Abstract
More than 30 mutations in ACTA2, which encodes α-smooth muscle actin, have been identified to cause autosomal dominant thoracic aortic aneurysm and dissection. The mutation R256H is of particular interest because it also causes patent ductus arteriosus and moyamoya disease. R256H is one of the more prevalent mutations and, based on its molecular location near the strand-strand interface in the actin filament, may affect F-actin stability. To understand the molecular ramifications of the R256H mutation, we generated Saccharomyces cerevisiae yeast cells expressing only R256H yeast actin as a model system. These cells displayed abnormal cytoskeletal morphology and increased sensitivity to latrunculin A. After cable disassembly induced by transient exposure to latrunculin A, mutant cells were delayed in reestablishing the actin cytoskeleton. In vitro, mutant actin exhibited a higher than normal critical concentration and a delayed nucleation. Consequently, we investigated regulation of mutant actin by formin, a potent facilitator of nucleation and a protein needed for normal vascular smooth muscle cell development. Mutant actin polymerization was inhibited by the FH1-FH2 fragment of the yeast formin, Bni1. This fragment strongly capped the filament rather than facilitating polymerization. Interestingly, phalloidin or the presence of wild type actin reversed the strong capping behavior of Bni1. Together, the data suggest that the R256H actin mutation alters filament conformation resulting in filament instability and misregulation by formin. These biochemical effects may contribute to abnormal histology identified in diseased arterial samples from affected patients.
Highlights
The biochemical mechanisms underlying ␣-smooth muscle actin-mediated vascular disease are unknown
Effect of Mutations on Cytoskeletal Morphology and Cell Growth—Our studies began with analyses of the actin cytoskeleton
We examined the sensitivity of wild type and R256H cells to latrunculin A, a drug that sequesters actin monomers and reversibly promotes rapid depolymerization of actin filaments [22, 35, 36]
Summary
The biochemical mechanisms underlying ␣-smooth muscle actin-mediated vascular disease are unknown. More than 30 mutations in ACTA2, which encodes ␣-smooth muscle actin, have been identified to cause autosomal dominant thoracic aortic aneurysm and dissection. We investigated regulation of mutant actin by formin, a potent facilitator of nucleation and a protein needed for normal vascular smooth muscle cell development. The data suggest that the R256H actin mutation alters filament conformation resulting in filament instability and misregulation by formin These biochemical effects may contribute to abnormal histology identified in diseased arterial samples from affected patients. Mutations in ␣-smooth muscle actin, encoded by ACTA2, are the most common genetic cause of familial thoracic aortic. The single actin isoform in yeast and ability to correlate in vivo findings with in vitro analyses have allowed us to successfully use this system to understand the biochemical mechanisms underlying TAAD [13] but other actinmediated diseases (14 –16). We describe analyses of formin regulation of mutant actin filament formation
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
