bims-unfpre Biomed news
on Unfolded protein response
Issue of 2019‒03‒03
seventeen papers selected by
Susan Logue
Apoptosis Research Centre

  1. Sci Rep. 2019 Mar 01. 9(1): 3245
    Bagratuni T, Sklirou AD, Kastritis E, Liacos CI, Spilioti C, Eleutherakis-Papaiakovou E, Kanellias N, Gavriatopoulou M, Terpos E, Trougakos IP, Dimopoulos MA.
      Despite recent biomedical improvements in treating Multiple Myeloma (MM), the disease still remains incurable. Toll like receptors (TLRs) provide a link between innate and adaptive immune responses and hence potentially correlate inflammation to cancer. Although the regulatory role of TLRs in MM has been under investigation the underlying mechanisms remain unclear. In this study we assayed the function of TLR4 in MM cell lines and in MM patients' samples. We found that lipopolysaccharide-mediated TLR4 activation increased MM cells proliferation and decreased endoplasmic reticulum (ER) stress-induced apoptosis. Furthermore, we observed that either the endogenous CHOP expression or the ER stress-mediated CHOP induction, were suppressed by TLR4 activation or its overexpression in MM cell lines; TLR4 induction also suppressed ER stress-induced apoptotic signals. In support, TLR4 gene expression silencing in MM cell lines significantly decreased cell proliferation and promoted CHOP and ATF4 upregulation. TLR4 activation was also able to partially abrogate the effect of bortezomib in MM cell lines by suppressing PERK, ATF4 and phospho-eIF2A. We suggest that TLR4-mediated disruption of ER stress responses contributes to MM cells proliferation and suppresses ER-dependent death signals.
  2. Sci Rep. 2019 Feb 27. 9(1): 2903
    Patel V, Bidault G, Chambers JE, Carobbio S, Everden AJT, Garcés C, Dalton LE, Gribble FM, Vidal-Puig A, Marciniak SJ.
      Phosphorylation of the translation initiation factor eIF2α within the mediobasal hypothalamus is known to suppress food intake, but the role of the eIF2α phosphatases in regulating body weight is poorly understood. Mice deficient in active PPP1R15A, a stress-inducible eIF2α phosphatase, are healthy and more resistant to endoplasmic reticulum stress than wild type controls. We report that when female Ppp1r15a mutant mice are fed a high fat diet they gain less weight than wild type littermates owing to reduced food intake. This results in healthy leaner Ppp1r15a mutant animals with reduced hepatic steatosis and improved insulin sensitivity, albeit with a possible modest defect in insulin secretion. By contrast, no weight differences are observed between wild type and Ppp1r15a deficient mice fed a standard diet. We conclude that female mice lacking the C-terminal PP1-binding domain of PPP1R15A show reduced dietary intake and preserved glucose tolerance. Our data indicate that this results in reduced weight gain and protection from diet-induced obesity.
  3. Science. 2019 03 01. 363(6430): 993-998
    Grootjans J, Krupka N, Hosomi S, Matute JD, Hanley T, Saveljeva S, Gensollen T, Heijmans J, Li H, Limenitakis JP, Ganal-Vonarburg SC, Suo S, Luoma AM, Shimodaira Y, Duan J, Shih DQ, Conner ME, Glickman JN, Fuhler GM, Palm NW, de Zoete MR, van der Woude CJ, Yuan GC, Wucherpfennig KW, Targan SR, Rosenstiel P, Flavell RA, McCoy KD, Macpherson AJ, Kaser A, Blumberg RS.
      Immunoglobulin A (IgA) is the major secretory immunoglobulin isotype found at mucosal surfaces, where it regulates microbial commensalism and excludes luminal factors from contacting intestinal epithelial cells (IECs). IgA is induced by both T cell-dependent and -independent (TI) pathways. However, little is known about TI regulation. We report that IEC endoplasmic reticulum (ER) stress induces a polyreactive IgA response, which is protective against enteric inflammation. IEC ER stress causes TI and microbiota-independent expansion and activation of peritoneal B1b cells, which culminates in increased lamina propria and luminal IgA. Increased numbers of IgA-producing plasma cells were observed in healthy humans with defective autophagy, who are known to exhibit IEC ER stress. Upon ER stress, IECs communicate signals to the peritoneum that induce a barrier-protective TI IgA response.
