Abstract 18902: DNA Methylation Changes in the b-MHC Locus Mediate Gene Regulation by Antisense RNA in Mouse Postnatal Cardiomyocyte Development
Objective: Expression of the b-MHC gene in ventricular cardiomyocytes is rapidly suppressed during early neonatal development in rodents. Antisense, non-coding RNA at this locus is developmentally expressed and inversely responsive to expression of the b-MHC gene. However, the mechanism mediating b-MHC gene suppression remains unknown. DNA methylation within promoter region is believed to silence gene expression, although the effects of DNA methylation on transcriptional activity in other regions during development remain controversial. Since the transcription-targeted DNA methylation has been implicated, we hypothesized that DNA methylation mediates the suppression of b-MHC expression under the transcription of antisense non-coding RNA.
Methods and Results: We analyzed the relationship between transcriptional and epigenetic status of a-MHC and b-MHC at birth, at 7, 14, and 21 days and 8 weeks after birth in purified mouse cardiomyocytes. Transcriptional status was analyzed using strand-specific quantitative RT-PCR, while DNA methylation status across the MHC gene locus was assessed by bisulfite sequencing and the novel genome-wide analytical method by next-generation sequencing. Curiously, the b-MHC gene promoter region was always unmethylated, whereas the “gene body” region, which is a genomic region corresponding to pre-mRNA sequence, of b-HMC showed a dynamic change in DNA methylation status during postnatal development that correlated with the transcription of antisense b-MHC RNA. Propylthiouracil treatment, which suppresses the expression of antisense RNA, reactivate the expression of b-MHC and reduced DNA methylation in the “gene body” region of b-MHC gene in the neonate cardiomyocytes. In addition, the treatment by DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine, increased the expression of b-MHC in adult cardiomyocytes.
Conclusions: The DNA methylation in the gene body region of the b-MHC gene changes dramatically during postnatal rodent development, and suppresses the transcription of b-MHC in adult cardiomyocytes. This demonstrates that antisense RNA transcription-targeted DNA methylation in the gene body region could regulate b-MHC gene expression during cardiomyocyte development.
- © 2010 by American Heart Association, Inc.