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We have studied the effect of glutamine deprivation on the expression of genes encoding for ubiquitin specific peptidases (USP) and ubiquitin activating enzyme E1-like protein/autophagy related 7 (GSA7/ATG7) in U87 glioma cells in relation to inhibition of inositol requiring enzyme-1 (IRE1). It was shown that exposure of control glioma cells (transfected by empty vector) upon glutamine deprivation led to suppression of USP1 and ATG7 mRNA expression and up-regulated USP25 mRNA. At the same time, glutamine deprivation did not significantly change USP4, USP10, USP14, and USP22 gene expressions in these cells. Inhibition of ІRE1 signaling enzyme function in U87 glioma cells increased effect of glutamine deprivation on the expression of USP1 gene and introduced sensitivity of USP4 and USP14 genes to this condition. Therefore, glutamine deprivation affected the expression level of most studied genes in gene specific manner in relation to the functional activity of IRE1 enzyme, a central mediator of endoplasmic reticulum stress, which controls cell proliferation and tumor growth.

We have studied the effect of hypoxia on the expression level of mRNA of the basic enzymes of pentose-phosphate cycle (G6PD, TKT, TALDO1, PGLS and RPIA) and glucose-6-phosphate isomerase (GPI) in U87 glioma cells in relation to inhibition of IRE1 (inositol requiring enzyme 1). It was shown that hypoxia leads to up-regulation of the expression of GPI and PGLS genes and to down-regulation of TALDO1 and RPIA genes in control glioma cells. Changes for GPI gene were more significant than for other genes. At the same time, inhibition of IRE1 modified the effect of hypoxia on the expression of all studied genes. In particular, it increased sensitivity to hypoxia of G6PD and TKT genes expression and suppressed the effect of hypoxia on the expression of GPI and RPIA genes. Additionally, inhibition of IRE1 eliminated hypoxic regulation of PGLS gene and did not change significantly effect of hypoxia on the expression of TALDO1 gene in glioma cells. Present study demonstrated that hypoxia, which often contributes to tumor growth, affects the expression of most studied genes and inhibition of IRE1 modified the hypoxic regulation of pentose-phosphate cycle gene expressions in a gene specific manner and thus possibly contributes to slower glioma growth, but several aspects of this regulation warrant further investigation.

Objective. The aim of the present study was to examine the effect of inhibition of endoplasmic reticulum stress signaling, mediated by IRE1 (inositol requiring enzyme 1), which is a central mediator of the unfolded protein response on the expression of genes encoded estrogen related proteins (NRIP1/RIP140, TRIM16/EBBP, ESRRA/NR3B1, FAM162A/E2IG5, PGRMC2/PMBP, and SLC39A6/LIV-1) and their hypoxic regulation in U87 glioma cells for evaluation of their possible significance in the control of glioma cells proliferation.

To study the effect of glucose deprivation on the expression of genes encoding for LONP1/PRSS15 and cathepsins in U87 glioma cells in relation to inhibition of inositol requiring enzyme-1 (IRE1) was the aim of the research. It was shown that glucose deprivation up-regulated the expression of CTSA, CTSB, CTSD, CTSK, CTSL, CTSO, and LONP1 genes and did not change the expression of CTSC, CTSF, and CTSS genes in control glioma cells (transfected by empty vector). Inhibition of ІRE1 signaling enzyme function in U87 glioma cells modified effect of glucose deprivation on the expression of most studied genes: removed the effect of glucose deprivation on CTSA and CTSO genes, introduces on CTSC and CTSS genes, reduced – on CTSK gene, and enhanced – on CTSL gene. Therefore, glucose deprivation affect the expression level of most studied genes in relation to the functional activity of IRE1 enzyme, a central mediator of endoplasmic reticulum stress, which control cell proliferation and tumor growth.

