β-adrenergic regulation of the L-type Ca2+ channel does not require phosphorylation of α1C Ser1700

Sympathetic nervous system triggered activation of protein kinase A, which phosphorylates several targets within cardiomyocytes, augments inotropy, chronotropy, and lusitropy. An important target of β-adrenergic stimulation is the sarcolemmal L-type Ca(2+) channel, CaV1.2, which plays a key role in cardiac excitation-contraction coupling. The molecular mechanisms of β-adrenergic regulation of CaV1.2 in cardiomyocytes, however, are incompletely known. Recently, it has been postulated that proteolytic cleavage at Ala(1800) and protein kinase A phosphorylation of Ser(1700) are required for β-adrenergic modulation of CaV1.2.

To assess the role of Ala(1800) in the cleavage of α1C and the role of Ser(1700) and Thr(1704) in mediating the adrenergic regulation of CaV1.2 in the heart.

Using a transgenic approach that enables selective and inducible expression in mice of FLAG-epitope-tagged, dihydropyridine-resistant CaV1.2 channels harboring mutations at key regulatory sites, we show that adrenergic regulation of CaV1.2 current and fractional shortening of cardiomyocytes do not require phosphorylation of either Ser(1700) or Thr(1704) of the α1C subunit. The presence of Ala(1800) and the (1798)NNAN(1801) motif in α1C is not required for proteolytic cleavage of the α1C C-terminus, and deletion of these residues did not perturb adrenergic modulation of CaV1.2 current.

These results show that protein kinase A phosphorylation of α1C Ser(1700) does not have a major role in the sympathetic stimulation of Ca(2+) current and contraction in the adult murine heart. Moreover, this new transgenic approach enables functional and reproducible screening of α1C mutants in freshly isolated adult cardiomyocytes in a reliable, timely, cost-effective manner.