AKAP79 Anchoring in L-type Calcium Channel Regulation

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DESCRIPTION (provided by applicant): CAMP-dependent kinase (PKA) phosphorylates the L-type calcium channel (LTCC) at a single serine residue in the C-terminus of the alpha1 subunit and in doing so increases the number of channels that will be activated by depolarization. In muscle PKA action on the LTCC is dependent upon its binding to A kinase anchoring proteins (AKAPs). In neurons LTCCs localize to similar subcellular locations as AKAP79, an AKAP that not only targets PKA to its substrates, but also the phosphatase calcineurin (CaN). The goal of the proposed research is to determine whether PKA and CaN together influence the activity of the LTCC in a manner that requires AKAP79 anchoring. I will disrupt AKAP79 anchoring of PKA and CaN and study how this affects the ability of these enzymes to regulate the channel by measuring changes in LTCC peak current using whole cell electrophysiology. Secondly, I will measure the rates of changes in intermolecular proximity between members of the channel-AKAP-PKA/-CaN complex using fluorescence resonance energy transfer and correlate these changes with LTCC regulation by simultaneously recording calcium channel currents.