Depletion of γ-glutamylcyclotransferase inhibits breast cancer cell growth via cellular senescence induction mediated by CDK inhibitor upregulation

Abstract

BackgroundChromosome 7 open reading frame 24 (C7orf24) was originally identified as a highly expressed protein in various types of cancer, and later shown to be a γ-glutamylcyclotransferase (GGCT). GGCT depletion in cancer cells has anti-proliferative effects in vitro and in vivo, and it is therefore considered a promising candidate as a therapeutic target. However, the cellular events induced by GGCT depletion remain unclear.MethodsGGCT was depleted by siRNA in MCF7, MDA-MB-231, PC3, A172, Hela, and LNCaP cells. Induction of cellular senescence was evaluated with senescence-associated β-galactosidase (SA-β-Gal) staining. Expression levels of p21WAF1/CIP1 and p16INK4A were assessed by qRT-PCR and Western blotting. Effects of simultaneous double knockdown of p21WAF1/CIP1 and p16INK4A together with GGCT on cell cycle regulation and cell growth was measured by flow cytometry, and trypan blue dye exclusion test.ResultsWe found that GGCT knockdown induces significant cellular senescence in various cancer cells. Cyclin dependent kinase inhibitor p21WAF1/CIP1 and/or p16INK4A were upregulated in all cell lines tested. Simultaneous knockdown of p21WAF1/CIP1 recovered the cell cycle arrest, attenuated cellular senescence induction, and rescued the subsequent growth inhibition in GGCT-silenced MCF7 breast cancer cells. In contrast, in GGCT silenced MDA-MB-231 breast cancer cells, GGCT depletion upregulated p16INK4A, which played a regulatory role in senescence induction, instead of p21WAF1/CIP1.ConclusionsOur findings demonstrate that induction of cellular senescence mediated by the upregulation of cyclin-dependent kinase inhibitors is a major event underlying the anti-proliferative effect of GGCT depletion in breast cancer cells, highlighting the potential of GGCT blockade as a therapeutic strategy to induce cellular senescence.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-016-2779-y) contains supplementary material, which is available to authorized users.

Mechanisms of Tumor Growth Inhibition by Depletion of γ-Glutamylcyclotransferase (GGCT): A Novel Molecular Target for Anticancer Therapy

“…NF-Y and Sp1 bind to the CCAAT boxes and to the GC box, respectively, to positively regulate transcription of the GGCT gene. Since promoters with several NF-Y-binding CCAAT boxes were found in genes related to the cell cycle, Ohno et al suggested that GGCT may also play a role in the cell cycle [ 18 ], as confirmed by other studies [ 19 , 20 , 21 , 22 ].…”

“…Dong et al also reported that GGCT knockdown blocked cell cycle progression at the G0/G1 phase in both HCT116 and in SW1116 colorectal cancer cells [ 21 ]. Matsumura et al observed G0/G1 arrest in MCF7 breast cancer cells [ 22 ]. In contrast, Zhang et al analyzed cell cycle change using a gastric cancer cell line, MGC80-3, and reported a significant reduction of cell cycle progression in shGGCT cells, with a reduction of the cell population in the G0/G1 phase and a corresponding increase in the G2/M phase when compared with the control cells [ 20 ].…”

“…In contrast, Li et al reported no change in cell apoptosis after interference of GGCT expression [ 16 ]. Matsumura et al showed that apoptotic signaling molecules such as caspases and PARP were not activated by GGCT knockdown in MCF7 cells [ 22 ]. Therefore, further studies are needed to clarify the precise cell death mechanism by GGCT depletion by considering the mutation background of specific cell types.…”

Structural and Functional Analyses of Human ChaC2 in Glutathione Metabolism

“…Recently, it has been reported that the downregulation of GGCT elicits an anti-proliferative effect in some cancer cell lines, such as breast cancer cell lines MCF-7, MCF-7/ADR, and MDA-MB-231 [44][45][46]. Interestingly, ChaC1 was suggested as a novel biomarker because the overexpression of mammalian ChaC1 and its related transcript variants has been previously found to promote cell proliferation in breast cancer cell lines Hs578T and BT-20 [18,47,48].…”

Interleukin-22 derived from cervical cancer-associated fibroblasts accelerates senescence of normal fibroblasts and promotes expression of tumorigenesis-related factors in HeLa cells

“…Senescence is a critical and irreversible cellular program characterized by changes in the behavior, function, and morphology of cells and tissues [27]. Various stimuli, such as oncogene activation, DNA lesion, cytotoxic stimulation, and inflammatory stress, can lead to senescence.…”