Role of Antisense RNA in Coordinating Cardiac Myosin Heavy Chain Gene Switching

Abstract

A novel mechanism of regulation of cardiac ␣ and ␤ myosin heavy chain gene by naturally occurring antisense transcription was elucidated via pre-mRNA analysis. Herein, we report the expression of an antisense ␤ myosin heavy chain RNA in the normal rodent myocardium. The pattern of expression of the antisense ␤MHC RNA (␤ RNA) under altered thyroid state and in diabetes directly correlates with that of the ␣ pre-mRNA/mRNA, whereas it negatively correlates with the ␤ mRNA expression. Rapid amplification of the 5 end shows that this antisense transcript originates 2 kb downstream of the ␤ gene, and it is transcribed across the entire ␤ gene from the opposite strand. Our results demonstrate that the ␤-␣ myosin heavy chain intergenic DNA possesses a bidirectional transcriptional activity, one direction transcribing the ␣ gene, and the opposite direction transcribing the antisense ␤ RNA. This process turns on the ␣ expression, and it simultaneously turns off that of the ␤ and thus coordinates ␣ and ␤ expression in an opposite fashion. Comparative analyses of the intergenic DNA sequence across five mammalian species revealed a conserved region that is proposed to be a common regulatory region for the ␣ and antisense ␤ promoter. This finding unravels the mechanism of cardiac ␣-␤ gene switching and implicates the role of cardiac myosin gene organization with their function.Cardiac muscle expresses two myosin heavy chain (MHC) 1 isoforms designated as ␣ (high ATPase) and ␤ (low ATPase) that are encoded by two distinct genes located in close proximity on the same chromosome (1-4). The MHC is the molecular motor driving muscle contraction, and its phenotypic composition regulates the intrinsic contraction properties of the heart (5, 6). Cardiac MHC isoform expression is developmentally regulated (5), and it can change totally in either direction under certain pathophysiological states (7-9). For example during the first 3 weeks of postnatal life of rodents there is a complete switch from a predominant ␤MHC expression at birth (Ͼ90%) to a predominant ␣MHC expression at 3 weeks of age (Ͼ95%). Throughout adult life, the ␣MHC expression predominates in a normal rodent heart, with ␤MHC expression gradually increasing as the animal gets older. At any time during life, the pattern of MHC expression can be altered. Hypothyroidism and diabetes are associated with a switch in the cardiac MHC gene expression from a predominant ␣MHC to a predominant ␤MHC. In contrast, thyroid hormones treatment increases the ␣MHC expression while down-regulating the ␤MHC expression. The exact molecular mechanisms causing this tightly coordinated regulation of these two genes remains unclear. Thyroid hormone has been shown to be a major regulator of MHC gene expression, and its regulation is thought to occur mainly via transcriptional processes regulating each gene independently in a well coordinated fashion (8). Several thyroid responsive elements have been located on the promoter of the ␣MHC gene, whereas the localized action of thyroid hormone on the ...

Factors controlling cardiac myosin-isoform shift during hypertrophy and heart failure

“…This intergenic transcription proceeds in the direction of the βMHC gene, thus yielding β-antisence RNA. Although the exact role of this anti-sense transcript is not yet understood, it has been found to be associated with MHC isoform switching in the heart in response to pathologic stimuli, such as hemodynamic overload and diabetes [35][36][37]. A similar anti-sense mediated regulation of the MHC genes has also been suggested for isoform switching in slow skeletal muscle fibers [38].…”

“…Recent studies have suggested that the genomic order and tandem organization of the MHC genes on the same chromosome might be important for the coordinated regulation of these two genes. The intergenic sequences between the β-and αMHC genes have been shown to synthesize βMHC anti-sense RNA, which seems to correlate with the antithetic expression of these two transcripts in response to various stimuli [35]. Other studies have documented a role of microRNA-208 in antithetic expression of α-and βMHC isoforms in stress conditions.…”

The four Rs of RNA-directed evolution

“…For example, a bidirectional promoter in the cardiac myosin locus that activates read-out of MYHC6 also generates an antisense transcript that switches off the upstream gene MYHC7 (ref. 56). Expression of MYHC6 is thus accompanied by silencing of MYHC7.…”

Dynamics of Myosin Heavy Chain Gene Regulation in Slow Skeletal Muscle

“…Recently, we reported that in cardiac muscle, a switch from ␣ to ␤ MHC is apparently regulated by a mechanism whereby decreased ␣ promoter activity is associated with decreased expression of naturally occurring antisense RNA transcripts (NAT), antithetical to the upstream ␤ gene, resulting in enhanced ␤ mRNA accumulation (14). This antisense ␤ transcript starts in the intergenic region between ␤ and ␣, and its transcriptional activity is positively correlated with that of the downstream ␣ MHC gene.…”

“…In view of the unique mechanism of isoform switching from ␣ to ␤ MHC involving NATs in cardiac muscle (14), the purpose of this study was to determine whether a different group of tandemly linked MHC genes in a different type of muscle tissue exhibits a similar pattern of regulation. Therefore, we tested the hypothesis that in skeletal muscle MHC NATs are expressed.…”