry, and Hoechst/PI staining assays were used to examine the effects of GNL3 knockdown on cell proliferation, migration, invasion and apoptosis in MG63 and U20S cells. The relative activity of MMP9 was detected using Gelatin zymography assay. Western blot was performed to detect the expression of related proteins.ResultsWe found that silencing of GNL3 reduced the growth, migration, and invasion abilities of MG63 and U20S cells. Moreover, silencing GNL3 triggered cell cycle arrest in MG63 and U20S cells, as well as promoted cell apoptosis. In addition, depletion of GNL3 was observed to reduce the activity of MMP9 and suppress the process of epithelial–mesenchymal transition (EMT) through up-regulation of E-cadherin and down-regulation of N-cadherin. Furthermore, we found that X-box-binding protein 1 (XBP1) could bind to GNL3 using dual-luciferase reporter assay, and XBP1 overexpression could restore the inhibitory effects on proliferation, invasion, and EMT in MG63 and U20S cells caused by GNL3 knockdown.ConclusionThese data suggest that GNL3 functions as an oncogene in the progression of osteosarcoma by regulation of EMT, and XBP1 is also involved in its mechanism.