  4. J Innate Immun. 2019 Feb 20. 1-15
    Colli ML, Paula FM, Marselli L, Marchetti P, Roivainen M, Eizirik DL, Op de Beeck A.
      Type 1 diabetes (T1D) is an autoimmune disease characterized by islet inflammation and progressive pancreatic β cell destruction. The disease is triggered by a combination of genetic and environmental factors, but the mechanisms leading to the triggering of early innate and late adaptive immunity and consequent progressive pancreatic β cell death remain unclear. The insulin-producing β cells are active secretory cells and are thus particularly sensitive to endoplasmic reticulum (ER) stress. ER stress plays an important role in the pathologic pathway leading to autoimmunity, islet inflammation, and β cell death. We show here that group B coxsackievirus (CVB) infection, a putative causative factor for T1D, induces a partial ER stress in rat and human β cells. The activation of the PERK/ATF4/CHOP branch is blunted while the IRE1α branch leads to increased spliced XBP1 expression and c-Jun N-terminal kinase (JNK) activation. Interestingly, JNK1 activation is essential for CVB amplification in both human and rat β cells. Furthermore, a chemically induced ER stress preceding viral infection increases viral replication, in a process dependent on IRE1α activation. Our findings show that CVB tailors the unfolded protein response in β cells to support their replication, preferentially triggering the pro-viral IRE1α/XBP1s/JNK1 pathway while blocking the pro-apoptotic PERK/ATF4/CHOP pathway.
    Keywords:  Endoplasmic reticulum stress; Enterovirus; IRE1α; Type 1 diabetes; c-Jun N-terminal kinase
  5. Appl Clin Genet. 2019 ;12 11-18
    Kumrah R, Mathew B, Vignesh P, Singh S, Rawat A.
      Inborn errors of immunity usually not only result in immunodeficiency but may also manifest as immune dysregulation in the form of autoinflammation, autoimmunity, or sometimes malignancy. One of the most recently discovered monogenic disorder of immune dysregulation is COPA syndrome. COPA syndrome is an inherited autoimmune disorder caused by mutations in COPA gene. COPA gene encodes for α subunit of the COP1 protein, which is involved in the reverse vesicular protein transport from Golgi apparatus to the endoplasmic reticulum (ER). The inheritance pattern of COPA syndrome is autosomal dominant, and the patients typically present with interstitial lung disease with pulmonary hemorrhage and subsequently develop arthritis. Immunological features involve autoantibody formation, elevated expression of IL-1β and IL-6, and increase in the number of Th17 cells. Molecular pathophysiology of COPA syndrome is not clearly understood. However, it is known that accumulation of unfolded proteins in ER leads to ER stress, which is an indirect result of aberrant vesicular transport of proteins from Golgi apparatus to ER and defective cellular autophagy. ER stress induces inflammation and is responsible for pathogenesis of a large number of chronic inflammatory diseases. Unfolded protein response process responds to improperly folded proteins and defends against stress in ER to ensure the fidelity of the protein folding. It maintains the expression of stress-response genes and causes initiation of inflammatory signaling pathways essential for the innate immunity. Mutation in COPA gene associated with defective protein sorting to ER has unearthed a new primary immunodeficiency disease with a unique clinical phenotype. This review highlights the clinical and molecular aspects of COPA syndrome.
