PROJECT SUMMARY Hydrogen sulfide (H2S) is a gaseous signaling molecule which has neuroprotective effects. H2S is generated endogenously from cysteine and/or homocysteine via the reverse transsulfuration pathway. H2S production is diminished in Alzheimer's disease (AD), the most common cause of dementia in the elderly, and H2S donors have been reported to confer beneficial effects. However, the mechanistic basis for this decrease has still not been elucidated. H2S regulates several physiological processes via a posttranslational modification designated as sulfhydration, wherein the -SH groups of reactive cysteine residues are converted to persulfide or -SSH groups. Prior studies conducted by us and others show that during aging, which is the biggest risk factor for developing AD, sulfhydration is decreased in the brain. Preliminary studies conducted by us show that Tau, the microtubule associated protein, and a component of neurofibrillary tangles in AD, binds cystathionine γ-lyase (CSE), one of the biosynthetic enzymes for H2S and activates it. In this study we propose to characterize the interaction of the three biosynthetic enzymes for H2S, CSE, cystathionine β-synthase (CBS) and 3-mercaptopyruvate sulfurtransferase (3-MST), both in vitro in cell culture as well as in vivo in the mouse brain. We also seek to elucidate the effects of H2S on the activity of Tau. In Aim 1, we will study the binding of CSE, CBS and 3-MST to wild type Tau and its isoforms. In Aim 2, we will evaluate the sulfhydration status of Tau and map the cysteine residues on tau which are sulfhydrated. Deciphering the signaling functions of the H2S/Tau pathway will not only identify basic physiological functions of tau, but also pinpoint nodes for therapeutic intervention in AD, which are also relevant for other tauopathies.