Epigenetic silencing of HTATIP2 in glioblastoma enhances nuclear translocation of the DNA-repair protein MPG affecting treatment resistance

Abstract

DNA methylome analysis of glioblastoma (GBM) identified the HIV-1 Tat interactive protein 2 (HTATIP2) gene as aberrantly methylated and silenced. HTATIP2 is a negative regulator of importin β-mediated (KPNB1) cytoplasmic-nuclear translocation of proteins and its deregulation may alter the functionality of cancer relevant nuclear proteins. We propose N-Methylpurine-DNA Glycosylase (MPG), responsible for removing alkylated bases and initiating base excision repair (BER), as a potential GBM relevant candidate. Here we investigated the role of epigenetic silencing of HTATIP2 on the subcellular localization of MPG, and MPG-mediated DNA repair. Induction of HTATIP2 expression in GBM cells lead to a significant shift of predominantly nuclear to cytoplasmic MPG, while depletion of endogenous levels of HTATIP2 resulted in enhanced nuclear MPG localization. We observed exclusion of MPG from the area exhibiting co-localization of HTATIP2 and KPNB1 in proximity to the nuclear membrane, suggesting competition of HTATIP2 with MPG to bind to KPNB1. In accordance, pharmacologic inhibition of KPNB1 similarly induced cytoplasmic retention of MPG as HTATIP2 expression. Reduced nuclear MPG localization, induced by HTATIP2 expression or depletion of MPG, yielded less P-H2AX-positive cells upon treatment with an alkylating agent. This suggested reduced MPG-mediated formation of apurinic/apyrimidinic (AP) sites, leaving behind unrepaired DNA lesions, hence, reflecting a reduced capacity of BER in response to the alkylating agent. Taken together, these results suggest that epigenetic silencing of HTATIP2 may increase nuclear localization of MPG, thereby increasing the capacity of the tumor cells to repair treatment related lesions and eventually contributing to treatment resistance.