Myocardin regulates BMP10 expression and is required for heart development.
ABSTRACT Myocardin is a muscle lineage-restricted transcriptional coactivator that has been shown to transduce extracellular signals to the nucleus required for SMC differentiation. We now report the discovery of a myocardin/BMP10 (where BMP10 indicates bone morphogenetic protein 10) signaling pathway required for cardiac growth, chamber maturation, and embryonic survival. Myocardin-null (Myocd) embryos and embryos harboring a cardiomyocyte-restricted mutation in the Myocd gene exhibited myocardial hypoplasia, defective atrial and ventricular chamber maturation, heart failure, and embryonic lethality. Cardiac hypoplasia was caused by decreased cardiomyocyte proliferation accompanied by a dramatic increase in programmed cell death. Defective chamber maturation and the block in cardiomyocyte proliferation were caused in part by a block in BMP10 signaling. Myocardin transactivated the Bmp10 gene via binding of a serum response factor-myocardin protein complex to a nonconsensus CArG element in the Bmp10 promoter. Expression of p57kip2, a BMP10-regulated cyclin-dependent kinase inhibitor, was induced in Myocd-/- hearts, while BMP10-activated cardiogenic transcription factors, including NKX2.5 and MEF2c, were repressed. Remarkably, when embryonic Myocd-/- hearts were cultured ex vivo in BMP10-conditioned medium, the defects in cardiomyocyte proliferation and p57kip2 expression were rescued. Taken together, these data identify a heretofore undescribed myocardin/BMP10 signaling pathway that regulates cardiomyocyte proliferation and apoptosis in the embryonic heart.
- SourceAvailable from: Jianyun Yan
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- "Cre recombinase from bacteriophage P1 recognizes specific 34-bp LoxP sequences and excises LoxPflanked DNA at high efficiency (Branda and Dymecki, 2004; Sauer and Henderson, 1988). Conditional knockout with Cre-LoxP technology allows inactivation of genes in a tissue-specific manner, and has greatly advanced our knowledge on genes' function in heart development and disease (Huang et al., 2012; Jiao et al., 2006; Song et al., 2007). To achieve myocardial specific deletion, a number of Cre deleter mouse lines have been created based on the myocardial genes including myosin light chain 2v (MLC2v), cardiac myosin heavy chain (Myh6) and cardiac Troponin T (Tnnt2) (Agah et al., 1997; Chen et al., 1998; Jiao et al., 2003; Moses et al., 2001). "
ABSTRACT: Tnnt2, encoding thin-filament sarcomeric protein cardiac troponin T, plays critical roles in heart development and function in mammals. To develop an inducible genetic deletion strategy in myocardial cells, we generated a new Tnnt2:MerCreMer (Tnnt2(MerCreMer/+) ) knock-in mouse. Rosa26 reporter lines were used to examine the specificity and efficiency of the inducible Cre recombinase. We found that Cre was specifically and robustly expressed in the cardiomyocytes at embryonic and adult stages following tamoxifen induction. The knock-in allele on Tnnt2 locus does not impact cardiac function. These results suggest that this new Tnnt2(MerCreMer/+) mouse could be applied towards the temporal genetic deletion of genes of interests in cardiomyocytes with Cre-LoxP technology. The Tnnt2(MerCreMer/+) mouse model also provides a useful tool to trace myocardial lineage during development and repair after cardiac injury. This article is protected by copyright. All rights reserved. © 2015 Wiley Periodicals, Inc.genesis 05/2015; 53(6). DOI:10.1002/dvg.22861 · 2.04 Impact Factor
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ABSTRACT: Heart disease remains a leading cause of death worldwide. Owing to the limited regenerative capacity of heart tissue, cardiac regenerative therapy has emerged as an attractive approach. Direct reprogramming of human cardiac fibroblasts (HCFs) into cardiomyocytes may hold great potential for this purpose. We reported previously that induced cardiomyocyte-like cells (iCMs) can be directly generated from mouse cardiac fibroblasts in vitro and vivo by transduction of three transcription factors: Gata4, Mef2c, and Tbx5, collectively termed GMT. In the present study, we sought to determine whether human fibroblasts also could be converted to iCMs by defined factors. Our initial finding that GMT was not sufficient for cardiac induction in HCFs prompted us to screen for additional factors to promote cardiac reprogramming by analyzing multiple cardiac-specific gene induction with quantitative RT-PCR. The addition of Mesp1 and Myocd to GMT up-regulated a broader spectrum of cardiac genes in HCFs more efficiently compared with GMT alone. The HCFs and human dermal fibroblasts transduced with GMT, Mesp1, and Myocd (GMTMM) changed the cell morphology from a spindle shape to a rod-like or polygonal shape, expressed multiple cardiac-specific proteins, increased a broad range of cardiac genes and concomitantly suppressed fibroblast genes, and exhibited spontaneous Ca(2+) oscillations. Moreover, the cells matured to exhibit action potentials and contract synchronously in coculture with murine cardiomyocytes. A 5-ethynyl-2'-deoxyuridine assay revealed that the iCMs thus generated do not pass through a mitotic cell state. These findings demonstrate that human fibroblasts can be directly converted to iCMs by defined factors, which may facilitate future applications in regenerative medicine.Proceedings of the National Academy of Sciences 07/2013; 110. DOI:10.1073/pnas.1304053110 · 9.81 Impact Factor
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ABSTRACT: Author Summary miRNAs are small non-coding RNAs involved in posttranscriptional regulation of protein-coding genes. In the mammalian genome, two distinct gene clusters code for miR-1 and miR-133a. Primary sequences of mature miR-1 or miR-133a are identical and both gene clusters show similar expression in the heart and skeletal muscle. We have generated compound mutant mice of both miR-1/133a gene clusters resulting in early arrest of heart development while single cluster mutants showed normal morphology but reacted differently to pressure overload. Compound mutant cardiomyocytes were characterized by an immature, mixed smooth muscle-heart muscle phenotype, indicating that miR1-/133a are responsible for specification of the cardiomyogenic lineage. Our search for miR1-/133a targets identified myocardin, which was strongly up-regulated in mutant hearts, while several other putative miR-1/133a targets that have been described before were not altered, indicating that miR-1/133a target control strongly depends on the cellular context. Overexpression of myocardin in embryonic hearts recapitulated major aspects of the miR-1/133a mutant phenotype, suggesting that loss of myocardin suppression is the primary reason for incorrect heart muscle specification in the mutants. In addition, we found that myocardin overexpression stimulated expression of miR-1/133a, which argues for a negative feedback loop required for adjustment of myocardin concentrations in the heart.PLoS Genetics 09/2013; 9(9):e1003793. DOI:10.1371/journal.pgen.1003793 · 8.17 Impact Factor