We investigated the impact of IRE1/ERN1 (inositol requiring enzyme 1/endoplasmic reticulum to nucleus signaling 1) knockdown on hypoxic regulation of the expression of a subset of proliferation and migration-related genes in U87 glioma cells. It was shown that hypoxia leads to up-regulation of the expression of MEST and SNAI2, to down-regulation – of MYBL1, TCF8 and GTF2F2 genes at the mRNA level in control glioma cells. At the same time hypoxia did not affect the expression of TCF3 and GTF2B transcription factor genes. In turn, inhibition of IRE1 modified the effect of hypoxia on the expression of all studied genes, except MYBL1 and GTF2B. For instance, IRE1 knockdown decreased sensitivity to hypoxia of the expression of MEST, TCF8 and SNAI2 genes and increased sensitivity to hypoxia of GTF2F2 expression. At the same time, IRE1 inhibition introduced sensitivity to hypoxia of the expression of TCF3 gene in glioma cells. The present study demonstrated that the inhibition of IRE1 in glioma cells affected the hypoxic regulation of the expression of studied genes in various directions, though hypoxic conditions did not abolish the effect of IRE1 inhibition on the expression of respective genes. To the contrary, in case of SNAI2, GTF2F2 and MEST hypoxic conditions magnified the effect of IRE1 inhibition on the expression of respective genes in glioma cells.

We have studied the effect of glucose deprivation on the expression of genes encoding for ubiquitin specific peptidases — USP and autophagy related 7 ATG7 in U87 glioma cells in relation to inhibition of inositol requiring enzyme-1 (IRE1). It was shown that glucose deprivation was downregulated the expression of USP1 and USP10 genes and up-regulated USP4 and USP25 genes in control (transfected by empty vector) glioma cells. At the same time, the expression level of USP14, USP22, and ATG7 genes in these cells did not significantly change upon glucose deprivation condition. Inhibition of ІRE1 signaling enzyme function in U87 glioma cells modified effect of glucose deprivation on the expression of most studied genes. Therefore, glucose deprivation affected the expression level of most ubiquitin specific peptidases genes in relation to the functional activity of IRE1 enzyme, which controls cell proliferation and tumor growth as a central mediator of endoplasmic reticulum stress.

Tumor necrosis factor (TNF) superfamily receptors and TNF apoptosis inducing ligands play an important role in the realization of TNF function and control tumor growth. The TNF-related pathways are controlled by endoplasmic reticulum stress signaling, which has a crucial role in the control of cell proliferation and tumor growth. Furthermore, the inhibition of IRE1 (inositol requiring enzyme-1), which is a central mediator of endoplasmic reticulum stress sand mainly responsible for cell proliferation and apoptosis, leads to suppression of tumor growth through specific changes in the expression of genes encoding transcription factors, tumor suppressors, angiogenesis and apoptosis related proteins, including TNF superfamily receptors and TNF apoptosis inducing ligands. Therefore, changes in the expression level of TNF-related genes encoding TNF superfamily receptors and apoptosis inducing ligands possibly reflect metabolic reprogramming of cancer cells upon inhibition of IRE1-mediated endoplasmic reticulum stress signaling and correlate with suppression of glioma cell proliferation.

Objective. The aim of the present investigation was to examine the effect of inhibition of endoplasmic reticulum stress signaling, mediated by IRE1 (inositol requiring enzyme 1), which is a central mediator of the unfolded protein response on the expression of genes encoding glucocorticoid receptor (NR3C1) and some related proteins (SGK1, SGK3, NCOA1, NCOA2, ARHGAP35, NNT) and their hypoxic regulation in U87 glioma cells for evaluation of their possible significance in the control of the glioma growth.

Objective. The aim of the present study was to investigate the effect of hypoxia on the expression of genes encoding insulin-like growth factors (IGF1 and IGF2), their receptor (IGF1R), binding protein-4 (IGFBP4), and stanniocalcin 2 (STC2) in U87 glioma cells in relation to inhibition of endoplasmic reticulum stress signaling mediated by IRE1 (inositol requiring enzyme 1) for evaluation of their possible significance in the control of tumor growth.

