A type 2A phosphatase-sensitive phosphorylation site controls modal gating of L-type Ca2+ channels in human vascular smooth-muscle cells.

The patch-clamp technique was employed to investigate phosphorylation/dephosphorylation-dependent modulation of L-type Ca 2+ channels in smooth-muscle cells isolated from human umbilical vein. Okadaic acid, an inhibitor of phosphoprotein phosphatases type 1 (PP1) and 2A (PP2A), increased the probability of channels being in the open state ( P o ) in intact cells. This increase in P o was due mainly to promotion of long-lasting channel openings, i.e. promotion of ‘mode 2’ gating behaviour. Exposure of the cytoplasmic side of excised patches of membrane to the purified catalytic subunit of PP2A (PP2A c ) resulted in the opposite modulation of channel function. PP2A c (0.2 unit/ml) reduced the P o of Ca 2+ channels mainly via suppression of ‘mode 2’ gating. This effect of PP2A c was completely prevented by 1 µ M okadaic acid. The catalytic subunit of PP1 (0.2 unit/ml), however, barely affected channel activity. Our results provide evidence for a PP2A-sensitive regulatory site that controls modal gating of L-type Ca 2+ channels in smooth muscle.