Intergenic region between TATA-box binding protein and proteasome subunit C3 genes of Medaka function as the bidirectional promoter in vitro and in vivo

Abstract

In the genome of eukaryotic organisms, each protein-coding gene has the unique promoter in the 5′-flanking region, and the direction of the promoter is usually controlled unidirectional. In this study, we revealed that the intergenic region between TATA-box binding protein (tbp) and proteasome subunit C3 (psmc3) genes in Medaka functions as bidirectional promoter in vitro and in vivo. The tbp and psmc3 genes were allocated as a head-to-head configuration with a 719bp intergenic region. A comparative analysis of gene arrangement surrounding loci of tbp in vertebrates also illustrated that it was unique in Acanthopterygii lineage. The transcription activities were about 1.2 times for tbp direction and 0.7 times for psmc3 direction against that of SV40 promoter in Medaka fibroblasts, respectively. A dual fluorescent reporter assay directly showed that the bidirectional promoter could express two divergent genes concurrently without disruption of RNA polymerase II elongation. In addition, an analysis of sequential deletion of this promoter suggested that the ETS binding site was necessary for maximum expression of downstream gene, and only the ETS binding site was shared from fish to mammals. In mammals, high correlation with CpG islands was observed in such bidirectional promoters, no association was found in the tbp/psmc3 bidirectional promoter in Medaka. These results suggest that molecular machineries of fish bidirectional promoter may be somehow different from those of mammals but the cis-acting element for binding ETS transcription factors is essential for divergent gene expression.