The Experts below are selected from a list of 3039 Experts worldwide ranked by ideXlab platform
Dazhi Wang - One of the best experts on this subject based on the ideXlab platform.
-
Generation of a Cre knock-in into the Myocardin locus to mark early cardiac and smooth muscle cell lineages.
Genesis, 2014Co-Authors: Ramón A. Espinoza-lewis, Dazhi WangAbstract:Summary The molecular events that control cell fate determination in cardiac and smooth muscle lineages remain elusive. Myocardin is an important transcription cofactor that regulates cell proliferation, differentiation, and development of the cardiovascular system. Here, we describe the construction and analysis of a dual Cre and enhanced green fluorescent protein (EGFP) knock-in mouse line in the Myocardin locus (MyocdKI). We report that the MyocdKI allele expresses the Cre enzyme and the EGFP in a manner that recapitulates endogenous Myocardin expression patterns. We show that Myocardin expression marks the earliest cardiac and smooth muscle lineages. Furthermore, this genetic model allows for the identification of a cardiac cell population, which maintains both Myocardin and Isl-1 expression, in E7.75–E8.0 embryos, highlighting the contribution and merging of the first and second heart fields during cardiogenesis. Therefore, the MyocdKI allele is a unique tool for studying cardiovascular development and lineage-specific gene manipulation. genesis 52:879–887, 2014. © 2014 Wiley Periodicals, Inc.
-
Opposite roles of Myocardin and atrogin-1 in L6 myoblast differentiation†
Journal of Cellular Physiology, 2013Co-Authors: Yulan Jiang, Dazhi Wang, Pavneet Singh, Yi‐xia Zhou, Xi-long ZhengAbstract:L6 rat myoblasts undergo differentiation and myotube formation when cultured in medium containing a low-concentration of serum, but the underlying mechanism is not well understood. The role of atrogin-1, an E3 ligase with well-characterized roles in muscle atrophy, has not been defined in muscle differentiation. Myocardin is a coactivator of serum response factor (SRF), which together promotes smooth muscle differentiation. Myocardin is transiently expressed in skeletal muscle progenitor cells with inhibitory effects on the expression of myogenin and muscle differentiation. It remains unknown whether Myocardin, which undergoes ubiquitination degradation, plays a role in L6 cell differentiation. The current study aimed to investigate the potential roles of Myocardin and atrogin-1 in differentiation of L6 cells. As reported by many others, shifting to medium containing 2% serum induced myotube formation of L6 cells. Differentiation was accompanied by up-regulation of atrogin-1 and down-regulation of Myocardin, suggesting that both may be involved in muscle differentiation. As expected, over-expression of atrogin-1 stimulated the expression of troponin T and myogenin and differentiation of the L6 myoblasts. Co-expression of Myocardin with atrogin-1 inhibited atrogin-1-induced myogenin expression. Over-expression of atrogin-1 decreased Myocardin protein level, albeit without affecting its mRNA level. Small-interfering RNA-mediated knockdown of atrogin-1 increased Myocardin protein. Consistently, ectopic expression of Myocardin inhibited myogenic differentiation. Unexpectedly, Myocardin decreased the expression of atrogin-1 without involving Foxo1. Taken together, our results have demonstrated that atrogin-1 plays a positive role in skeletal muscle differentiation through down-regulation of Myocardin.
-
acetylation of Myocardin is required for the activation of cardiac and smooth muscle genes
Journal of Biological Chemistry, 2012Co-Authors: Chunbo Wang, Dazhi Wang, Ruhang Tang, Huaqun Chen, Zheng Zhang, Mariko TatsuguchiAbstract:Abstract Myocardin belongs to the SAF-A/B, Acinus, PIAS (SAP) domain family of transcription factors and is specifically expressed in cardiac and smooth muscle. Myocardin functions as a transcriptional coactivator of SRF and is sufficient and necessary for smooth muscle gene expression. We have previously found that Myocardin induces the acetylation of nucleosomal histones surrounding SRF-binding sites in the control regions of cardiac and smooth muscle genes through recruiting chromatin-modifying enzyme p300, yet no studies have determined whether Myocardin itself is similarly modified. In this study, we show that Myocardin is a direct target for p300-mediated acetylation. p300 acetylates lysine residues at the N terminus of the Myocardin protein. Interestingly, a direct interaction between p300 and Myocardin, which is mediated by the C terminus of Myocardin, is required for the acetylation event. Acetylation of Myocardin by p300 enhances the association of Myocardin and SRF as well as the formation of the Myocardin-SRF-CArG box ternary complex. Conversely, acetylation of Myocardin decreases the binding of histone deacetylase 5 (HDAC5) to Myocardin. Acetylation of Myocardin is required for Myocardin to activate smooth muscle genes. Our study demonstrates that acetylation plays a key role in modulating Myocardin function in controlling cardiac and smooth muscle gene expression.