    Keywords:  COPA syndrome; Golgi apparatus; arthritis; autoimmunity; autoinflammation; endoplasmic reticulum stress; interleukins; interstitial lung disease; protein transport
  6. Cell Metab. 2019 Feb 13. pii: S1550-4131(19)30020-8. [Epub ahead of print]
    Harris IS, Endress JE, Coloff JL, Selfors LM, McBrayer SK, Rosenbluth JM, Takahashi N, Dhakal S, Koduri V, Oser MG, Schauer NJ, Doherty LM, Hong AL, Kang YP, Younger ST, Doench JG, Hahn WC, Buhrlage SJ, DeNicola GM, Kaelin WG, Brugge JS.
      Cells are subjected to oxidative stress during the initiation and progression of tumors, and this imposes selective pressure for cancer cells to adapt mechanisms to tolerate these conditions. Here, we examined the dependency of cancer cells on glutathione (GSH), the most abundant cellular antioxidant. While cancer cell lines displayed a broad range of sensitivities to inhibition of GSH synthesis, the majority were resistant to GSH depletion. To identify cellular pathways required for this resistance, we carried out genetic and pharmacologic screens. Both approaches revealed that inhibition of deubiquitinating enzymes (DUBs) sensitizes cancer cells to GSH depletion. Inhibition of GSH synthesis, in combination with DUB inhibition, led to an accumulation of polyubiquitinated proteins, induction of proteotoxic stress, and cell death. These results indicate that depletion of GSH renders cancer cells dependent on DUB activity to maintain protein homeostasis and cell viability and reveal a potentially exploitable vulnerability for cancer therapy.
    Keywords:  GCLC; HSF1; UPR; antioxidants; cancer; deubiquitinase; glutamate-cysteine ligase catalytic subunit; glutathione; heat shock factor 1; high-throughput screening; oxidative stress; proteotoxic stress; unfolded protein response
  7. J Cell Physiol. 2019 Mar 01.
    Chen YC, Chen BC, Huang HM, Lin SH, Lin CH.
      In the present study, we investigated the role of PKR-like endoplasmic reticular kinase (PERK), an endoplasmic reticulum (ER) stress kinase, in endothelin 1 (ET-1)- and thrombin-induced pulmonary fibrosis (PF), and the preventive effects of curcumin (CUR). Using the human embryonic WI-38 lung fibroblast cell line, ET-1 and thrombin induced the expression of ER stress-related proteins (CCAAT-enhancer-binding protein homologous protein, PERK, and binding immunoglobulin protein), a profibrogenic factor (cellular communication network factor 2 [CCN2]), and differentiation markers including α-smooth muscle actin (α-SMA), collagen I (Col I), and Col IV. Knockdown of PERK expression via small interfering RNA (siRNA) significantly reduced the increases in CCN2, α-SMA, Col I, and Col IV proteins in WI-38 cells according to western blot analysis and immunohistochemistry (IHC). Activation of c-Jun N-terminal kinase (JNK) was observed in ET-1- and thrombin-treated WI-38 cells, and the addition of a JNK inhibitor (SP) suppressed the induction of the indicated proteins by ET-1 and thrombin. Thapsigargin (TG), an ER stress inducer, elevated expressions of PERK and ER stress-related proteins with increased differentiation of WI-38 cells. Knockdown of PERK by siRNA or the PERK inhibitor glycogen synthesis kinase reduced expressions of the differentiation markers, α-SMA and Col IV, in WI-38 cells. CUR concentration-dependently inhibited ET-1- or thrombin-induced CCN2, α-SMA, and vimentin proteins with decreased levels of phosphorylated mitogen-activated protein kinase and PERK in WI-38 cells. An in vivo bleomycin-induced PF study showed that an intraperitoneal injection of CUR (30 mg/kg) reduced expressions of α-SMA, CCN2, Col IV, and vimentin in lung tissues via IHC staining using specific antibodies. This study is the first to demonstrate that PERK activation contributes to pulmonary fibroblast differentiation elicited by ET-1 or thrombin, and the inhibitory activity of CUR against PF is demonstrated herein.
