Functional validation reveals the novel missense V419L variant in TGFBR2 associated with Loeys-Dietz syndrome (LDS) impairs canonical TGF-β signaling. Cold Spring Harb Mol Case Stud 2017 Jul;3(4)
Date
07/07/2017Pubmed ID
28679693Pubmed Central ID
PMC5495030DOI
10.1101/mcs.a001727Scopus ID
2-s2.0-85037598912 (requires institutional sign-in at Scopus site) 7 CitationsAbstract
TGF-β-related heritable connective tissue disorders are characterized by a similar pattern of cardiovascular defects, including aortic root dilatation, mitral valve prolapse, vascular aneurysms, and vascular dissections and exhibit incomplete penetrance and variable expressivity. Because of the phenotypic overlap of these disorders, panel-based genetic testing is frequently used to confirm the clinical findings. Unfortunately in many cases, variants of uncertain significance (VUSs) obscure the genetic diagnosis until more information becomes available. Here, we describe and characterize the functional impact of a novel VUS in the TGFBR2 kinase domain (c.1255G>T; p.Val419Leu), in a patient with the clinical diagnosis of Marfan syndrome spectrum. We assessed the structural and functional consequence of this VUS using molecular modeling, molecular dynamic simulations, and in vitro cell-based assays. A high-quality homology-based model of TGFBR2 was generated and computational mutagenesis followed by refinement and molecular dynamics simulations were used to assess structural and dynamic changes. Relative to wild type, the V419L induced conformational and dynamic changes that may affect ATP binding, increasing the likelihood of adopting an inactive state, and, we hypothesize, alter canonical signaling. Experimentally, we tested this by measuring the canonical TGF-β signaling pathway activation at two points; V419L significantly delayed SMAD2 phosphorylation by western blot and significantly decreased TGF-β-induced gene transcription by reporter assays consistent with known pathogenic variants in this gene. Thus, our results establish that the V419L variant leads to aberrant TGF-β signaling and confirm the diagnosis of Loeys-Dietz syndrome in this patient.
Author List
Cousin MA, Zimmermann MT, Mathison AJ, Blackburn PR, Boczek NJ, Oliver GR, Lomberk GA, Urrutia RA, Deyle DR, Klee EWAuthors
Gwen Lomberk PhD Professor in the Surgery department at Medical College of WisconsinAngela Mathison PhD Assistant Professor in the Surgery department at Medical College of Wisconsin
Michael T. Zimmermann PhD Director, Assistant Professor in the Clinical and Translational Science Institute department at Medical College of Wisconsin
MESH terms used to index this publication - Major topics in bold
AdultHumans
Loeys-Dietz Syndrome
Male
Marfan Syndrome
Mutation
Mutation, Missense
Phosphorylation
Receptors, Transforming Growth Factor beta
Signal Transduction
Smad2 Protein
Transforming Growth Factor beta