-
synergistic activation of cardiac genes by Myocardin and tbx5
PLOS ONE, 2011Co-Authors: Chunbo Wang, Dazhi Wang, Qing Kenneth WangAbstract:Myocardial differentiation is associated with the activation and expression of an array of cardiac specific genes. However, the transcriptional networks that control cardiac gene expression are not completely understood. Myocardin is a cardiac and smooth muscle-specific expressed transcriptional coactivator of Serum Response Factor (SRF) and is able to potently activate cardiac and smooth muscle gene expression during development. We hypothesize that Myocardin discriminates between cardiac and smooth muscle specific genes by associating with distinct co-factors. Here, we show that Myocardin directly interacts with Tbx5, a member of the T-box family of transcription factors involved in the Holt-Oram syndrome. Tbx5 synergizes with Myocardin to activate expression of the cardiac specific genes atrial natriuretic factor (ANF) and alpha myosin heavy chain (α-MHC), but not that of smooth muscle specific genes SM22 or smooth muscle myosin heavy chain (SM-MHC). We found that this synergistic activation of shared target genes is dependent on the binding sites for Tbx5, T-box factor-Binding Elements (TBEs). Myocardin and Tbx5 physically interact and their interaction domains were mapped to the basic domain and the coil domain of Myocardin and Tbx5, respectively. Our analysis demonstrates that the Tbx5G80R mutation, which leads to the Holt-Oram syndrome in humans, failed to synergize with Myocardin to activate cardiac gene expression. These data uncover a key role for Tbx5 and Myocardin in establishing the transcriptional foundation for cardiac gene activation and suggest that the interaction of Myocardin and Tbx5 maybe involved in cardiac development and diseases.
-
induction of microrna 1 by Myocardin in smooth muscle cells inhibits cell proliferation
Arteriosclerosis Thrombosis and Vascular Biology, 2011Co-Authors: Jie Chen, Dazhi Wang, Yulan Jiang, Jingjing Li, Sarvan Kumar Radhakrishnan, Zhanpeng Huang, Elisabeth P C Kilsdonk, Xi-long ZhengAbstract:Objective— Myocardin is a cardiac- and smooth muscle–specific transcription co-factor that potently activates the expression of downstream target genes. Previously, we demonstrated that overexpression of Myocardin inhibited the proliferation of smooth muscle cells (SMCs). Recently, Myocardin was reported to induce the expression of microRNA-1 (miR-1) in cardiomyocytes. In this study, we investigated whether Myocardin induces miR-1 expression to mediate its inhibitory effects on SMC proliferation. Methods and Results— Using tetracycline-regulated expression (T-REx) inducible system expressing Myocardin in human vascular SMCs, we found that overexpression of Myocardin resulted in significant induction of miR-1 expression and inhibition of SMC proliferation, which was reversed by miR-1 inhibitors. Consistently, introduction of miR-1 into SMCs inhibited their proliferation. We isolated spindle-shaped and epithelioid human SMCs and demonstrated that spindle-shaped SMCs were more differentiated and less proliferative. Correspondingly, spindle-shaped SMCs had significantly higher expression levels of both Myocardin and miR-1 than epithelioid SMCs. We identified Pim-1, a serine/threonine kinase, as a target gene for miR-1 in SMCs. Western blot and luciferase reporter assays further confirmed that miR-1 targeted Pim-1 directly. Furthermore, neointimal lesions of mouse carotid arteries displayed downregulation of Myocardin and miR-1 with upregulation of Pim-1. Conclusion— Our data demonstrate that miR-1 participates in Myocardin-dependent of SMC proliferation inhibition.
Eric N Olson - One of the best experts on this subject based on the ideXlab platform.