    Keywords:  ERK; curcumin; endothelin 1; lung fibrosis; thrombin
  8. ACS Chem Biol. 2019 Mar 01.
    Grandjean JMD, Plate L, Morimoto RI, Bollong MJ, Powers ET, Wiseman RL.
      Cellular proteostasis is maintained by stress-responsive signaling pathways such as the heat shock response (HSR), the oxidative stress response (OSR), and the unfolded protein response (UPR). Activation of these pathways results in the transcriptional upregulation of select subsets of stress-responsive genes that restore proteostasis and adapt cellular physiology to promote recovery following various types of acute insult. The capacity for these pathways to regulate cellular proteostasis makes them attractive therapeutic targets to correct proteostasis defects associated with diverse diseases. High-throughput screening (HTS) using cell-based reporter assays is highly effective for identifying putative activators of stress-responsive signaling pathways. However, the development of these compounds is hampered by the lack of medium-throughput assays to define compound potency and selectivity for a given pathway. Here, we describe a targeted RNA sequencing (RNAseq) assay that allows cost effective, medium-throughput screening of stress-responsive signaling pathway activation. We demonstrate that this assay allows deconvolution of stress-responsive signaling activated by chemical genetic or pharmacologic agents. Furthermore, we use this assay to define the selectivity of putative OSR and HSR activating compounds previously identified by HTS. Our results demonstrate the potential for integrating this adaptable targeted RNAseq assay into screening programs focused on developing pharmacologic activators of stress-responsive signaling pathways.
  9. Sci Rep. 2019 Mar 01. 9(1): 3210
    Shi Z, Yu X, Yuan M, Lv W, Feng T, Bai R, Zhong H.
      Colon cancer is a major health problem worldwide. While chemotherapy remains a main approach for treating late-stage colon cancer patients, most, if not all, of them will develop drug resistance and die of uncontrollable disease progression eventually. Therefore, identification of mechanism of drug resistance and development of overcoming strategy hold great significance in management of colon cancer. In this study, we discovered that activation of the PERK branch of the unfolded protein response (UPR) pathways is required for colon cancer cells to survive treatment of 5-Fluorouracil (5-FU), one of the first-line chemotherapeutics for late-stage colon cancer patients. Genetic and pharmacological inhibition of PERK or its downstream factors greatly sensitize colon cancer cells to 5-FU. Most importantly, in vivo use of PERK inhibitor synergizes with 5-FU in suppressing the growth of colon cancer cells in mouse models. In summary, our findings established a promising way to overcome resistance to chemotherapy in colon cancer.
  10. J Biol Chem. 2019 Mar 01. pii: jbc.RA118.006801. [Epub ahead of print]
    Emrich SM, Yoast RE, Xin P, Zhang X, Pathak T, Nwokonko R, Gueguinou M, Subedi KP, Zhou Y, Ambudkar IS, Hempel N, Machaca K, Gill DL, Trebak M.
      Store-operated Ca2+ entry (SOCE) is a ubiquitous pathway for Ca2+ influx across the plasma membrane (PM). SOCE is mediated by the endoplasmic reticulum (ER)-associated Ca2+-sensing proteins stromal interaction molecule 1 (STIM1) and STIM2, which transition into an active conformation in response to ER Ca2+ store depletion, thereby interacting with and gating PM-associated ORAI1 channels. Although structurally homologous, STIM1 and STIM2 generate distinct Ca2+ signatures in response to varying strengths of agonist stimulation. The physiological functions of these Ca2+ signatures, particularly under native conditions, remain unclear. To investigate the structural properties distinguishing STIM1 and STIM2 activation of ORAI1 channels under native conditions, here we used CRISPR/Cas9 to generate STIM1-/-, STIM2-/-, and STIM1/2-/- knockouts in HEK293 and colorectal HCT116 cells. We show that depending on cell type, STIM2 can significantly sustain SOCE in response to maximal store depletion. Utilizing the SOCE modifier 2-aminoethoxydiphenyl borate (2-APB), we demonstrate that 2-APB-activated store-independent Ca2+ entry is mediated exclusively by endogenous STIM2. Using variants that either stabilize or disrupt intramolecular interactions of STIM C termini, we show that the increased flexibility of the STIM2 C terminus contributes to its selective store-independent activation by 2-APB. However, STIM1 variants with enhanced flexibility in C terminus failed to support its store-independent activation. STIM1/STIM2 chimeric constructs indicated that coordination between N-terminal sensitivity and C-terminal flexibility is required for specific store-independent STIM2 activation. Our results clarify the structural determinants underlying activation of specific STIM isoforms, insights that are potentially useful for isoform-selective drug targeting.