We have studied the glucose and glutamine deprivation effect on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells in relation to inhibition of inositol requiring enzyme-1 (IRE1). It was shown that glutamine deprivation down-regulated the expression of mitochondrial (NADP+)-dependent isocitrate dehydrogenase 2 (IDH2), malic enzyme 2 (ME2), mitochondrial aspartate aminotransferase (GOT2), and subunit B of succinate dehydrogenase (SDHB) genes in control glioma cells in gene specific manner. At the same time, the expression level of malate dehydrogenase 2 (MDH2) and subunit D of succinate dehydrogenase (SDHD) genes in these cells was not changed upon glutamine deprivation. It was also shown that inhibition of ІRE1 signaling enzyme function in U87 glioma cells modified the glutamine deprivation effect on the expression of all studied genes. Furthermore, the expression of the majority of studied genes was resistant to glucose deprivation, except IDH2 and SDHB genes, which expression levels were slightly down-regulated. Inhibition of IRE1 modified the effect of glucose deprivation on ME2, SDHB, SDHD, and GOT2 genes expression. Therefore, glucose and glutamine deprivation affected the expression level of the majority of nuclear genes encoding mitochondrial proteins in relation to the functional activity of IRE1 enzyme, which is a central mediator of endoplasmic reticulum stress and controls cell proliferation and tumor growth.

We have studied the expression of genes encoding proliferation related factors and enzymes such as IL13RA2, KRT18, CD24, ING1, ING2, MYL9, BET1, TRAPPC3, ENDOG, POLG, TSFM, and MTIF2 in U87 glioma cells upon glucose and glutamine deprivation in relation to inhibition of inositol requiring enzyme 1, a central mediator of endoplasmic reticulum stress. It was shown that glutamine deprivation leads to up-regulation of the expression of BET1, MYL9, and MTIF2 genes and down-regulation of CD24, ING2, ENDOG, POLG, and TSFM genes in control (with native IRE1) glioma cells. At the same time, glucose deprivation enhances the expression of MYL9 gene only and decreases – ING1, ING2, and MTIF2 genes in control glioma cells. Thus, effect of glucose and glutamine deprivation on gene expressions in glioma cells is gene-specific. Inhibition of inositol requiring enzyme 1 by dnIRE1 significantly modifies the effect of both glutamine and glucose deprivation on the expression of most studied genes with different direction and magnitude, especially for ING2, CD24, and MTIF2 genes. Present study demonstrates that IRE1 knockdown modifies glucose and glutamine deprivation effects on the expression of proliferation related genes and possibly contributes to slower tumor growth of these glioma cells after inhibition of IRE1 signaling enzyme.

We have studied hypoxic regulation of the expression of different insulin-like growth factor binding­ protein genes in U87 glioma cells in relation to inhibition of IRE1 (inositol requiring enzyme-1), a central mediator of endoplasmic reticulum stress, which controls cell proliferation and tumor growth. We have de­monstrated that hypoxia leads to up-regulation of the expression of IGFBP6, IGFBP7, IGFBP10/CYR61, WISP1, and WISP2 genes and down-regulation – of IGFBP9/NOV gene at the mRNA level in control glioma cells, being more significant changes for IGFBP10/CYR61 and WISP2 genes. At the same time, inhibition of IRE1 modifies the effect of hypoxia on the expression of all studied genes: eliminates sensitivity to hypoxia the expression of IGFBP7 and IGFBP9/NOV genes, suppresses effect of hypoxia on IGFBP6, IGFBP10/CYR61, and WISP2 genes, and slightly enhances hypoxic regulation of WISP1 gene expression in glioma cells. We have also demonstrated that the expression of all studied genes in glioma cells is regulated by IRE1 signaling enzyme upon normoxic condition, because inhibition of IRE1 significantly up-regulates IGFBP7, IGFBP10/CYR61, WISP1, and WISP2 genes and down-regulates IGFBP6 and IGFBP9/NOV genes as compared to control glioma cells. The present study demonstrates that hypoxia, which contributes to tumor growth, affects all studied IGFBP and WISP gene expressions and that inhibition of IRE1 preferentially abolishes or suppresses the hypoxic regulation of these gene expressions and thus possibly contributes to slower glioma growth. Moreover, inhibition of IRE1, which correlates with suppression of cell proliferation and glioma growth, is down-regulated expression of pro-proliferative IGFBP genes, attesting to the fact that endoplasmic reticulum stress is a necessary component of malignant tumor growth.

