Ca2+ responses to thyrotropin-releasing hormone and angiotensin II:: the role of plasma membrane integrity and effect of G11α protein overexpression on homologous and heterologous desensitization

The molecular mechanisms involved in GPCR-initiated signaling cascades where the two receptors share the same signaling cascade, such as thyrotropin-releasing hormone (TRH) and angiotensin II (ANG II), are still far from being understood. Here, we analyzed hormone-induced Ca2+ responses and the process of desensitization in HEK-293 cells, which express endogenous ANG II receptors. These cells were transfected to express exogenously high levels of TRH receptors (clone E2) or both TRH receptors and G, a protein (clone E2MII). We observed that the characteristics of the Ca2+ response, as well as the process of desensitization, were both strongly dependent on receptor number and G, Ice protein level. Whereas treatment of E2 cells with TRH or ANG II led to significant desensitization of the Ca2+ response to subsequent addition of either hormone, the response was not desensitized in E2MII cells expressing high levels of G, a. In addition, stimulation of both cell lines with THR elicited a clear heterologous desensitization to subsequent stimulation with ANG II. On the other hand, ANG II did not affect a subsequent response to TRH. ANG II-mediated signal transduction was strongly dependent on plasma membrane integrity modified by cholesterol depletion, but signaling through TRH receptors was altered only slightly under these conditions. It may be concluded that the level of expression of G-protein-coupled receptors and their cognate G-proteins strongly influences not only the magnitude of the Ca2+ response but also the process of desensitization and resistance to subsequent hormone addition. Copyright (c) 2007 John Wiley & Sons, Ltd.