    Keywords:  Calcium signaling; Ion sensor; SOCE; STIM2; calcium channel; calcium imaging; calcium intracellular release; calcium release-activated calcium channel protein 1 (ORAI1); calcium transport; cation channel; stromal interaction molecule 1 (STIM1)
  11. J Cell Biol. 2019 Feb 26. pii: jcb.201808119. [Epub ahead of print]
    Hariri H, Speer N, Bowerman J, Rogers S, Fu G, Reetz E, Datta S, Feathers JR, Ugrankar R, Nicastro D, Henne WM.
      Lipid droplets (LDs) serve as cytoplasmic reservoirs for energy-rich fatty acids (FAs) stored in the form of triacylglycerides (TAGs). During nutrient stress, yeast LDs cluster adjacent to the vacuole/lysosome, but how this LD accumulation is coordinated remains poorly understood. The ER protein Mdm1 is a molecular tether that plays a role in clustering LDs during nutrient depletion, but its mechanism of function remains unknown. Here, we show that Mdm1 associates with LDs through its hydrophobic N-terminal region, which is sufficient to demarcate sites for LD budding. Mdm1 binds FAs via its Phox-associated domain and coenriches with fatty acyl-coenzyme A ligase Faa1 at LD bud sites. Consistent with this, loss of MDM1 perturbs free FA activation and Dga1-dependent synthesis of TAGs, elevating the cellular FA level, which perturbs ER morphology and sensitizes yeast to FA-induced lipotoxicity. We propose that Mdm1 coordinates FA activation adjacent to the vacuole to promote LD production in response to stress, thus maintaining ER homeostasis.
  12. Curr Alzheimer Res. 2019 Feb 28.
    Rozpędek W, Pytel D, Popławski T, Walczak A, Gradzik K, Wawrzynkiewicz A, Wojtczak R, Mucha B, Diehl JA, Majsterek I.
      There is a body of evidence that neurodegenerative disease entities are directly correlated with the perturbations on the molecular level. Hence, the ER stress-mediated Unfolded Protein Response (UPR) is activated resulting in PERK-dependent phosphorylation of the Eukaryotic initiation factor 2 (eIF2α). Thus, the levels of ATF4 and CHOP proteins are significantly increased, which subsequently switches the pro-adaptive branch of the UPR into the pro-apoptotic directly leading to neuronal loss and initiation of the neurodegenerative process. The aim of the presented study was the evaluation of the biological activity of highly specific, small-molecule inhibitors of the PERK-dependent UPR signaling pathway. The study was conducted on rat astrocytic DI TNC1 cell line. The level of p-eIF2α was measured by Western blot technique, the cytotoxicity of the investigated compound was assessed by the MTT assay and using the FITC-conjugated Annexin V (Annexin V-FITC) to indicate apoptosis and propidium iodide (PI) to indicate necrosis. The effect of tested compound on cell cycle progression was measured by flow cytometry, where the PI-labelled nuclei were analysed for DNA content. As a result one of the investigated compound LDN-0060609 triggers a significant inhibition of the eIF2α phosphorylation in DI TNC1 cell line. Moreover, we showed that compound LDN-0060609 is non-cytotoxic and has no effect on cell cycle progression. In conclusion, LDN-0060609 may constitute a novel, targeted treatment approach against neurodegenerative diseases, including Alzheimer's disease (AD), which pathogenesis and progression are closely associated with the overactivation of the PERK-dependent UPR signaling pathway.
