RESUMO
BACKGROUND: Hypertrophic cardiomyocyte growth and dysfunction accompany various forms of heart disease. The mechanisms responsible for transcriptional changes that affect cardiac physiology and the transition to heart failure are not well understood. The intercalated disc (ID) is a specialized intercellular junction coupling cardiomyocyte force transmission and propagation of electrical activity. The ID is gaining attention as a mechanosensitive signaling hub and hotspot for causative mutations in cardiomyopathy. METHODS: Transmission electron microscopy, confocal microscopy, and single-molecule localization microscopy were used to examine changes in ID structure and protein localization in the murine and human heart. We conducted detailed cardiac functional assessment and transcriptional profiling of mice lacking myocardin-related transcription factor (MRTF)-A and MRTF-B specifically in adult cardiomyocytes to evaluate the role of mechanosensitive regulation of gene expression in load-induced ventricular remodeling. RESULTS: We found that MRTFs localize to IDs in the healthy human heart and accumulate in the nucleus in heart failure. Although mice lacking MRTFs in adult cardiomyocytes display normal cardiac physiology at baseline, pressure overload leads to rapid heart failure characterized by sarcomere disarray, ID disintegration, chamber dilation and wall thinning, cardiac functional decline, and partially penetrant acute lethality. Transcriptional profiling reveals a program of actin cytoskeleton and cardiomyocyte adhesion genes driven by MRTFs during pressure overload. Indeed, conspicuous remodeling of gap junctions at IDs identified by single-molecule localization microscopy may partially stem from a reduction in Mapre1 expression, which we show is a direct mechanosensitive MRTF target. CONCLUSIONS: Our study describes a novel paradigm in which MRTFs control an acute mechanosensitive signaling circuit that coordinates cross-talk between the actin and microtubule cytoskeleton and maintains ID integrity and cardiomyocyte homeostasis in heart disease.
Assuntos
Insuficiência Cardíaca/metabolismo , Hipertrofia Ventricular Esquerda/metabolismo , Mecanotransdução Celular , Miócitos Cardíacos/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Idoso , Animais , Animais Recém-Nascidos , Células COS , Estudos de Casos e Controles , Chlorocebus aethiops , Conexina 43/genética , Conexina 43/metabolismo , Feminino , Regulação da Expressão Gênica , Insuficiência Cardíaca/genética , Insuficiência Cardíaca/patologia , Insuficiência Cardíaca/fisiopatologia , Humanos , Hipertrofia Ventricular Esquerda/genética , Hipertrofia Ventricular Esquerda/patologia , Hipertrofia Ventricular Esquerda/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Confocal , Microscopia Eletrônica de Transmissão , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Pessoa de Meia-Idade , Miócitos Cardíacos/ultraestrutura , Células NIH 3T3 , Imagem Individual de Molécula , Transativadores/deficiência , Transativadores/genética , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética , Função Ventricular Esquerda , Remodelação VentricularRESUMO
OBJECTIVE: The chromatin remodeling enzyme BRG1 (brahma-related gene 1) transcriptionally regulates target genes important for early blood vessel development and primitive hematopoiesis. However, because Brg1 deletion in vascular progenitor cells results in lethal anemia by embryonic day 10.5 (E10.5), roles for BRG1 in embryonic vascular development after midgestation are unknown. In this study, we sought to determine whether endothelial cell BRG1 regulates genes important for vascular development or maintenance later in embryonic development. APPROACH AND RESULTS: Using mice with temporally inducible deletion of endothelial BRG1 (Brg1fl/fl;Cdh5(PAC)-CreERT2 ), we found that Brg1 excision between E9.5 and 11.5 results in capillary dilation and lethal hemorrhage by E14.5. This phenotype strongly resembles that seen when the SRF (serum response factor) transcription factor is deleted from embryonic endothelial cells. Although expression of Srf and several of its known endothelial cell target genes are downregulated in BRG1-depleted endothelial cells, we did not detect binding of BRG1 at these gene promoters, indicating that they are not direct BRG1 target genes. Instead, we found that BRG1 binds to the promoters of the SRF cofactors Mrtfa and Mrtfb (myocardin-related transcription factors A and B) in endothelial cells, and these genes are downregulated in Brg1-deficient endothelial cells. CONCLUSIONS: BRG1 promotes transcription of endothelial Mrtfa and Mrtfb, which elevates expression of SRF and SRF target genes that establish embryonic capillary integrity. These data highlight a new and temporally specific role for BRG1 in embryonic vasculature and provide novel information about epigenetic regulation of Mrtf expression and SRF signaling in developing blood vessels.
Assuntos
Capilares/metabolismo , DNA Helicases/metabolismo , Células Endoteliais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Neovascularização Fisiológica , Proteínas Nucleares/metabolismo , Transativadores/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Animais , Antígenos CD/genética , Sítios de Ligação , Caderinas/genética , Capilares/embriologia , Linhagem Celular , DNA Helicases/deficiência , DNA Helicases/genética , Epigênese Genética , Genótipo , Idade Gestacional , Integrases/genética , Camundongos Knockout , Morfogênese , Proteínas Nucleares/deficiência , Proteínas Nucleares/genética , Fenótipo , Regiões Promotoras Genéticas , Interferência de RNA , Fator de Resposta Sérica/genética , Fator de Resposta Sérica/metabolismo , Transdução de Sinais , Transativadores/genética , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética , TransfecçãoRESUMO
Myocardin-Related Transcription Factors A and B (MRTF-A and MRTF-B) are highly homologous proteins that function as powerful coactivators of serum response factor (SRF), a ubiquitously expressed transcription factor essential for cardiac development. The SRF/MRTF complex binds to CArG boxes found in the control regions of genes that regulate cytoskeletal dynamics and muscle contraction, among other processes. While SRF is required for heart development and function, the role of MRTFs in the developing or adult heart has not been explored. Through cardiac-specific deletion of MRTF alleles in mice, we show that either MRTF-A or MRTF-B is dispensable for cardiac development and function, whereas deletion of both MRTF-A and MRTF-B causes a spectrum of structural and functional cardiac abnormalities. Defects observed in MRTF-A/B null mice ranged from reduced cardiac contractility and adult onset heart failure to neonatal lethality accompanied by sarcomere disarray. RNA-seq analysis on neonatal hearts identified the most altered pathways in MRTF double knockout hearts as being involved in cytoskeletal organization. Together, these findings demonstrate redundant but essential roles of the MRTFs in maintenance of cardiac structure and function and as indispensible links in cardiac cytoskeletal gene regulatory networks.