Nuclear Translocation of Glutaminase GLS2 in Human Cancer Cells Associates with Proliferation Arrest and Differentiation

Abstract

Glutaminase (GA) catalyzes the first step in mitochondrial glutaminolysis playing a key role in cancer metabolic reprogramming. Humans express two types of GA isoforms: GLS and GLS2. GLS isozymes have been consistently related to cell proliferation, but the role of GLS2 in cancer remains poorly understood. GLS2 is repressed in many tumor cells and a better understanding of its function in tumorigenesis may further the development of new therapeutic approaches. We analyzed GLS2 expression in HCC, GBM and neuroblastoma cells, as well as in monkey COS-7 cells. We studied GLS2 expression after induction of differentiation with phorbol ester (PMA) and transduction with the fulllength cDNA of GLS2. In parallel, we investigated cell cycle progression and levels of p53, p21 and c-Myc proteins. Using the baculovirus system, human GLS2 protein was overexpressed, purified and analyzed for posttranslational modifications employing a proteomics LC-MS/MS platform. We have demonstrated a dual targeting of GLS2 in human cancer cells. Immunocytochemistry and subcellular fractionation gave consistent results demonstrating nuclear and mitochondrial locations, with the latter being predominant. Nuclear targeting was confirmed in cancer cells overexpressing c-Myc-and GFP-tagged GLS2 proteins. We assessed the subnuclear location finding a widespread distribution of GLS2 in the nucleoplasm without clear overlapping with specific nuclear substructures. GLS2 expression and nuclear accrual notably increased by treatment of SH-SY5Y cells with PMA and it correlated with cell cycle arrest at G2/M, upregulation of tumor suppressor p53 and p21 protein. A similar response was obtained by overexpression of GLS2 in T98G glioma cells, including downregulation of oncogene c-Myc. Furthermore, human GLS2 was identified as being hypusinated by MS analysis, a posttranslational modification which may be relevant for its nuclear targeting and/or function. Our studies provide evidence for a tumor suppressor role of GLS2 in certain types of cancer. The data imply that GLS2 can be regarded as a highly mobile and multilocalizing protein translocated to both mitochondria and nuclei. Upregulation of GLS2 in cancer cells induced an antiproliferative response with cell cycle arrest at the G2/M phase.Altered metabolism is a hallmark of cancer 1 and the term "metabolic reprogramming" has been coined to describe the whole range of metabolic abnormalities accompanying tumorigenesis and metastasis 2 . Increased

Glutaminase 2 functions as a tumor suppressor gene in gastric cancer

“…In the current study, we found that the expression of GLS2 mRNA in the tissues of gastric cancer was significantly lower than in adjacent non-cancer tissue, suggesting that GLS2 downregulation might be associated with the development and progression of gastric cancer. Our data were consistent with the previous reports that GLS2 was repressed in many tumor cells, such as HCCs and glioblastomas ( 16 , 30 ). Szeliga et al demonstrated that GLS2 displayed a significant downregulation in glioblastoma due to DNA demethylation, while CpG methylation was absent in GLS2-expressing adjacent non-cancer brain tissues ( 20 ).…”

A Novel Ferroptosis-Related Gene Signature Robustly Predicts Clinical Prognosis And Tumor Immunity in Patients With Kidney Renal Clear Cell Carcinoma

“…It is reported that miR-9 can suppress ferroptosis via downregulating GOT1 expression in melanoma (46). In cancer cells, GLS2 upregulation can induce an antiproliferative response with cell cycle arrest (47). Notably, high expression of risk genes, such as CD44, FANCD2 and CHAC1 as well as low expression of NCOA4 were found to be correlated with worse OS in KIRC (21,22), in line with our results.…”

Impact of the acidic environment on gene expression and functional parameters of tumors in vitro and in vivo

“…Several genes found to be regulated by acidosis after 24 h has been described to affect functional parameter of tumor progression, such as proliferation (e.g., Gls2 [ 37 ]), cell death (e.g., Txnip, Gls2 [ 38 , 39 ] tumor cell migration (e.g., Per3, Ikbke, Txnip [ 40 – 42 ] and adhesion (e.g., Txnip [ 43 ]) or mitochondrial activity (e.g., Txnip [ 44 ]). Therefore the impact of acidification on these cell functions was analyzed in the tumor models.…”