Role of G-Proteins in Mechanical Signal Transduction

Abstract

Cells have highly specialized forced sensing mechanisms such as those involved in hearing and touch which can convert a mechanical stimulus into electrical and biochemical responses. Fluid flow itself is another such stimulus that exists in the cellular environment. Indirect evidence suggests that flow regulation of vascular tone via the endothelium and smooth muscle cells involves mechanical signal transduction via G proteins. Human Umbilical Vein Endothelial cells (hUVECs) grown on glass slides were subjected to fluid flow in parallel plate flow chambers. Fluid shear-induced GTP binding in hUVECs was assayed by preincubating the cells with a GTP photoreactive analogue, 8-azido GTP (α-32_p) and subjecting to UV crosslinking after fluid flow (10 dynes/cm²) for 10 sec. Autoradiographic analysis of these cells lysate show a 42 kDa protein being labeled by GTP upon flow stimulation. Further this protein was identified with anti Gα subunits Gαq and Gαi3 may be involved in the flow induced signal transduction pathway in endothelial cells. In other supporting experiments, Gproteins purified from bovine brain were reconstituted into PE:PS liposomes and subjected to fluid shear. There was an increase in Gprotein mediated GTP hydolysis upon the onset of shear. These experiments suggest the involvement of Gproteins as mechanochemical signal transducers. This work is supported by HL40696-07.