Regulation of the Antisense Beta Myosin Heavy Chain Promoter

PURPOSE: Heart muscle expresses two genes encoding MHC isoforms, alpha and beta, which are arranged in tandem 4.5 Kb apart. Rat hearts typically express a predominance of the alpha MHC isoform and have high intrinsic contractility. Hypothyroid (PTU), hypertensive, and diabetic (STZ) rats express a significantly higher proportion of the beta isoform and thus have a lower rate of tension development, but a greater energy economy of force production. This antithetical expression of the alpha and beta genes is highly coordinated, but the mechanism underlying this regulation is poorly understood. We discovered an antisense (AS) beta RNA transcript that starts in the middle of the 4.5Kb (intergenic spacer, IGS) and extends upstream to the beta MHC gene promoter region, which makes the AS transcript fully complementary to the beta MHC gene. The expression of the AS transcript was positively correlated with levels of alpha MHC gene sense mRNA and negatively correlated with the beta MHC gene sense mRNA. METHODS: A promoter corresponding to the −2285/ −945 region of the IGS was inserted into a luciferase plasmid in the 3′ to 5′ AS direction, and was injected into rat ventricle. RESULTS: This AS promoter was activated in control heart and decreased greatly in response to PTU and STZ and increased 9x in hyperthyroid rats. Mutating a putative retinoic acid receptor (RAR) (a known thyroid hormone receptor cofactor) binding site abolished reporter activity in Cont, PTU, and STZ hearts. We examined the effect of a RAR repeat sequence on promoter activity as well as the transcriptional activity of RAR in vivo. CONCLUSIONS: We have shown that the RAR is a critical element in the AS RNA regulation.