Inhibition of IRE1 (inositol requiring enzyme-1), the major signaling pathway of endoplasmic reticulum stress, significantly decreases glioma cell proliferation and tumor growth. We have studied the expression of TNFα-related genes and effect of glucose deprivation on these gene expressions in U87 glioma cells overexpressing dominant-negative IRE1 defective in both kinase and endonuclease (dn-IRE1) activity of IRE1 with hopes of elucidating its contribution to IRE1 mediated glioma growth. We have demonstrated that glucose deprivation condition leads to down-regulation of the expression of TNFRSF11B, TNFRSF1A, TNFRSF10D/TRAILR4, and LITAF genes and up-regulation of TNFRSF10B/TRAILR2/DR5 gene at the mRNA level in control glioma cells. At the same time, the expression of TNFRSF21/DR6, TNFAIP1, TNFAIP3, TRADD, and CD70/TNFSF7 genes in control glioma cells is resistant to glucose deprivation condition. The inhibition of IRE1 modifies the effect of glucose deprivation on LITAF, TNFRSF21, TNFRSF11B, and TRADD gene expressions and induces sensitivity to glucose deprivation condition the expression of TNFRSF10B, TNFRSF1A, and CD70 genes. We have also demonstrated that the expression of all studied genes is affected in glioma cells by inhibition of IRE1, except TNFRSF1A gene, as compared to control glioma cells. Moreover, the changes in the expression of TNFRSF1A, TNFRSF10D/TRAILR4, and LITAF genes induced by glucose deprivation condition have opposite orientation to that induced by inhibition of IRE1. The present study demonstrates that fine-tuning of the expression of TNFα-induced proteins and TNF receptor superfamily genes, which related to cell death and proliferation, is regulated by IRE1, an effector of endoplasmic reticulum stress, as well as depends on glucose deprivation in gene specific manner. Thus, the inhibition of kinase and endoribonuclease activity of IRE1 correlates with deregulation of TNFα-induced protein genes and TNF receptor superfamily genes in gene specific manner and thus slower the tumor growth.

We have studied the glucose and glutamine deprivation effect on the expression of nuclear genes encoding mitochondrial proteins in U87 glioma cells in relation to inhibition of inositol requiring enzyme-1 (IRE1). It was shown that glutamine deprivation down-regulated the expression of mitochondrial (NADP+)-dependent isocitrate dehydrogenase 2 (IDH2), malic enzyme 2 (ME2), mitochondrial aspartate aminotransferase (GOT2), and subunit B of succinate dehydrogenase (SDHB) genes in control glioma cells in gene specific manner. At the same time, the expression level of malate dehydrogenase 2 (MDH2) and subunit D of succinate dehydrogenase (SDHD) genes in these cells was not changed upon glutamine deprivation. It was also shown that inhibition of ІRE1 signaling enzyme function in U87 glioma cells modified the glutamine deprivation effect on the expression of all studied genes. Furthermore, the expression of the majority of studied genes was resistant to glucose deprivation, except IDH2 and SDHB genes, which expression levels were slightly down-regulated. Inhibition of IRE1 modified the effect of glucose deprivation on ME2, SDHB, SDHD, and GOT2 genes expression. Therefore, glucose and glutamine deprivation affected the expression level of the majority of nuclear genes encoding mitochondrial proteins in relation to the functional activity of IRE1 enzyme, which is a central mediator of endoplasmic reticulum stress and controls cell proliferation and tumor growth. [ABSTRACT FROM AUTHOR]

