Tsymbal D. Role of signaling enzyme IRE1 in the regulation of expression of pro- and anti-proliferative genes in glioma cells

Object: mechanisms of IRE1-dependent expression of transcription factors involved in control of cell proliferation, as well as of tumor suppressors, upon hypoxia, glucose or glutamine deficiency in U87 glioma cells. Aim: to identify a possible role of transcription factors and tumor suppressors, involved in control of cell proliferation, in IRE1-dependent tumor growth by investigation of their expression in U87 glioma cells with complete or partial inhibition of IRE1, a key sensory-signaling enzyme of endoplasmic reticulum stress, as well as upon hypoxia, glucose or glutamine deficiency in medium. Methods: cell cultivation, RNA and proteins extraction, spectrophotometric assessment of RNA quantity and quality, cDNA synthesis, polymerase chain reaction, nucleic acid electrophoresis, restriction of plasmid DNA, methods of molecular cloning, liposome transfection of cells, Western blot, luciferase reporter assay, statistical methods of data processing. It was first shown that the expression levels of HOXC6, ATF3, TBX3, TBX2, MAZ, SNAI2/SLUG, TCF3, TCF8, MYBL1, MYBL2, FOXF1, EPAS1, E2F8, MEST, CD24, KRT18, ING1, ING2, IL13RA2 and MYRL2 genes in U87 glioma cells depend on the functional activity of IRE1. Changes in the expression of studied genes fall in line with previously published data about suppressed proliferation of glioma cells with IRE1 knockdown. Detected changes in gene expression may play a key role in IRE1-dependent control of cells’ proliferation. Transcription factors HOXC6 and TBX2, as well as subunit А2 of interleukin 13 receptor are promising targets for further research in field of development of new chemotherapeutic agents for glioma treatment. Obtained results broaden our understanding of molecular mechanisms, which constitute the basis of IRE1-mediated control of cancer cells’ proliferation.