  13. Sci Rep. 2019 Feb 26. 9(1): 2794
    Scussolini M, Bauckneht M, Cossu V, Bruno S, Orengo AM, Piccioli P, Capitanio S, Yosifov N, Ravera S, Morbelli S, Piana M, Sambuceti G, Caviglia G, Marini C.
      The favourable kinetics of 18F-fluoro-2-deoxyglucose (FDG) permits to depict cancer glucose consumption by a single evaluation of late tracer uptake. This standard procedure relies on the slow radioactivity loss, usually attributed to the limited tumour expression of G6P-phosphatase (G6Pase). However, this classical interpretation intrinsically represents an approximation since, as in all tissues, cancer G6Pase activity is remarkable and is confined to the endoplasmic reticulum (ER), whose lumen must be reached by phosphorylated FDG to explain its hydrolysis and radioactivity release. The present study tested the impact of G6Pase sequestration on the mathematical description of FDG trafficking and handling in cultured cancer cells. Our data show that accounting for tracer access to the ER configures this compartment as the preferential site of FDG accumulation. This is confirmed by the reticular localization of fluorescent FDG analogues. Remarkably enough, reticular accumulation rate of FDG is dependent upon extracellular glucose availability, thus configuring the same ER as a significant determinant of cancer glucose metabolism.
  14. Nat Cell Biol. 2019 Mar;21(3): 384-396
    Şentürk M, Lin G, Zuo Z, Mao D, Watson E, Mikos AG, Bellen HJ.
      Although the aetiology of amyotrophic lateral sclerosis (ALS) remains poorly understood, impaired proteostasis is a common feature of different forms of ALS. Mutations in genes encoding ubiquilins, UBQLN2 and UBQLN4, cause familial ALS. The role of ubiquilins in proteasomal degradation is well established, but their role in autophagy-lysosomal clearance is poorly defined. Here, we describe a crosstalk between endoplasmic reticulum stress, mTOR signalling and autophagic flux in Drosophila and mammalian cells lacking ubiquilins. We found that loss of ubiquilins leads to endoplasmic reticulum stress, impairs mTORC1 activity, promotes autophagy and causes the demise of neurons. We show that ubiquilin mutants display defective autophagic flux due to reduced lysosome acidification. Ubiquilins are required to maintain proper levels of the V0a/V100 subunit of the vacuolar H+-ATPase and lysosomal pH. Feeding flies acidic nanoparticles alleviates defective autophagic flux in ubiquilin mutants. Hence, our studies reveal a conserved role for ubiquilins as regulators of autophagy by controlling vacuolar H+-ATPase activity and mTOR signalling.
  15. Front Physiol. 2019 ;10 70
    Maamoun H, Benameur T, Pintus G, Munusamy S, Agouni A.
      Type-2 diabetes prevalence is continuing to rise worldwide due to physical inactivity and obesity epidemic. Diabetes and fluctuations of blood sugar are related to multiple micro- and macrovascular complications, that are attributed to oxidative stress, endoplasmic reticulum (ER) activation and inflammatory processes, which lead to endothelial dysfunction characterized, among other features, by reduced availability of nitric oxide (NO) and aberrant angiogenic capacity. Several enzymatic anti-oxidant and anti-inflammatory agents have been found to play protective roles against oxidative stress and its downstream signaling pathways. Of particular interest, heme oxygenase (HO) isoforms, specifically HO-1, have attracted much attention as major cytoprotective players in conditions associated with inflammation and oxidative stress. HO operates as a key rate-limiting enzyme in the process of degradation of the iron-containing molecule, heme, yielding the following byproducts: carbon monoxide (CO), iron, and biliverdin. Because HO-1 induction was linked to pro-oxidant states, it has been regarded as a marker of oxidative stress; however, accumulating evidence has established multiple cytoprotective roles of the enzyme in metabolic and cardiovascular disorders. The cytoprotective effects of HO-1 depend on several cellular mechanisms including the generation of bilirubin, an anti-oxidant molecule, from the degradation of heme; the induction of ferritin, a strong chelator of free iron; and the release of CO, that displays multiple anti-inflammatory and anti-apoptotic actions. The current review article describes the major molecular mechanisms contributing to endothelial dysfunction and altered angiogenesis in diabetes with a special focus on the interplay between oxidative stress and ER stress response. The review summarizes the key cytoprotective roles of HO-1 against hyperglycemia-induced endothelial dysfunction and aberrant angiogenesis and discusses the major underlying cellular mechanisms associated with its protective effects.
