Gene expression in gamma-glutamylcyclotransferase-overexpressing mouse embryonic fibroblast NIH3T3 cells.

Although hypoxia inducible factor-1a (HIF-1a) was originally identified as a transcriptional factor to adapt hypoxia, HIF-1a has also been shown to regulate metabolic pathways specific for cancer, including aerobic glycolysis, namely “Warburg effect”. However, the mechanisms by which HIF-1a mediates the metabolic pathway under normoxia are not fully understood. Here, we identified gamma-glutamylcyclotransferase (GGCT) as a novel regulator of HIF-1a. GGCT is a highly expressed protein in various cancer tissues. Knockdown of GGCT exerts anticancer effects both in vitro and in vivo; thus, it is considered a promising therapeutic target. In this study, we show that depleting GGCT downregulates the protein and mRNA expressions of HIF-1a in PC3 prostate cancer cells, and that overexpressing GGCT upregulates them in mouse fibroblast NIH3T3 cells. We also show that depleting GGCT downregulates HIF-1a downstream target genes involved in glycolysis, whereas overexpressing GGCT exhibits upregulation pattern in these genes. Furthermore, we suggest that GGCT induces the metabolic switch from citric acid cycle to glycolysis under normoxia. Additionally, we show that GGCT regulates expression of HIF-1a protein via AMPK-mTORC1-4EBP1 pathway in PC3 cells. These results indicate the GGCT plays a regulatory role in the expression of HIF-1a followed by glycolytic switch in cancer cells. To test the effect of overexpression of GGCT on the gene expression, mouse embryonic fibroblast cell line NIH3T3 was transfected with GGCT expression retroviral vector or control vector. Three independent experiments were performed and GGCT-overexpressing NIH3T3 cells were compared to control cells.