title: Nuclear RhoA signaling regulates MRTF-dependent SMC-specific transcription.
abstract: We have previously shown that RhoA-mediated actin polymerization stimulates smooth muscle cell (SMC)-specific transcription by regulating the nuclear localization of the myocardin-related transcription factors (MRTFs). On the basis of the recent demonstration that nuclear G-actin regulates MRTF nuclear export and observations from our laboratory and others that the RhoA effector, mDia2, shuttles between the nucleus and cytoplasm, we investigated whether nuclear RhoA signaling plays a role in regulating MRTF activity. We identified sequences that control mDia2 nuclear-cytoplasmic shuttling and used mDia2 variants to demonstrate that the ability of mDia2 to fully stimulate MRTF nuclear accumulation and SMC-specific gene transcription was dependent on its localization to the nucleus. To test whether RhoA signaling promotes nuclear actin polymerization, we established a fluorescence recovery after photobleaching (FRAP)-based assay to measure green fluorescent protein-actin diffusion in the nuclear compartment. Nuclear actin FRAP was delayed in cells expressing nuclear-targeted constitutively active mDia1 and mDia2 variants and in cells treated with the polymerization inducer, jasplakinolide. In contrast, FRAP was enhanced in cells expressing a nuclear-targeted variant of mDia that inhibits both mDia1 and mDia2. Treatment of 10T1/2 cells with sphingosine 1-phosphate induced RhoA activity in the nucleus and forced nuclear localization of RhoA or the Rho-specific guanine nucleotide exchange factor (GEF), leukemia-associated RhoGEF, enhanced the ability of these proteins to stimulate MRTF activity. Taken together, these data support the emerging idea that RhoA-dependent nuclear actin polymerization has important effects on transcription and nuclear structure.
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina.
authors.forename: Dean P
authors.initials: DP
authors.lastname: Staus
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina.
authors.forename: Laura
authors.initials: L
authors.lastname: Weise-Cross
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina.
authors.forename: Kevin D
authors.initials: KD
authors.lastname: Mangum
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina.
authors.forename: Matt D
authors.initials: MD
authors.lastname: Medlin
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina.
authors.forename: Lee
authors.initials: L
authors.lastname: Mangiante
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina.
authors.forename: Joan M
authors.initials: JM
authors.lastname: Taylor
authors.affiliation: Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill, North Carolina cmack@med.unc.edu.
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Mack
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D021581:Active Transport, Cell Nucleus; D000818:Animals; D001013:Aorta, Thoracic / Q000201:enzymology; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D002467:Cell Nucleus / Q000201:enzymology*; D002522:Chlorocebus aethiops; D036681:Fluorescence Recovery After Photobleaching; D000080865:Formins; D005786:Gene Expression Regulation; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D032389:Myocytes, Smooth Muscle / Q000201:enzymology*; D009252:NADPH Dehydrogenase / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D064067:Rho Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D020741:rho GTP-Binding Proteins / Q000378:metabolism*; D020742:rhoA GTP-Binding Protein
chemical_list: D000199:Actins; C577223:Arhgef12 protein, mouse; D002352:Carrier Proteins; C491464:Diap1 protein, mouse; D000080865:Formins; D008869:Microtubule-Associated Proteins; C511173:Mrtfa protein, mouse; D064067:Rho Guanine Nucleotide Exchange Factors; D015534:Trans-Activators; D014157:Transcription Factors; C503315:myocardin-related transcription factor B, mouse; D049452:Green Fluorescent Proteins; C508552:Dia2 protein, mouse; D009252:NADPH Dehydrogenase; C526380:RhoA protein, mouse; D020741:rho GTP-Binding Proteins; D020742:rhoA GTP-Binding Protein
keywords: RhoA; diaphanous formins; myocardin-related factors; serum response factor; smooth muscle