Marguerite Lemonnier

Characterization of a Cardiac-specific Enhancer, Which Directs α-Cardiac Actin Gene Transcription in the Mouse Adult Heart

Expression of the mouse α-cardiac actin gene in skeletal and cardiac muscle is regulated by enhancers lying 5′ to the proximal promoter. Here we report the characterization of a cardiac-specific enhancer located within -2.354/-1.36 kbp of the gene, which is active in cardiocytes but not in C2 skeletal muscle cells. In vivo it directs reporter gene expression to the adult heart, where the proximal promoter alone is inactive. An 85-bp region within the enhancer is highly conserved between human and mouse and contains a central AT-rich site, which is essential for enhancer activity. This site binds myocyte enhancer factor (MEF)2 factors, principally MEF2D and MEF2A in cardiocyte nuclear extracts. These results are discussed in the context of MEF2 activity and of the regulation of the α-cardiac actin locus.

Cell history determines the maintenance of transcriptional differences between left and right ventricular cardiomyocytes in the developing mouse heart.

The molecular mechanisms that establish and maintain transcriptional differences between cardiomyocytes in the left and right ventricular chambers are unkown. We have previously analysed a myosin light chain 3f transgene containing an nlacZ reporter gene, which is transcribed in left but not right ventricular cardiomyocytes. In this report we examine the mechanisms involved in maintaining regionalised transgene expression. Primary cardiomyocytes prepared from left and right ventricular walls of transgenic mice were found to maintain transgene expression status in culture. However, similar cultures prepared from nontransgenic mice or rats show uniform expression after transient transfection of Mlc3f constructs, suggesting that the mechanism responsible for differential expression of the transgene between left and right ventricular cells does not operate on transiently introduced molecules. These data suggest that developmental cell history determines transgene expression status. Maintenance of transgene expression status is regulated by a cell-autonomous mechanism that is independent of DNA methylation, trichostatin A-sensitive histone deacetylation and miss-expression of transcription factors that are expressed in the left or right ventricles of the embryonic heart. Parallels between Mlc3f transgene repression in right ventricular cardiomyocytes and polycomb-mediated silencing in Drosophila suggest that Mlc3f regulatory sequences included on the transgene may contain a cellular memory module that is switched into an on or off state during early cardiogenesis. Epigenetic mechanisms may therefore be involved in maintaining patterning of the mammalian myocardium.

Denervated chicken breast muscle displays discoordinate regulation and differential patterns of expression of αf and β tropomyosin genes

SummaryThe expression of the α fast (αf) and β tropomyosin (TM) genes has been analysed with muscle-specific and common cDNA probes after unilateral nerve section of the pectoralis major muscle (PM) in 4-week-old chickens. The following were observed in denervated muscles. (1) The βTM mRNA, which was repressed during development, reaccumulates in a biphasic curve with the increase in the βTM protein lagging behind the changes in its mRNA. Accordingly, no βTM is seen in products translated in vitro from total and polyA+ RNA obtained 1 week after denervation. No such translation block is seen with RNA obtained from control or muscles denervated for 6 weeks. (2) No changes in the αfTM mRNA and corresponding protein are observed. (3) RNA processing of the two genes is not changed. (4) In the contralateral muscles, transitory increases in αf and βTM mRNAs are observed while the corresponding proteins remain unchanged. Our data suggest that muscle fibres display early and long-term responses to the loss of neural input which might result from a combination of changes produced by regenerative processes and reprogramming of existing fibres. Moreover, in contrast to normal development, no reciprocal changes of αf and βTM expression are seen in denervated muscles.