title: Endocardial cell epithelial-mesenchymal transformation requires Type III TGFβ receptor interaction with GIPC.
abstract: An early event in heart valve formation is the epithelial-mesenchymal transformation (EMT) of a subpopulation of endothelial cells in specific regions of the heart tube, the endocardial cushions. The Type III TGFβ receptor (TGFβR3) is required for TGFβ2- or BMP-2-stimulated EMT in atrioventricular endocardial cushion (AVC) explants in vitro but the mediators downstream of TGFβR3 are not well described. Using AVC and ventricular explants as an in vitro assay, we found an absolute requirement for specific TGFβR3 cytoplasmic residues, GAIP-interacting protein, C terminus (GIPC), and specific Activin Receptor-Like Kinases (ALK)s for TGFβR3-mediated EMT when stimulated by TGFβ2 or BMP-2. The introduction of TGFβR3 into nontransforming ventricular endocardial cells, followed by the addition of either TGFβ2 or BMP-2, results in EMT. TGFβR3 lacking the entire cytoplasmic domain, or only the 3C-terminal amino acids that are required to bind GIPC, fails to support EMT in response to TGFβ2 or BMP-2. Overexpression of GIPC in AVC endocardial cells enhanced EMT while siRNA-mediated silencing of GIPC in ventricular cells overexpressing TGFβR3 significantly inhibited EMT. Targeting of specific ALKs by siRNA revealed that TGFβR3-mediated EMT requires ALK2 and ALK3, in addition to ALK5, but not ALK4 or ALK6. Taken together, these data identify GIPC, ALK2, ALK3, and ALK5 as signaling components required for TGFβR3-mediated endothelial cell EMT.
authors.affiliation: Department of Pharmacology, Vanderbilt University Medical Center, Nashville, TN 37232-6600, USA. Todd.Townsend@Vanderbilt.edu
authors.forename: Todd A
authors.initials: TA
authors.lastname: Townsend
authors.forename: Jamille Y
authors.initials: JY
authors.lastname: Robinson
authors.forename: Tam
authors.initials: T
authors.lastname: How
authors.forename: Daniel M
authors.initials: DM
authors.lastname: DeLaughter
authors.forename: Gerard C
authors.initials: GC
authors.lastname: Blobe
authors.forename: Joey V
authors.initials: JV
authors.lastname: Barnett
mesh_terms: D029404:Activin Receptors / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D055396:Bone Morphogenetic Protein 2 / Q000494:pharmacology / Q000502:physiology; D002642:Chick Embryo; D054089:Endocardial Cushions / Q000166:cytology / Q000502:physiology*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D058750:Epithelial-Mesenchymal Transition; D049452:Green Fluorescent Proteins / Q000096:biosynthesis; D008969:Molecular Sequence Data; D054730:Protein Interaction Domains and Motifs; D011509:Proteoglycans / Q000737:chemistry / Q000378:metabolism*; D018125:Receptors, Transforming Growth Factor beta / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis; D046509:Tissue Culture Techniques; D053781:Transforming Growth Factor beta2 / Q000494:pharmacology / Q000502:physiology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D055396:Bone Morphogenetic Protein 2; D011509:Proteoglycans; D018125:Receptors, Transforming Growth Factor beta; D011994:Recombinant Proteins; D053781:Transforming Growth Factor beta2; C071736:betaglycan; D049452:Green Fluorescent Proteins; D029404:Activin Receptors