A bidirectional promoter reporter vector for the analysis of the p53/WDR79 dual regulatory element

Abstract

Analysis of numerous genomes has identified a class of regulatory regions that contain a head-to-head arrangement (5′ to 5′) on opposite strands of DNA. Often these regulatory regions have fewer than 1000 base pairs separating their corresponding transcription start sites and have been termed as being “bidirectional”. This bidirectional arrangement and the divergent gene pairs under the control of these regulatory regions appear to be a common feature within genomes. Establishing methods to study these bidirectional transcriptional promoters, and understanding how they are regulated will allow researchers to gain more insight into the roles that divergent transcription plays in the expression and maintenance of protein coding genes. Recently, the p53 tumor suppressor gene was shown to have a bidirectional gene partner, WDR79. The transcription start sites (TSSs) of human and murine p53 and WDR79 genes are separated by approximately 800 and 930 bp, respectively, in a head-to-head fashion, and fit the criteria of what is designated to be a putative bidirectional regulatory region. However, further testing is needed to demonstrate that the region between these genes contains a functional bidirectional promoter. Here, we have developed a bidirectional reporter vector, termed pLucRLuc, to study the transcriptional output of each promoter. This bidirectional reporter vector will allow researchers to determine the output of transcripts mediated by the bidirectional promoters. By focusing our studies on the transcriptional regulation of p53 and its bidirectional gene partner, WDR79, we hope to elucidate key factors that can control and regulate the expression of the p53 and WDR79 genes. Here, we demonstrate that pLucRLuc is a vector capable of expressing reporter genes under the control of bidirectional promoters in multiple human and murine cell lines and that the regulatory region upstream of the p53 and WDR79 TSSs is a bidirectional promoter controlled by common regulatory factors.