T lymphocyte receptors for antigen, the alpha beta/T3 complex, play critical roles in generation and regulation of immune responses. First, by virtue of clonally-distributed specificity for antigen, these receptors dictate the specificity of the immune response. Second, they act as signal transducers which upon antigen binding mediate the activation of proto-oncogenes and genes encoding IL2 and IL2 receptors. Available evidence indicAtes that the alpha beta/T3 complex transduces signals via the activation of phosphoinositide hydrolysis yielding diacylglycerol and inositol trisphosphate second messenger moleculars. Although the molecular bAsis of alpha beta/T3 coupling to phosphoinositide hydrolysis is unknown, available evidence suggests that a GTP-binding protein may be involved. The multiple components of T3 have extensive hydrophilic carboxyl terminal sequences which extend into the cytoplasm and thus could interact directly with elements of second messenger generating systems such as GTP-binding proteins. Phosphorylation of T3 components following receptor ligation is consistent with the notion that T3 components are involved in receptor coupling to phosphoinositide hydrolysis. We have recently developed an isolated plasma membrane system in which alpha beta/T3 coupling to phosphoinositide hydrolysis can be demonstrated. We propose to use this system, which allows access to both faces of the plasma membrane, to study the role of specific T3 components in this coupling. We will assess the ability of antibodies against cytosolic portions of T3 components to block stimulus-response coupling. In addition, we will determine the modulation on coupling. Finally, we will utilize the system to compare signaling in mature T cells and mature thymocytes to signaling in immature thymocytes, which is impaired. The proposed studies should yield important insight into the molecular basis of alpha beta/T3 mediAted signal transduction.
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