We have studied the expression of genes encoding proliferation related factors and enzymes such as IL13RA2, KRT18, CD24, ING1, ING2, MYL9, BET1, TRAPPC3, ENDOG, POLG, TSFM , and MTIF2 in U87 glioma cells upon glucose and glutamine deprivation in relation to inhibition of inositol requiring enzyme 1, a central mediator of endoplasmic reticulum stress. It was shown that glutamine deprivation leads to up-regulation of the expression of BET1, MYL9 , and MTIF2 genes and down-regulation of CD24, ING2, ENDOG, POLG , and TSFM genes in control (with native IRE1) glioma cells. At the same time, glucose deprivation enhances the expression of MYL9 gene only and decreases - ING1, ING2, and MTIF2 genes in control glioma cells. Thus, effect of glucose and glutamine deprivation on gene expressions in glioma cells is gene-specific. Inhibition of inositol requiring enzyme 1 by dnIREI significantly modifies the effect of both glutamine and glucose deprivation on the expression of most studied genes with different direction and magnitude, especially for ING2, CD2 4, and MTIF2 genes. Present study demonstrates that IRE1 knockdown modifies glucose and glutamine deprivation effects on the expression of proliferation related genes and possibly contributes to slower tumor growth of these glioma cells after inhibition of IRE1 signaling enzyme. [ABSTRACT FROM AUTHOR]

Objective: The aim of the present study was to investigate the effect of inhibition of endoplasmic reticulum stress signaling mediated by IRE1/ERN1 (inositol-requiring enzyme 1/endoplasmic reticulum to nucleus signaling 1) on the expression of genes encoding different groups of insulin-like growth binding proteins (IGFBP6 and IGFBP7) and CCN family (IGFBP8/CTGF/CCN2, IGFBP9/NOV/CCN3, IGFBP10/CYR61/CCN1, WISP1/CCN4, and WISP2/CCN5) and its sensitivity to glucose deprivation in U87 glioma cells., Methods: The expression of IGFBP6, IGFBP7, IGFBP8, IGFBP9, IGFBP10, WISP1, and WISP2 genes was studied by qPCR in control U87 glioma cells (wild-type) and its subline with IRE1 signaling enzyme loss of function upon glucose deprivation., Results: The expression of IGFBP8, IGFBP9, and WISP2 genes was up-regulated in control glioma cells upon glucose deprivation with most significant changes for IGFBP9 gene. At the same time, the expression of IGFBP6, IGFBP10, and WISP1 genes was resistant to glucose deprivation in these glioma cells, but the IGFBP7 gene expression was down-regulated. The inhibition of both enzymatic activities (kinase and endoribonuclease) of IRE1 in glioma cells modified the sensitivity of most studied gene expressions to glucose deprivation condition: introduced sensitivity of IGFBP10 and WISP1 genes to glucose deprivation, enhanced the effect of this deprivation on IGFBP7 and IGFBP9 gene expressions, and reduced this effect on WISP2 gene and induced suppressive effect of glucose deprivation on the expression of IGFBP8 gene. Furthermore, the inhibition of IRE1 strongly affected the expression of all studied genes in glioma cells upon regular growing condition in gene specific manner: up-regulated the expression levels of IGFBP7, IGFBP8, IGFBP10, WISP1, and WISP2 genes and down-regulated the IGFBP6 and IGFBP9 genes., Conclusions: The data of this investigation demonstrate that the expression of IGFBP7, IGFBP8, IGFBP9, and WISP2 genes are sensitive to glucose deprivation in U87 glioma cells and that inhibition of IRE1 signaling enzyme function may significantly affect the expression of all studied genes in the presence of glucose as well as modify the effect of glucose deprivation on the expression of most studied genes. These data also show that proteins encoded by these genes may participate in the regulation of metabolic and proliferative processes via IGF/INS receptors and possibly other signaling pathways as well, via IRE1 signaling, which is a central mediator of the unfolded protein response and an important component of the tumor growth and metabolic diseases.