-
Myocardin is differentially required for the development of smooth muscle cells and cardiomyocytes
American Journal of Physiology-heart and Circulatory Physiology, 2011Co-Authors: Mark H Hoofnagle, Eric N Olson, Ronald L Neppl, Erica Berzin, G Teg C Pipes, Brian W Wamhoff, Avril V Somlyo, Gary K OwensAbstract:Myocardin is a serum response factor (SRF) coactivator exclusively expressed in cardiomyocytes and smooth muscle cells (SMCs). However, there is highly controversial evidence as to whether Myocardin is essential for normal differentiation of these cell types, and there are no data showing whether cardiac or SMC subtypes exhibit differential Myocardin requirements during development. Results of the present studies showed the virtual absence of Myocardin−/− visceral SMCs or ventricular myocytes in chimeric Myocardin knockout (KO) mice generated by injection of Myocardin−/− embryonic stem cells (ESCs) into wild-type (WT; i.e., Myocardin+/+ ESC) blastocysts. In contrast, Myocardin−/− ESCs readily formed vascular SMC, albeit at a reduced frequency compared with WT ESCs. In addition, Myocardin−/− ESCs competed equally with WT ESCs in forming atrial myocytes. The ultrastructural features of Myocardin−/− vascular SMCs and cardiomyocytes were unchanged from their WT counterparts as determined using a unique X-ray microprobe transmission electron microscopic method developed by our laboratory. Myocardin−/− ESC-derived SMCs also showed normal contractile properties in an in vitro embryoid body SMC differentiation model, other than impaired thromboxane A2 responsiveness. Together, these results provide novel evidence that Myocardin is essential for development of visceral SMCs and ventricular myocytes but is dispensable for development of atrial myocytes and vascular SMCs in the setting of chimeric KO mice. In addition, results suggest that as yet undefined defects in development and/or maturation of ventricular cardiomyocytes may have contributed to early embryonic lethality observed in conventional Myocardin KO mice and that observed deficiencies in development of vascular SMC may have been secondary to these defects.
-
Myocardin sumoylation transactivates cardiogenic genes in pluripotent 10t1 2 fibroblasts
Molecular and Cellular Biology, 2007Co-Authors: Zhigao Wang, Eric N Olson, Jun Wang, Ankang Li, Xinhua Feng, Robert J SchwartzAbstract:Myocardin, a serum response factor (SRF)-dependent cofactor, is a potent activator of smooth muscle gene activity but a poor activator of cardiogenic genes in pluripotent 10T1/2 fibroblasts. Posttranslational modification of GATA4, another Myocardin cofactor, by sumoylation strongly activated cardiogenic gene activity. Here, we found that Myocardin's activity was strongly enhanced by SUMO-1 via modification of a lysine residue primarily located at position 445 and that the conversion of this residue to arginine (K445R) impaired Myocardin transactivation. PIAS1 was involved in governing Myocardin activity via its E3 ligase activity that stimulated Myocardin sumoylation on an atypical sumoylation site(s) and by its physical association with Myocardin. Myocardin initiated the expression of cardiac muscle-specified genes, such as those encoding cardiac α-actin and α-myosin heavy chain, in an SRF-dependent manner in 10T1/2 fibroblasts, but only in the presence of coexpressed SUMO-1/PIAS1. Thus, SUMO modification acted as a molecular switch to promote Myocardin's role in cardiogenic gene expression.
-
Myocardin sumoylation transactivates cardiogenic genes in pluripotent 10T1/2 fibroblasts.
Molecular and Cellular Biology, 2006Co-Authors: Jun Wang, Zhigao Wang, Eric N Olson, Ankang Li, Xinhua Feng, Robert J SchwartzAbstract:Myocardin, a serum response factor (SRF)-dependent cofactor, is a potent activator of smooth muscle gene activity but a poor activator of cardiogenic genes in pluripotent 10T1/2 fibroblasts. Posttranslational modification of GATA4, another Myocardin cofactor, by sumoylation strongly activated cardiogenic gene activity. Here, we found that Myocardin's activity was strongly enhanced by SUMO-1 via modification of a lysine residue primarily located at position 445 and that the conversion of this residue to arginine (K445R) impaired Myocardin transactivation. PIAS1 was involved in governing Myocardin activity via its E3 ligase activity that stimulated Myocardin sumoylation on an atypical sumoylation site(s) and by its physical association with Myocardin. Myocardin initiated the expression of cardiac muscle-specified genes, such as those encoding cardiac α-actin and α-myosin heavy chain, in an SRF-dependent manner in 10T1/2 fibroblasts, but only in the presence of coexpressed SUMO-1/PIAS1. Thus, SUMO modification acted as a molecular switch to promote Myocardin's role in cardiogenic gene expression.
