Genomic organization and characterization of the promoter of rat malonyl-CoA decarboxylase gene

Abstract

Malonyl-CoA decarboxylase (MCD) catalyzes the decarboxylation of malonyl-CoA, an elongating agent for fatty acid synthesis and also known as a fuel-sensing mediator. In order to elucidate the genome organization, we isolated a 2020 bp rat MCD cDNA from rat brain cDNA library and isolated the corresponding rat genomic clones from the rat genomic PAC library. Sequencing and comparison of these clones showed that the MCD genome consists of five exons and four introns spanning approximately 17 kb. The proximal upstream region is GC-rich, lacks a TATA box, and contains a variety of putative transcriptional regulatory elements within 2 kb. A major transcriptional initiation site was identified by a primer extension at a site 157 nucleotides upstream of the translational initiation site. To investigate the transcriptional regulation of MCD, a series of 5′-deletion constructs of the 5′-flanking region were generated and cloned upstream from the luciferase reporter gene. By comparing promoter activity in CV-1 cells, we suggest that an area of −15 bp 5′ from the first exon acted as a basal promoter for MCD and that there are positive cis-regulatory elements in the region from −55 to −325 bp and negative regulator elements in the region −1380 to −2240 bp.