Regulation of CBP by Synaptic Activity

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DESCRIPTION (provided by applicant): Activity-regulated gene expression plays a fundamental role in both the remodeling of synaptic circuitry and in neuronal survival. Activity-dependent increases in intracellular Ca2+ trigger the expression of hundreds or perhaps thousands of genes. The cAMP response element (CRE) was identified as a major Ca2+ responsive element via analysis of immediate early gene (IEGs) promoters and many if not most Ca2+ responsive promoters contain CREs. The basic leucine zipper transcription factor, CREB, binds to the CRE as a dimer and can be activated by Ca2+, cAMP/PKA, and growth factor signaling pathways. The transcription factor CREB is a major target of activity-induced Ca2+ influx and is a key regulator of both neuronal survival and adaptive synaptic plasticity. Nevertheless, the mechanisms by which Ca2+ activates CREB have not been clearly elucidated. It is well established that PKA phosphorylates Ser 133 of CREB and creates a phospho-serine docking motif that recruits its coactivator CBP (or the homologue, p300). Recruitment of CBP is thought to enhance transcriptional activation either via its association with the general transcriptional machinery or via its intrinsic histone acetyltransferase activity. Although activity-induced Ca2+ influx is widely believed to regulate CREB function in an analogous manner, the signaling cascades that promote CBP recruitment and transcriptional activation have not been clearly defined. A major theme of this proposal is to determine whether the PKA-CREB-CBP paradigm also pertains to Ca2+-activated transcription. Thus, an initial goal of this study will be to determine whether neuronal activity induces the recruitment of CBP to CREB and whether CBP recruitment is required for transcriptional activation. Four specific aims are proposed. (1) Determine whether synaptic activity induces the recruitment of CBP to CREB in neurons and whether recruitment of CBP is sufficient for full transcriptional activation. (2) Decipher the activity-regulated signaling cascades that promote the phosphorylation and activation of CREB and CBP. (3) Determine how synaptic activity and Ca2+ influx regulate CBP function. (4) Analyze the regulation of CBP and CREB function in mice deficient for CaM kinase IV. This study seeks to dissect the biochemical events result in synaptic activity-mediated phosphorylation of CREB, recruitment of CBP, and "activation" of CBP-dependent transcription. Gaining insight into the mechanisms of activity-mediated transcription has relevance in the treatment of neurodegenerative pathologies that may result from excitotoxic cell death or apoptosis. Through a mentored career development plan, the Principal Investigator will gain an enhanced capacity to be productive in the field of molecular neurobiology with a focus on gene regulation. This will be accomplished by conducting the above described research, attending and presenting at local and national seminars and meetings, and participating in courses teaching molecular biology and neurophysiology techniques.