-
Myocardin is a direct transcriptional target of mef2 tead and foxo proteins during cardiovascular development
Development, 2006Co-Authors: Esther E Creemers, James A Richardson, Lillian B Sutherland, John Mcanally, Eric N OlsonAbstract:Myocardin is a transcriptional co-activator of serum response factor (Srf), which is a key regulator of the expression of smooth and cardiac muscle genes. Consistent with its role in regulating cardiovascular development, Myocardin is the earliest known marker specific to both the cardiac and smooth muscle lineages during embryogenesis. To understand how the expression of this early transcriptional regulator is initiated and maintained, we scanned 90 kb of genomic DNA encompassing the Myocardin gene for cis-regulatory elements capable of directing Myocardin transcription in cardiac and smooth muscle lineages in vivo. Here, we describe an enhancer that controls cardiovascular expression of the mouse Myocardin gene during mouse embryogenesis and adulthood. Activity of this enhancer in the heart and vascular system requires the combined actions of the Mef2 and Foxo transcription factors. In addition, the Tead transcription factor is required specifically for enhancer activation in neural-crest-derived smooth muscle cells and dorsal aorta. Notably, Myocardin also regulates its own enhancer, but in contrast to the majority of Myocardin target genes, which are dependent on Srf, Myocardin acts through Mef2 to control its enhancer. These findings reveal an Srf-independent mechanism for smooth and cardiac muscle-restricted transcription and provide insight into the regulatory mechanisms responsible for establishing the smooth and cardiac muscle phenotypes during development.
-
coactivation of mef2 by the sap domain proteins Myocardin and mastr
Molecular Cell, 2006Co-Authors: Esther E Creemers, Jiyeon Oh, Lillian B Sutherland, Ana C Barbosa, Eric N OlsonAbstract:Myocardin is a cardiac- and smooth muscle-specific SAP domain transcription factor that functions as a coactivator for serum response factor (SRF), which controls genes involved in muscle differentiation and cell proliferation. The DNA binding domain of SRF, which interacts with Myocardin, shares homology with the MEF2 transcription factor, which also controls muscle and growth-associated genes. Here we show that alternative splicing produces a cardiac-enriched isoform of Myocardin containing a unique peptide sequence that confers the ability to interact with and stimulate the transcriptional activity of MEF2. This MEF2 binding motif is also contained in a previously unknown SAP domain transcription factor, referred to as MASTR, which functions as a MEF2 coactivator. This unique protein-protein interaction motif expands the regulatory potential of Myocardin, and its presence in MASTR reveals a new mechanism for the control of MEF2 activity.
Jiliang Zhou - One of the best experts on this subject based on the ideXlab platform.
-
the transcription factor tead1 represses smooth muscle specific gene expression by abolishing Myocardin function
Journal of Biological Chemistry, 2014Co-Authors: Xiaobo Wang, Guoqing Hu, Yong Wang, Jiliang ZhouAbstract:Abstract The TEAD (transcriptional enhancer activator domain) proteins share an evolutionarily conserved DNA-binding TEA domain, which binds to the MCAT cis-acting regulatory element. Previous studies have shown that TEAD proteins are involved in regulating the expression of smooth muscle α-actin. However, it remains undetermined whether TEAD proteins play a broader role in regulating expression of other genes in vascular smooth muscle cells. In this study, we show that the expression of TEAD1 is significantly induced during smooth muscle cell phenotypic modulation and negatively correlates with smooth muscle-specific gene expression. We further demonstrate that TEAD1 plays a novel role in suppressing expression of smooth muscle-specific genes, including smooth muscle α-actin, by abolishing the promyogenic function of Myocardin, a key mediator of smooth muscle differentiation. Mechanistically, we found that TEAD1 competes with Myocardin for binding to serum response factor (SRF), resulting in disruption of Myocardin and SRF interactions and thereby attenuating expression of smooth muscle-specific genes. This study provides the first evidence demonstrating that TEAD1 is a novel general repressor of smooth muscle-specific gene expression through interfering with Myocardin binding to SRF.
-
SOX9 and Myocardin counteract each other in regulating vascular smooth muscle cell differentiation.
Biochemical and Biophysical Research Communications, 2012Co-Authors: Zhonghui Xu, Xiaobo Wang, Jiliang Zhou, Guangdong Ji, Jianbin Shen, Li LiAbstract:Transdifferentiation of vascular smooth muscle cells (VSMC) into chondrogenic cells contributes significantly to vascular calcification during the pathogenesis of atherosclerosis. However, the transcriptional mechanisms that control such phenotypic switch remain unclear. This process is characterized by the induction of Sox9 and Col2a1 genes accompanied by the repression of Myocardin (Myocd) and SMC differentiation markers such as SM22, SM α-actin and SM-MHC. Here we explore the regulatory role of SOX9, the master regulator for chondrogenesis, in modulating SMC marker gene expression. qRT-PCR and luciferase assays show that over-expression of SOX9 inhibits SMC gene transcription and promoter activities induced by Myocardin, the master regulator of smooth muscle differentiation. Such suppression is independent of the CArG box in the SMC promoters but dependent on Myocardin. EMSA assay further shows that SOX9 neither participates in SRF (serum response factor) binding to the CArG box nor interacts with SRF, while co-immunoprecipitation demonstrates an association of SOX9 with Myocardin. Conversely, Myocardin suppresses SOX9-mediated chondrogenic gene Col2a1 expression. These findings provide the first mechanistic insights into the important regulatory role of SOX9 and Myocardin in controlling the transcription program during SMC transdifferentiation into chondrocytes.
