Molecular and functional characterization of the ZNF509 in the regulation of cell proliferation

Abstract

Cell proliferation is a process which is precisely controlled by interaction between a number of regulatory proteins. In cancer cells, the aberrant expression of oncogenes and/or tumor suppressor genes causes dramatic changes in regulatory programs controlling cell proliferation. Recently, the POK family of transcription factors was characterized as important oncogenes or tumor suppressors that regulate the expression of cell cycle control genes.In this study, ZNF509, a novel POK family of transcription factors that is induced by p53, was characterized to be a transcriptional activator of the CDKN1A cell cycle arrest gene. It was shown that ZNF509 directly bound to the CDKN1A proximal promoter by EMSA, Oligonucleotide pull-down, and ChIP assays. Interaction of ZNF509 and MIZ-1 activated the transcription of CDKN1A synergistically. This interaction between ZNF509 and MIZ-1 recruited the co-activator p300, and the ZNF509-MIZ-1-p300 complex increased histone Ac-H3 and -H4 levels at the CDKN1A proximal promoter. Knockdown of endogenous ZNF509 expression caused decrease in CDKN1A transcription, and resulted in increasing cell proliferation.ZNF509 not only arrested cell cycle progression but also inhibited apoptosis when cells were exposed to DNA damaging agents. Expression of p53 was increased by etoposide treatment regardless of ZNF509 expression and, in turn, p53 increases PUMA gene expression. Expression of PUMA gene decreased moderately only in the presence of ZNF509. IP, GST pull-down, Oligonucleotide pull-down, and ChIP assays showed that ZNF509 directly interacted with p53 via its zinc-fingers, resulting in inhibition p53 binding to regulatory elements of the PUMA promoter and suppression of PUMA transcription.These results suggest that ZNF509 serves important functions in cell fate decision directly toward cell survival. First, ZNF509 potently activates transcription of CDKN1A through interaction with MIZ-1. Second, ZNF509 inhibits apoptosis by repressing the PUMA gene expression by inhibiting DNA binding of p53 to the PUMA promoter.