DESCRIPTION (provided by applicant): Oxidative stress may be the single most prevalent cause of neuronal dysfunction in neurodegenerative disorders, and its prevalence underscores the need to clarify mechanisms causing and attenuating the deleterious effects, the overall goals of this project. We report exciting and novel results: (1) DNA damaging agents that elevate p53 cause a novel mechanism of activation of the pro-apoptotic glycogen synthase kinase-3b (GSK3b). (2) Oxidative stress induces RGS2 (Regulator of G-protein Signaling 2) expression, a G-protein GTPase-activating protein, providing a mechanistic basis for impaired signaling. (3) Stimulation of muscarinic receptors greatly attenuates oxidative stress-induced apoptosis, remarkably as effectively as a general caspase inhibitor. These results provide important new insights about mechanisms that contribute to oxidative stress-induced impairments and about mechanisms capable of attenuating the deleterious effects. Specific Aim 1 will test the hypothesis that oxidative stress and DNA damage activate p53-mediated signaling encompassing recruitment of GSK3b by a novel activation mechanism. We will test the hypotheses that p53-induced activation of GSK3b leads to inhibition of survival-promoting transcription factor substrates of GSK3b, and promotes responses to p53, identify the p53-binding domain on GSK3b, determine if p53 binding alters the association of GSK3b with other proteins, identify the GSK3b-binding domain on p53 and determine if GSK3b binding alters p53 functions. Specific Aim 2 will test the hypothesis that oxidative stress and DNA damage induce the expression of RGS2 which attenuates muscarinic receptor-coupled signaling and facilitates oxidative stress-induced apoptosis. We will identify the signal mediating H202-induced increases in RGS2, Determine if H202-induced increases in RGS2 impair muscarinic receptor-coupled signaling, and test if IGS2 expression is pro-apoptotic role after oxidative stress. Specific Aim 3 will test the hypothesis that stimulated muscarinic receptors protect cells from oxidative stress, identify the blocked site in -1202-inducedsignaling, test the hypothesis that muscarinic receptors provide protection from other apoptotic conditions, identify the signaling pathways activated by muscarinic receptors providing protection, and test the hypothesis that activation of Rho family small G-proteins is protective.