-
transforming growth factor β1 induced transcript 1 protein a novel marker for smooth muscle contractile phenotype is regulated by serum response factor Myocardin protein
Journal of Biological Chemistry, 2011Co-Authors: Xiaobo Wang, Guoqing Hu, Courtney Betts, Erin Y Harmon, Rebecca S Keller, Livingston Van De Water, Jiliang ZhouAbstract:Abstract Serum response factor (SRF) plays a central role in regulating expression of smooth muscle-specific genes partly by associating with the potent tissue-specific cofactor Myocardin. Previous studies have shown that transforming growth factor-β1-induced transcript 1 (TGFB1I1, also known as Hic-5) is a TGF-β-responsive gene and is involved in the cellular response to vascular injury, but the regulation of TGFB1I1 expression remains elusive. In this report, we demonstrated that TGFB1I1 is a novel marker for the smooth muscle contractile phenotype and is regulated by SRF/Myocardin. We found that TGFB1I1 is specifically expressed in smooth muscle cells (SMCs) and in smooth muscle-rich tissues. Furthermore, TGFB1I1 expression is significantly down-regulated in a variety of models for smooth muscle phenotypic modulation. The TGFB1I1 promoter contains an evolutionarily conserved CArG element, and this element is indispensible for Myocardin-induced transactivation of TGFB1I1 promoter. By oligonucleotide pulldown and chromatin immunoprecipitation assays, we found that SRF binds to this CArG element in vitro and in vivo. Ectopic expression of Myocardin is sufficient to induce endogenous TGFB1I1 expression in multiple cell lines whereas knocking-down Myocardin or SRF significantly attenuated TGFB1I1 expression in SMCs. Furthermore, our data demonstrated that SRF is essential for TGF-β-mediated induction of TGFB1I1. Finally, silencing of TGFB1I1 expression significantly promotes SMC proliferation. Collectively, this study provides the first evidence that TGFB1I1 is not only an SRF/Myocardin-regulated smooth muscle marker but also critical for maintaining smooth muscle contractile phenotype by inhibiting smooth muscle proliferation.
-
modulation of Myocardin function by the ubiquitin e3 ligase ubr5
Journal of Biological Chemistry, 2010Co-Authors: Guoqing Hu, Xiaobo Wang, Darren N Saunders, Michelle J Henderson, Amanda J Russell, Paul B Herring, Jiliang ZhouAbstract:Fully differentiated mature smooth muscle cells (SMCs) are characterized by the presence of a unique repertoire of smooth muscle-specific proteins. Although previous studies have shown Myocardin to be a critical transcription factor for stimulating expression of smooth muscle-specific genes, the mechanisms regulating Myocardin activity are still poorly understood. We used a yeast two-hybrid screen with Myocardin as bait to search for factors that may regulate the transcriptional activity of the Myocardin. From this screen we identified a HECT domain-containing protein UBR5 (ubiquitin protein ligase E3 component n-recognin 5) as a Myocardin-binding protein. Previous studies have shown that HECT domain-containing proteins are ubiquitin E3 ligases that play an important role in protein degradation. UBR5 has, however, also been shown to regulate transcription independent of its E3 ligase activity. In the current study we demonstrated that UBR5 localized in the nuclei of SMCs and forms a complex with Myocardin in vivo and in vitro. We also show that UBR5 specifically enhanced trans-activation of smooth muscle-specific promoters by the Myocardin family of proteins. In addition, UBR5 significantly augmented the ability of Myocardin to induce expression of endogenous SMC marker genes independent on its E3 ligase function. Conversely, depletion of endogenous UBR5 by small interfering RNA in fibroblast cells attenuated Myocardin-induced smooth muscle-specific gene expression, and UBR5 knockdown in SMCs resulted in down-regulation of smooth muscle-specific genes. Furthermore, we found that UBR5 can attenuate Myocardin protein degradation resulting in increased Myocardin protein expression without affecting Myocardin mRNA expression. The effects of UBR5 on Myocardin requires only the HECT and UBR1 domains of UBR5. This study reveals an unexpected role for the ubiquitin E3 ligase UBR5 as an activator of smooth muscle differentiation through its ability to stabilize Myocardin protein.
