Transforming growth factor-beta-stimulated endocardial cell transformation is dependent on Par6c regulation of RhoA.
Valvular heart disease due to congenital abnormalities or pathology is a major cause of mortality and morbidity. Understanding the cellular processes and molecules that regulate valve formation and remodeling is required to develop effective therapies. In the developing heart, epithelial-mesenchymal transformation (EMT) in a subpopulation of endocardial cells in the atrioventricular cushion (AVC) is an important step in valve formation. Transforming growth factor-beta (TGFbeta) has been shown to be an important regulator of AVC endocardial cell EMT in vitro and mesenchymal cell differentiation in vivo. Recently Par6c (Par6) has been shown to function downstream of TGFbeta to recruit Smurf1, an E3 ubiquitin ligase, which targets RhoA for degradation to control apical-basal polarity and tight junction dissolution. We tested the hypothesis that Par6 functions in a pathway that regulates endocardial cell EMT. Here we show that the Type I TGFbeta receptor ALK5 is required for endocardial cell EMT. Overexpression of dominant negative Par6 inhibits EMT in AVC endocardial cells, whereas overexpression of wild-type Par6 in normally non-transforming ventricular endocardial cells results in EMT. Overexpression of Smurf1 in ventricular endocardial cells induces EMT. Decreasing RhoA activity using dominant negative RhoA or small interfering RNA in ventricular endocardial cells also increases EMT, whereas overexpression of constitutively active RhoA in AVC endothelial cells blocks EMT. Manipulation of Rac1 or Cdc42 activity is without effect. These data demonstrate a functional role for Par6/Smurf1/RhoA in regulating EMT in endocardial cells.
Townsend
Todd A
TA
Department of Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37232-6600, USA.
Wrana
Jeffrey L
JL
Davis
George E
GE
Barnett
Joey V
JV
D000818:Animals; D002642:Chick Embryo; D003094:Collagen / Q000737:chemistry; D004699:Endocardium / Q000166:cytology* / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D005799:Genes, Dominant; D006352:Heart Ventricles / Q000378:metabolism*; D008024:Ligands; D008648:Mesoderm / Q000378:metabolism; D008954:Models, Biological; D034741:RNA, Small Interfering / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
D008024:Ligands; D034741:RNA, Small Interfering; D016212:Transforming Growth Factor beta; D003094:Collagen; C498342:SMURF1 protein, human; D044767:Ubiquitin-Protein Ligases; D020764:cdc42 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein