| Summary: | Activity of cardiac Cav1.2 channels is enhanced by cyclic AMP-PKA signaling. In this study, we studied the effects of PKA phosphorylation on the binding of calmodulin to the fragment peptide of the proximal C-terminal tail of α1C subunit (CT1, a.a. 1509–1789 of guinea-pig). In the pull-down assay, in vitro PKA phosphorylation significantly decreased calmodulin binding to CT1 (61%) at high [Ca2+]. The phosphoresistant (CT1SA) and phosphomimetic (CT1SD) CT1 mutants, in which three PKA sites (Ser1574, 1626, 1699) were mutated to Ala and Asp, respectively, bound with calmodulin with 99% and 65% amount, respectively, compared to that of wild-type CT1. In contrast, at low [Ca2+], calmodulin-binding to CT1SD was higher (33–35%) than that to CT1SA. The distal C-terminal region of α1C (CT3, a.a. 1942–2169) is known to interact with CT1 and inhibit channel activity. CT3 bound to CT1SD was also significantly less than that to CT1SA. In inside-out patch, PKA catalytic subunit (PKAc) facilitated Ca2+ channel activity at both high and low Ca2+ condition. Altogether, these results support the hypothesis that PKA phosphorylation may enhance channel activity and attenuate the Ca2+-dependent inactivation, at least partially, by modulating calmodulin-CT1 interaction both directly and indirectly via CT3-CT1 interaction. Keywords: Ca2+ channels, Cav1.2, Calmodulin, PKA, Phosphorylation
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