-
the swi snf chromatin remodeling complex regulates Myocardin induced smooth muscle specific gene expression
Arteriosclerosis Thrombosis and Vascular Biology, 2009Co-Authors: Jiliang Zhou, Min Zhang, Hong Fang, Omar Elmounayri, Jennifer M Rodenberg, Anthony N Imbalzano, Paul B HerringAbstract:Objective— Regulatory complexes comprising Myocardin and serum response factor (SRF) are critical for the transcriptional regulation of many smooth muscle–specific genes. However, little is known about the epigenetic mechanisms that regulate the activity of these complexes. In the current study, we investigated the role of SWI/SNF ATP-dependent chromatin remodeling enzymes in regulating the myogenic activity of Myocardin. Methods and Results— We found that both Brg1 and Brm are required for maintaining expression of several smooth muscle–specific genes in primary cultures of aortic smooth muscle cells. Furthermore, the ability of Myocardin to induce expression of smooth muscle–specific genes is abrogated in cells expressing dominant negative Brg1. In SW13 cells, which lack endogenous Brg1 and Brm1, Myocardin is unable to induce expression of smooth muscle–specific genes. Whereas, reconstitution of wild-type, or bromodomain mutant forms Brg1 or Brm1, into SW13 cells restored their responsiveness to Myocardin. SWI/SNF complexes were found to be required for Myocardin to increase SRF binding to the promoters of smooth muscle–specific genes. Brg1 and Brm directly bind to the N terminus of Myocardin, in vitro, through their ATPase domains and Brg1 forms a complex with SRF and Myocardin in vivo in smooth muscle cells. Conclusion— These data demonstrate that the ability of Myocardin to induce smooth muscle–specific gene expression is dependent on its interaction with SWI/SNF ATP-dependent chromatin remodeling complexes.
Zhigao Wang - One of the best experts on this subject based on the ideXlab platform.
-
Myocardin sumoylation transactivates cardiogenic genes in pluripotent 10t1 2 fibroblasts
Molecular and Cellular Biology, 2007Co-Authors: Zhigao Wang, Eric N Olson, Jun Wang, Ankang Li, Xinhua Feng, Robert J SchwartzAbstract:Myocardin, a serum response factor (SRF)-dependent cofactor, is a potent activator of smooth muscle gene activity but a poor activator of cardiogenic genes in pluripotent 10T1/2 fibroblasts. Posttranslational modification of GATA4, another Myocardin cofactor, by sumoylation strongly activated cardiogenic gene activity. Here, we found that Myocardin's activity was strongly enhanced by SUMO-1 via modification of a lysine residue primarily located at position 445 and that the conversion of this residue to arginine (K445R) impaired Myocardin transactivation. PIAS1 was involved in governing Myocardin activity via its E3 ligase activity that stimulated Myocardin sumoylation on an atypical sumoylation site(s) and by its physical association with Myocardin. Myocardin initiated the expression of cardiac muscle-specified genes, such as those encoding cardiac α-actin and α-myosin heavy chain, in an SRF-dependent manner in 10T1/2 fibroblasts, but only in the presence of coexpressed SUMO-1/PIAS1. Thus, SUMO modification acted as a molecular switch to promote Myocardin's role in cardiogenic gene expression.
-
Myocardin sumoylation transactivates cardiogenic genes in pluripotent 10T1/2 fibroblasts.
Molecular and Cellular Biology, 2006Co-Authors: Jun Wang, Zhigao Wang, Eric N Olson, Ankang Li, Xinhua Feng, Robert J SchwartzAbstract:Myocardin, a serum response factor (SRF)-dependent cofactor, is a potent activator of smooth muscle gene activity but a poor activator of cardiogenic genes in pluripotent 10T1/2 fibroblasts. Posttranslational modification of GATA4, another Myocardin cofactor, by sumoylation strongly activated cardiogenic gene activity. Here, we found that Myocardin's activity was strongly enhanced by SUMO-1 via modification of a lysine residue primarily located at position 445 and that the conversion of this residue to arginine (K445R) impaired Myocardin transactivation. PIAS1 was involved in governing Myocardin activity via its E3 ligase activity that stimulated Myocardin sumoylation on an atypical sumoylation site(s) and by its physical association with Myocardin. Myocardin initiated the expression of cardiac muscle-specified genes, such as those encoding cardiac α-actin and α-myosin heavy chain, in an SRF-dependent manner in 10T1/2 fibroblasts, but only in the presence of coexpressed SUMO-1/PIAS1. Thus, SUMO modification acted as a molecular switch to promote Myocardin's role in cardiogenic gene expression.