    Keywords:  ER stress; angiogenesis; diabetes; endothelial dysfunction; heme oxygenase-1 (Ho-1); hyperglycemia; oxidative stress
  16. Nat Commun. 2019 Feb 28. 10(1): 984
    Pain C, Kriechbaumer V, Kittelmann M, Hawes C, Fricker M.
      The endoplasmic reticulum (ER) is a highly dynamic polygonal membrane network composed of interconnected tubules and sheets (cisternae) that forms the first compartment in the secretory pathway involved in protein translocation, folding, glycosylation, quality control, lipid synthesis, calcium signalling, and metabolon formation. Despite its central role in this plethora of biosynthetic, metabolic and physiological processes, there is little quantitative information on ER structure, morphology or dynamics. Here we describe a software package (AnalyzER) to automatically extract ER tubules and cisternae from multi-dimensional fluorescence images of plant ER. The structure, topology, protein-localisation patterns, and dynamics are automatically quantified using spatial, intensity and graph-theoretic metrics. We validate the method against manually-traced ground-truth networks, and calibrate the sub-resolution width estimates against ER profiles identified in serial block-face SEM images. We apply the approach to quantify the effects on ER morphology of drug treatments, abiotic stress and over-expression of ER tubule-shaping and cisternal-modifying proteins.
  17. Int J Oncol. 2019 Feb 28.
    Chaiyawat P, Sungngam P, Teeyakasem P, Sirikaew N, Klangjorhor J, Settakorn J, Diskul-Na-Ayudthaya P, Chokchaichamnankit D, Srisomsap C, Svasti J, Pruksakorn D.
      Oncogenic drivers of osteosarcoma remain controversial due to the complexity of the genomic background of the disease. There are limited novel therapeutic options, and the survival rate of patients with osteosarcoma has not improved in decades. Genomic instability leads to complexity in various pathways, which is potentially revealed at the protein level. Therefore, the present study aimed to identify the mechanisms involved in the oncogenesis of osteosarcoma using proteomics and bioinformatics tools. As clinical specimens from patients are the most relevant disease‑related source, expression patterns of proteins in osteosarcoma tissues were compared with soft tissue callus from donors containing high numbers of osteoblastic cells. Two‑dimensional electrophoresis and liquid chromatography‑tandem mass spectrometry (LC‑MS/MS) successfully identified 33 differentially expressed proteins in the osteosarcoma tissues compared with the soft tissue callus. Among these proteins, 29 proteins were significantly upregulated in osteosarcoma. A functionally grouped network of the overexpressed proteins, that was created using the ClueGo and CluePedia applications, demonstrated that the unfolded protein response (UPR) pathway was activated mainly through the activating transcription factor 6 arm in osteosarcoma. The results of proteomics analysis were confirmed by elevated expression of UPR‑related chaperone proteins, including 78 kDa glucose‑related protein (GRP78), endoplasmin, calreticulin and prelamin‑A/C, in the patient‑derived primary cells and osteosarcoma cell lines. Furthermore, the expression of GRP78, a master regulator of the UPR, was enhanced in the osteosarcoma tissues of patients that were resistant to double regimen of doxorubicin and a platinum‑based drug. The findings of the present study suggest that targeting the UPR pathway may be promising for the treatment of osteosarcoma.