Tripartite interactions of PKA catalytic subunit and C-terminal domains of cardiac Ca2+ channel may modulate its β-adrenergic regulation

BackgroundThe beta-adrenergic augmentation of cardiac contraction, by increasing the conductivity of L-type voltage-gated CaV1.2 channels, is of great physiological and pathophysiological importance. Stimulation of beta-adrenergic receptors (beta AR) activates protein kinase A (PKA) through separation of regulatory (PKAR) from catalytic (PKAC) subunits. Free PKAC phosphorylates the inhibitory protein Rad, leading to increased Ca2+ influx. In cardiomyocytes, the core subunit of CaV1.2, CaV1.2 alpha 1, exists in two forms: full-length or truncated (lacking the distal C-terminus (dCT)). Signaling efficiency is believed to emanate from protein interactions within multimolecular complexes, such as anchoring PKA (via PKAR) to CaV1.2 alpha 1 by A-kinase anchoring proteins (AKAPs). However, AKAPs are inessential for beta AR regulation of CaV1.2 in heterologous models, and their role in cardiomyocytes also remains unclear.ResultsWe show that PKAC interacts with CaV1.2 alpha 1 in heart and a heterologous model, independently of Rad, PKAR, or AKAPs. Studies with peptide array assays and purified recombinant proteins demonstrate direct binding of PKAC to two domains in CaV1.2 alpha 1-CT: the proximal and distal C-terminal regulatory domains (PCRD and DCRD), which also interact with each other. Data indicate both partial competition and possible simultaneous interaction of PCRD and DCRD with PKAC. The beta AR regulation of CaV1.2 alpha 1 lacking dCT (which harbors DCRD) was preserved, but subtly altered, in a heterologous model, the Xenopus oocyte.ConclusionsWe discover direct interactions between PKAC and two domains in CaV1.2 alpha 1. We propose that these tripartite interactions, if present in vivo, may participate in organizing the multimolecular signaling complex and fine-tuning the beta AR effect in cardiomyocytes.