-
Myocardin induces cardiomyocyte hypertrophy
Circulation Research, 2006Co-Authors: Weibing Xing, Zhigao Wang, Eric N Olson, Shijie Li, Tongcun Zhang, Christopher L Antos, Yibin Wang, Dazhi WangAbstract:In response to stress signals, postnatal cardiomyocytes undergo hypertrophic growth accompanied by activation of a fetal gene program, assembly of sarcomeres, and cellular enlargement. We show that hypertrophic signals stimulate the expression and transcriptional activity of Myocardin, a cardiac and smooth muscle-specific coactivator of serum response factor (SRF). Consistent with a role for Myocardin as a transducer of hypertrophic signals, forced expression of Myocardin in cardiomyocytes is sufficient to substitute for hypertrophic signals and induce cardiomyocyte hypertrophy and the fetal cardiac gene program. Conversely, a dominant-negative mutant form of Myocardin, which retains the ability to associate with SRF but is defective in transcriptional activation, blocks cardiomyocyte hypertrophy induced by hypertrophic agonists such as phenylephrine and leukemia inhibitory factor. Myocardin-dependent hypertrophy can also be partially repressed by histone deacetylase 5, a transcriptional repressor of Myocardin. These findings identify Myocardin as a nuclear effector of hypertrophic signaling pathways that couples stress signals to a transcriptional program for postnatal cardiac growth and remodeling.
-
phenotypic modulation of smooth muscle cells through interaction of foxo4 and Myocardin
Developmental Cell, 2005Co-Authors: Zhigao Wang, Hiromi Yanagisawa, Eric N OlsonAbstract:Summary Smooth muscle cells (SMCs) modulate their phenotype between proliferative and differentiated states in response to physiological and pathological cues. Insulin-like growth factor-I stimulates differentiation of SMCs by activating phosphoinositide-3-kinase (PI3K)-Akt signaling. Foxo forkhead transcription factors act as downstream targets of Akt and are inactivated through phosphorylation by Akt. We show that Foxo4 represses SMC differentiation by interacting with and inhibiting the activity of Myocardin, a transcriptional coactivator of smooth muscle genes. PI3K/Akt signaling promotes SMC differentiation, at least in part, by stimulating nuclear export of Foxo4, thereby releasing Myocardin from its inhibitory influence. Accordingly, reduction of Foxo4 expression in SMCs by siRNA enhances Myocardin activity and SMC differentiation. We conclude that signal-dependent interaction of Foxo4 with Myocardin couples extracellular signals with the transcriptional program for SMC differentiation.
-
modulation of smooth muscle gene expression by association of histone acetyltransferases and deacetylases with Myocardin
Molecular and Cellular Biology, 2005Co-Authors: Zhigao Wang, Dazhi Wang, Shijie Li, James A Richardson, Weibing Xing, Jiyeon Oh, Chun Li Zhang, Eric N OlsonAbstract:Differentiation of smooth muscle cells is accompanied by the transcriptional activation of an array of muscle-specific genes controlled by serum response factor (SRF). Myocardin is a cardiac and smooth muscle-specific expressed transcriptional coactivator of SRF and is sufficient and necessary for smooth muscle gene expression. Here, we show that Myocardin induces the acetylation of nucleosomal histones surrounding SRF-binding sites in the control regions of smooth muscle genes. The promyogenic activity of Myocardin is enhanced by p300, a histone acetyltransferase that associates with the transcription activation domain of Myocardin. Conversely, class II histone deacetylases interact with a domain of Myocardin distinct from the p300-binding domain and suppress smooth muscle gene activation by Myocardin. These findings point to Myocardin as a nexus for positive and negative regulation of smooth muscle gene expression by changes in chromatin acetylation.
Paul B Herring - One of the best experts on this subject based on the ideXlab platform.
-
modulation of Myocardin function by the ubiquitin e3 ligase ubr5
Journal of Biological Chemistry, 2010Co-Authors: Guoqing Hu, Xiaobo Wang, Darren N Saunders, Michelle J Henderson, Amanda J Russell, Paul B Herring, Jiliang ZhouAbstract:Fully differentiated mature smooth muscle cells (SMCs) are characterized by the presence of a unique repertoire of smooth muscle-specific proteins. Although previous studies have shown Myocardin to be a critical transcription factor for stimulating expression of smooth muscle-specific genes, the mechanisms regulating Myocardin activity are still poorly understood. We used a yeast two-hybrid screen with Myocardin as bait to search for factors that may regulate the transcriptional activity of the Myocardin. From this screen we identified a HECT domain-containing protein UBR5 (ubiquitin protein ligase E3 component n-recognin 5) as a Myocardin-binding protein. Previous studies have shown that HECT domain-containing proteins are ubiquitin E3 ligases that play an important role in protein degradation. UBR5 has, however, also been shown to regulate transcription independent of its E3 ligase activity. In the current study we demonstrated that UBR5 localized in the nuclei of SMCs and forms a complex with Myocardin in vivo and in vitro. We also show that UBR5 specifically enhanced trans-activation of smooth muscle-specific promoters by the Myocardin family of proteins. In addition, UBR5 significantly augmented the ability of Myocardin to induce expression of endogenous SMC marker genes independent on its E3 ligase function. Conversely, depletion of endogenous UBR5 by small interfering RNA in fibroblast cells attenuated Myocardin-induced smooth muscle-specific gene expression, and UBR5 knockdown in SMCs resulted in down-regulation of smooth muscle-specific genes. Furthermore, we found that UBR5 can attenuate Myocardin protein degradation resulting in increased Myocardin protein expression without affecting Myocardin mRNA expression. The effects of UBR5 on Myocardin requires only the HECT and UBR1 domains of UBR5. This study reveals an unexpected role for the ubiquitin E3 ligase UBR5 as an activator of smooth muscle differentiation through its ability to stabilize Myocardin protein.
