Identification of proteins associating with GPI-anchored T-cadherin on the surface of vascular endothelial cells: the role for Grp78/BiP in T-cadherin-dependent cell survival

Department of Research, Cardiovascular Laboratories, Basel University Hospital, Hebelstrasse 20, CH 4031 Basel, Switzerland; Department of Vascular Biology and Thrombosis Research, Medical University of Vienna, Schwarzspanierstrasse 17, 1090 Vienna, Austria; Division of Cardiology, Luzern Kantonsspital, CH 6000 Luzern, Switzerland

^* To whom correspondence should be addressed. Email: maria.filippova{at}unibas.ch.

	Abstract

There is scant knowledge regarding how cell surface lipid-anchored T-cadherin (T-cad) transmits signals through the plasma membrane to its intracellular targets. This study aimed to identify membrane proteins colocalizing with atypical GPI-anchored T-cad on the surface of endothelial cells and to evaluate their role as signaling adaptors for T-cad. Application of coimmunoprecipitation from endothelial cells expressing c-myc-tagged T-cad and HPLC revealed putative association of T-cad with the following proteins: glucose-related protein GRP78, GABA-A receptor 1 subunit, integrin ₃ and two hypothetical proteins LOC124245 and FLJ32070. Association of Grp78 and integrin ₃ with T-cad on the cell surface was confirmed by surface biotinylation and reciprocal immunoprecipitation and by confocal microscopy. Use of anti-Grp78 blocking antibodies, Grp78 siRNA and coexpression of constitutively active Akt demonstrated an essential role for surface Grp78 in T-cad-dependent survival signal transduction via Akt in endothelial cells. The findings herein are relevant in the context of both the identification of transmembrane signaling partners for GPI-anchored T-cad, as well as the demonstration of a novel mechanism whereby Grp78 can influence endothelial cell survival as a cell surface signaling receptor rather than an intracellular chaperone.