beta1 integrin and organized actin filaments facilitate cardiomyocyte-specific RhoA-dependent activation of the skeletal alpha-actin promoter.
Activation of RhoA GTPase causes actin filament bundling into stress fibers, integrin clustering, and focal adhesion formation through its action on actin cytoskeleton organization. RhoA also regulates transcriptional activity of serum response factor (SRF). Recent studies in NIH 3T3 fibroblasts have shown that SRF activation by RhoA does not require an organized cytoskeleton and may be regulated by G-actin level. In cardiac myocytes, the organization of actin fibers into myofibrils is one of the primary characteristics of cardiac differentiation and hypertrophy. The primary purpose of this study was to examine if RhoA regulates SRF-dependent gene expression in neonatal cardiomyocytes in a manner different from that observed in fibroblasts. Our results show that RhoA-dependent skeletal alpha-actin promoter activation requires beta1 integrin and a functional cytoskeleton in cardiomyocytes but not in NIH 3T3 fibroblasts. Activation of the alpha-actin promoter by RhoA is greatly potentiated (up to 15-fold) by co-expression of the integrin beta1A or beta1D isoform but is significantly reduced by 70% with a co-expressed dominant negative mutant of beta1 integrin. Furthermore, clustering of beta1 integrin with anti-beta1 integrin antibodies potentiates synergistic RhoA and beta1 integrin activation of the alpha-actin promoter. Cytochalasin D and latrunculin B, inhibitors of actin polymerization, significantly reduced RhoA-induced activation of the alpha-actin promoter. Jasplakinolide, an actin polymerizing agent, mimics the synergistic effect of RhoA and beta1 integrin on the actin promoter. These observations support the concept that RhoA regulates SRF-dependent cardiac gene expression through cross-talk with beta1 integrin signal pathway via an organized actin cytoskeleton.
Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, Texas, USA.
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L
Wei
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Wang
J A
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Carson
J E
JE
Agan
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Imanaka-Yoshida
R J
RJ
Schwartz
D016475:3T3 Cells; D000199:Actins / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D015638:Cytochalasin D / Q000494:pharmacology; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D005786:Gene Expression Regulation / Q000235:genetics; D017930:Genes, Reporter; D019012:Integrin beta1 / Q000378:metabolism*; D051379:Mice; D009206:Myocardium / Q000166:cytology* / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015375:Receptors, Interleukin-2 / Q000235:genetics / Q000378:metabolism; D026362:Serum Response Factor; D015398:Signal Transduction / Q000502:physiology; D014162:Transfection; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D000199:Actins; D016895:Culture Media, Serum-Free; D004268:DNA-Binding Proteins; D019012:Integrin beta1; D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D015375:Receptors, Interleukin-2; D026362:Serum Response Factor; D015638:Cytochalasin D; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; C496052:Ptk2 protein, rat; D020742:rhoA GTP-Binding Protein