-
the swi snf chromatin remodeling complex regulates Myocardin induced smooth muscle specific gene expression
Arteriosclerosis Thrombosis and Vascular Biology, 2009Co-Authors: Jiliang Zhou, Min Zhang, Hong Fang, Omar Elmounayri, Jennifer M Rodenberg, Anthony N Imbalzano, Paul B HerringAbstract:Objective— Regulatory complexes comprising Myocardin and serum response factor (SRF) are critical for the transcriptional regulation of many smooth muscle–specific genes. However, little is known about the epigenetic mechanisms that regulate the activity of these complexes. In the current study, we investigated the role of SWI/SNF ATP-dependent chromatin remodeling enzymes in regulating the myogenic activity of Myocardin. Methods and Results— We found that both Brg1 and Brm are required for maintaining expression of several smooth muscle–specific genes in primary cultures of aortic smooth muscle cells. Furthermore, the ability of Myocardin to induce expression of smooth muscle–specific genes is abrogated in cells expressing dominant negative Brg1. In SW13 cells, which lack endogenous Brg1 and Brm1, Myocardin is unable to induce expression of smooth muscle–specific genes. Whereas, reconstitution of wild-type, or bromodomain mutant forms Brg1 or Brm1, into SW13 cells restored their responsiveness to Myocardin. SWI/SNF complexes were found to be required for Myocardin to increase SRF binding to the promoters of smooth muscle–specific genes. Brg1 and Brm directly bind to the N terminus of Myocardin, in vitro, through their ATPase domains and Brg1 forms a complex with SRF and Myocardin in vivo in smooth muscle cells. Conclusion— These data demonstrate that the ability of Myocardin to induce smooth muscle–specific gene expression is dependent on its interaction with SWI/SNF ATP-dependent chromatin remodeling complexes.
-
mechanisms responsible for the promoter specific effects of Myocardin
Journal of Biological Chemistry, 2005Co-Authors: Jiliang Zhou, Paul B HerringAbstract:Abstract Understanding the mechanism of smooth muscle cell (SMC) differentiation will provide the foundation for elucidating SMC-related diseases such as atherosclerosis, restenosis, and asthma. Recent studies have demonstrated that the interaction of SRF with the co-activator Myocardin is a critical determinant of smooth muscle development. It has been proposed that the specific transcriptional activation of smooth muscle-restricted genes (as opposed to other SRF-dependent genes) by Myocardin results from the presence of multiple CArG boxes in smooth muscle genes that facilitate Myocardin homodimer formation. This proposal was further tested in the current study. Our results show that the SMC-specific telokin promoter, which contains only a single CArG box, is strongly activated by Myocardin. Furthermore, Myocardin and a dimerization defective mutant Myocardin induce expression of endogenous telokin but not c-fos in 10T1/2 fibroblast cells. Knocking down Myocardin by small interfering RNA decreased telokin promoter activity and expression in A10 SMCs. A series of telokin and c-fos promoter chimeric and mutant reporter genes was generated to determine the mechanisms responsible for the promoter-specific effects of Myocardin. Data from these experiments demonstrated that the ets binding site in the c-fos promoter partially blocks the activation of this promoter by Myocardin. However, the binding of ets factors alone was not sufficient to explain the promoter-specific effects of Myocardin. Elements 3′ of the CArG box in the c-fos promoter act in concert with the ets binding site to block the ability of Myocardin to activate the promoter. Conversely, elements 5′ and 3′ of the CArG box in the telokin promoter act in concert with the CArG box to facilitate Myocardin stimulation of the promoter. Together these data suggest that the promoter specificity of Myocardin is dependent on complex combinatorial interactions of multiple cis elements and their trans binding factors.
