Conformational changes of focal adhesion protein vinculin in response to mechanical stress using an equibiaxial stretching device

L. SIGAUT; C. VON BILDERLING; M. BIANCHI; L. GASTALDI; L. I. PIETRASANTA

San Javier, Tucumán

Congreso; Reunión Anual de la Sociedad Argentina de Biofísica; 2012

Sociedad Argentina de Biofísica

Mechanical forces play an important role in the organization, growth, maturation, and function of living tissues. Mechanotransduction, the process by which cells sense and respond to mechanical signals, is mediated by a complex network of different proteins. Evidence from cells in culture suggests that vinculin is involved in transducing force across cell membranes in regulating cell adhesion and motility. When cells adhere and spread on the extracellular matrix (ECM) a portion of the cytoplasmic vinculin is recruited to specialized sites on the plasma membrane called focal adhesions (FA). At these sites, dynamic connections are made between the actin cytoskeleton and ECM through transmembrane integrin receptors. In the presence of a mechanical stimulus, FAs are dynamically assembled and disassembled. In this work, we introduce a stretching device that allows both live fluorescence imaging and mechanical equibiaxial stretching of the cells cultured on silicone elastic membranes. Careful characterization of the device described herein shows its capability of delivering up to 12% of two dimensional homogeneous strain to silicone membranes. Using a Förster resonance energy transfer probe that reports on changes in vinculin?s conformation, expressed in epithelial mammalian living cells (HC11), we are able to study these conformational changes in response to stretching.