Disruption of the plasma membrane integrity by cholesterol depletion impairs effectiveness of TRH receptor-mediated signal transduction via G(q)/G(11)alpha proteins.
We monitored the radioligand-binding characteristics of thyrotropin-releasing hormone (TRH) receptors, functional activity of G(q/11)alpha proteins, and functional status of the whole signaling cascade in HEK293 expressing high levels of TRH receptors and G(11)alpha. Our analyses indicated that disruption of plasma membrane microdomains by cholesterol depletion did not markedly influence the binding parameters of TRH receptors, but it altered efficacy of signal transduction. The functional coupling between TRH receptor and G(q/11)alpha was assessed by agonist-stimulated [(35)S]GTPgammaS binding, and results of these measurements pointed out to significantly lower potency of TRH to mediate G protein activation in the plasma membrane fraction isolated from cholesterol-depleted cells; there was a shift in sensitivity by one order of magnitude to the higher concentrations. A markedly lower sensitivity to stimulation with TRH was also observed in our experiments dealing with determination of hormone-induced Ca(2+) response. These data suggest that the intact structure of plasma membranes is an important optimum signal transduction initiated by TRH receptors and mediated by G(q/11)alpha proteins.
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D002784:Cholesterol / Q000502:physiology*; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D008024:Ligands; D021962:Membrane Microdomains / Q000378:metabolism*; D018025:Receptors, Thyrotropin-Releasing Hormone / Q000378:metabolism*; D015398:Signal Transduction; D013973:Thyrotropin-Releasing Hormone / Q000378:metabolism*
D008024:Ligands; D018025:Receptors, Thyrotropin-Releasing Hormone; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D013973:Thyrotropin-Releasing Hormone; D002784:Cholesterol; D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D002118:Calcium