The overall objective of this proposal is to understand molecular mechanisms that govern the duration of G protein signaling in photoreceptor cells. The photoreceptor G protein, transducin, plays a central role in vertebrate phototransduction where it conveys a signal from the activated receptor, photoexcited rhodopsin, to its effector, cGMP phosphodiesterase. Transducin is activated when photoexcited rhodopsin catalyses an exchange of GDP for GTP on the transducin alpha subunit and continues to stimulate PDE activity until the bound GTP is hydrolyzed. Studies conducted by this and other laboratories have shown that photoreceptors contain a multi-protein complex, which accelerates the slow intrinsic GTPase activity of transducin to a rate sufficient for timely photoresponse recovery. This complex includes the ninth member of the Regulators of G protein Signaling protein family (RGS9-1), type 5 G protein beta-subunit (Gbeta5L) and their membrane anchor (R9AP). Understanding the exact molecular mechanisms by which members of this protein ensemble contribute to the activation of transducin GTPase in a highly coordinated manner is the major goal of this proposal.
Aim 1 is to establish the mechanism by which R9AP potentiates the ability of the entire complex to activate transducin GTPase.
Aim 2 is to understand the nature of protein-protein interactions between RGS9-1, Gbeta5L and R9AP.
Aim 3 is to search for additional proteins interacting with the components of the GTPase activating complex in photoreceptors.
Aim 4 is to study phototransduction in rods from the animals where the content or composition of the GTPase activating complex is genetically modified. Addressing this problem is important not only for the understanding of the molecular mechanism of photoresponse recovery, but also for establishing general molecular principles of signal duration regulation in other intracellular signaling pathways. This work is relevant to understanding of how biochemical pathways responsible for ensuring normal photoreceptor activity may be affected in various inherited eye diseases.
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