PIAS1 activates the expression of smooth muscle cell differentiation marker genes by interacting with serum response factor and class I basic helix-loop-helix proteins.
Although a critical component of vascular disease is modulation of the differentiated state of vascular smooth muscle cells (SMC), the mechanisms governing SMC differentiation are relatively poorly understood. We have previously shown that E-boxes and the ubiquitously expressed class I basic helix-loop-helix (bHLH) proteins, including E2-2 and E12, are important in regulation of the SMC differentiation marker gene, the SM alpha-actin gene. The aim of the present study was to identify proteins that bind to class I bHLH proteins in SMC and modulate transcriptional regulation of SMC differentiation marker genes. Herein we report that members of the protein inhibitor of activated STAT (PIAS) family interact with class I bHLH factors as well as serum response factor (SRF). PIAS1 interacted with E2-2 and E12 based on yeast two-hybrid screens, mammalian two-hybrid assays, and/or coimmunoprecipitation assays. Overexpression of PIAS1 significantly activated the SM alpha-actin promoter and mRNA expression, as well as SM myosin heavy chain and SM22alpha, whereas a small interfering RNA for PIAS1 decreased activity of these promoters, as well as endogenous mRNA expression, and SRF binding to SM alpha-actin promoter within intact chromatin in cultured SMC. Of significance, PIAS1 bound to SRF and activated SM alpha-actin promoter expression in wild-type but not SRF(-/-) embryonic stem cells. These results provide novel evidence that PIAS1 modulates transcriptional activation of SMC marker genes through cooperative interactions with both SRF and class I bHLH proteins.
Kawai-Kowase
Keiko
K
Department of Molecular Physiology and Biological Physics, University of Virginia, 415 Lane Road, MR5, Room 1220, P.O. Box 801394, Charlottesville, VA 22908, USA.
Kumar
Meena S
MS
Hoofnagle
Mark H
MH
Yoshida
Tadashi
T
Owens
Gary K
GK
D000199:Actins / Q000235:genetics; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins; D002454:Cell Differentiation / Q000235:genetics; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015536:Down-Regulation / Q000235:genetics; D015870:Gene Expression; D005819:Genetic Markers / Q000235:genetics; D006801:Humans; D051379:Mice; D009124:Muscle Proteins / Q000235:genetics*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D050828:Protein Inhibitors of Activated STAT; D034741:RNA, Small Interfering / Q000235:genetics / Q000494:pharmacology; D051381:Rats; D026362:Serum Response Factor / Q000378:metabolism*; D025841:Small Ubiquitin-Related Modifier Proteins / Q000235:genetics / Q000378:metabolism*; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects / Q000235:genetics; D020798:Two-Hybrid System Techniques; D029961:Zebrafish Proteins
D000199:Actins; D051792:Basic Helix-Loop-Helix Proteins; D004268:DNA-Binding Proteins; D005819:Genetic Markers; D009124:Muscle Proteins; D050828:Protein Inhibitors of Activated STAT; D034741:RNA, Small Interfering; D026362:Serum Response Factor; D025841:Small Ubiquitin-Related Modifier Proteins; D051820:TCF Transcription Factors; D058125:Transcription Factor 7-Like 1 Protein; D058118:Transcription Factor 7-Like 2 Protein; D014157:Transcription Factors; D029961:Zebrafish Proteins; C498765:HAND2 protein, human; C095572:Hand1 protein, mouse; C498766:Hand2 protein, mouse; C498767:Hand2 protein, rat; C106529:PIAS1 protein, human; C544912:TCF7L1 protein, human; C544894:TCF7L2 protein, human; C544913:Tcf7l1 protein, mouse; C553060:Tcf7l1 protein, rat; C544895:Tcf7l2 protein, mouse; C553059:Tcf7l2 protein, rat; C410915:hand2 protein, zebrafish; C097341:helix-loop-helix protein, eHAND