Khan Mohammad Ashhar I, Chirasani Venkat R, Sarker Muzaddid, McCormick Laura, Campbell Sharon L
Department of Biochemistry & Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
Department of Biochemistry & Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA; R. L. Juliano Structural Bioinformatics Core, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
J Biol Chem. 2025 May 14;301(6):110232. doi: 10.1016/j.jbc.2025.110232.
Vinculin (Vcn) and its splice variant metavinculin (MVcn) are cell adhesion proteins that regulate cell morphology, adhesion, and motility. They function as scaffold proteins that anchor membrane receptors to filamentous actin (F-actin) at focal adhesions and cell-cell junctions. MVcn bears an extra 68 amino acid insert in the tail domain and is selectively expressed in cardiac and smooth muscle cells at substoichiometric levels relative to Vcn. Mutations in the MVcn tail domain (MVt) promote cardiomyopathy, yet how these mutations alter ligand interactions to promote defects in force transduction and reduced blood flow is unclear. One difference between Vcn and MVcn lies in the ability to reorganize F-actin, with MVcn negatively regulating Vcn-mediated F-actin bundling. Vcn associates with phosphatidylinositol 4,5-bisphosphate (PIP) through its tail domain (Vt) to drive recruitment, activation, and focal adhesion turnover. However, it remains unclear whether MVcn specifically associates with PIP-containing membranes and how such interactions might influence its functional interplay with Vcn in tissues where both isoforms coexist. To evaluate the interaction of MVt and MVt cardiomyopathy mutants with PIP membranes in comparison with Vt, we conducted mutagenesis, phospholipid-association assays, and computational modeling. We found that MVt shows reduced association for PIP-containing liposomes relative to Vt due to sequence differences within the insert region. Moreover, mutations in MVt that promote cardiomyopathies do not affect PIP-dependent lipid association. These findings suggest that MVcn differs from Vcn in driving PIP-mediated membrane association and sheds light on the coordinate role of Vcn and MVcn in membrane association as well as MVcn cardiomyopathy defects.
纽蛋白（Vcn）及其剪接变体元纽蛋白（MVcn）是调节细胞形态、黏附及运动的细胞黏附蛋白。它们作为支架蛋白，在黏着斑和细胞间连接中将膜受体锚定到丝状肌动蛋白（F-肌动蛋白）上。MVcn在尾部结构域有一个额外的68个氨基酸的插入片段，并且在心肌和平滑肌细胞中相对于Vcn以亚化学计量水平选择性表达。MVcn尾部结构域（MVt）的突变会引发心肌病，但这些突变如何改变配体相互作用以促进力传导缺陷和减少血流量尚不清楚。Vcn和MVcn的一个区别在于重组F-肌动蛋白的能力，MVcn对Vcn介导的F-肌动蛋白束形成起负调节作用。Vcn通过其尾部结构域（Vt）与磷脂酰肌醇4,5-二磷酸（PIP）结合，以驱动募集、激活和黏着斑周转。然而，尚不清楚MVcn是否特异性地与含PIP的膜结合，以及这种相互作用在两种异构体共存的组织中如何影响其与Vcn的功能相互作用。为了评估MVt和MVt心肌病突变体与含PIP膜的相互作用并与Vt进行比较，我们进行了诱变、磷脂结合测定和计算建模。我们发现，由于插入区域内的序列差异，MVt相对于Vt与含PIP脂质体的结合减少。此外，促进心肌病的MVt突变并不影响PIP依赖性脂质结合。这些发现表明，MVcn在驱动PIP介导的膜结合方面与Vcn不同，并揭示了Vcn和MVcn在膜结合中的协同作用以及MVcn心肌病缺陷。
