Reconstitution of ß-adrenergic regulation of CaV1.2: Rad-dependent and Rad-independent protein kinase A mechanisms.

ABSTRACT

L-type voltage-gated CaV1.2 channels crucially regulate cardiac muscle contraction. Activation of ß-adrenergic receptors (ß-AR) augments contraction via protein kinase A (PKA)-induced increase of calcium influx through CaV1.2 channels. To date, the full ß-AR cascade has never been heterologously reconstituted. A recent study identified Rad, a CaV1.2 inhibitory protein, as essential for PKA regulation of CaV1.2. We corroborated this finding and reconstituted the complete pathway with agonist activation of ß1-AR or ß2-AR in Xenopus oocytes. We found, and distinguished between, two distinct pathways of PKA modulation of CaV1.2 Rad dependent (∼80% of total) and Rad independent. The reconstituted system reproduces the known features of ß-AR regulation in cardiomyocytes and reveals several aspects the differential regulation of posttranslationally modified CaV1.2 variants and the distinct features of ß1-AR versus ß2-AR activity. This system allows for the addressing of central unresolved issues in the ß-AR-CaV1.2 cascade and will facilitate the development of therapies for catecholamine-induced cardiac pathologies.