Human dCTP pyrophosphatase 1 promotes breast cancer cell growth and stemness through the modulation on 5-methyl-dCTP metabolism and global hypomethylation.

F-F Song,Y Wang,J-Q Wang,P Ji,Y-B Tang,Y Zhang,L-L Xia,L Zhu,G-P Zhao,J Zhang,H-L Ge,Z-M Huang

doi:10.1038/oncsis.2015.10

Abstract

Human DCTPP1 (dCTP pyrophosphatase 1), also known as XTP3-transactivated protein A, belongs to MazG-like nucleoside triphosphate pyrophosphatase (NTP-PPase) superfamily. Being a newly identified pyrophosphatase, its relevance to tumorigenesis and the mechanisms are not well investigated. In the present study, we have confirmed our previous study that DCTPP1 was significantly hyperexpressed in breast cancer and further demonstrated its strong association with tumor progression and poor prognosis in breast cancer. Knockdown of DCTPP1 in breast cancer cell line MCF-7 cells remarkably retarded proliferation and colony formation in vitro. The capacity of mammosphere formation of MCF-7 was suppressed with the silence of DCTPP1, which was consistent with the enhanced mammosphere-forming ability in DCTPP1-overexpressed MDA-MB-231 cells. To further dissect the mechanisms of DCTPP1 in promoting tumor cell growth and stemness maintenance, its biochemical properties and biological functions were investigated. DCTPP1 displayed bioactive form with tetrameric structure similar to other MazG domain-containing pyrophosphatases based on structure simulation. A substrate preference for dCTP and its methylated or halogen-modified derivatives over the other canonical (deoxy-) NTPs was demonstrated from enzymatic assay. This substrate preference was also proved in breast cancer cells that the intracellular 5-methyl-dCTP level increased in DCTPP1-deficient MCF-7 cells but decreased in DCTPP1-overexpressed MDA-MB-231 cells. Moreover, global methylation level was elevated in DCTPP1-knockdown MCF-7 cells or mammosphere-forming MCF-7 cells but decreased significantly in DCTPP1-overexpressed MDA-MB-231 cells and its mammospheres. Our results thus indicated that human DCTPP1 was capable of modulating the concentration of intracellular 5-methyl-dCTP. This in turn affected global methylation, contributing to a known phenomenon of hypomethylation related to the cancer cell growth and stemness maintenance. Our current investigations point to the pathological functions of DCTPP1 overexpression in breast cancer cells with aberrant dCTP metabolism and epigenetic modification.

Highlights

Intrinsic physiological processes such as cell metabolism or extrinsic oxidative damage and pathogen infection[1,2] are prone to generate noncanonical nucleotides
These results indicate that dCTP pyrophosphatase 1 (DCTPP1) is strongly associated with breast cancer progression and might be indicative to evaluate the prognosis in breast cancer
Our findings demonstrated that human DCTPP1 was involved in promoting breast cancer cell proliferation and stemness maintenance, largely through controlling 5-methyl-dCTP metabolism and global DNA hypomethylation

Summary

INTRODUCTION

Intrinsic physiological processes such as cell metabolism or extrinsic oxidative damage and pathogen infection[1,2] are prone to generate noncanonical nucleotides. Its expression level is strongly correlated with histological grades and a poor prognosis.[21] There is evidence of elevated expression of dUTPase in NSCLC cell lines and fresh tumor specimens.[22] the expression of dUTPase in colon cancer is highly variable in quantity and diverse in intracellular localization.[23] Noteworthy, the expression of nucleic dUTPase isoform in normal cells is proliferation associated while dUTPase is mostly expressed in replicating cells of normal tissues.[23] All the above studies indicate that human NTP-PPases expression (Table 1). Our findings demonstrated that human DCTPP1 was involved in promoting breast cancer cell proliferation and stemness maintenance, largely through controlling 5-methyl-dCTP metabolism and global DNA hypomethylation.

RESULTS

DISCUSSION

Findings

MATERIALS AND METHODS