Sequence analysis of the human genome permitted cloning of five Ca2+-channel β2 splice variants (β2a-β2e) that differed only in their proximal amino-termini. The functional consequences of such β2-subunit diversity were explored in recombinant L-type channels reconstituted in HEK 293 cells. β2a and β2e targeted autonomously to the plasma membrane, whereas β2b-β2d localized to the cytosol when expressed in HEK 293 cells. The pattern of modulation of L-type channel voltage-dependent inactivation gating correlated with the subcellular localization of the component β2 variant - membrane-bound β2a and β2e subunits conferred slow(er) channel inactivation kinetics and displayed a smaller fraction of channels recovering from inactivation with fast kinetics, compared to β2b β2d channels. The varying effects of β2 subunits on inactivation gating were accounted for by a quantitative model in which L-type channels reversibly distributed between fast and slow forms of voltage-dependent inactivation - membrane-bound β2 subunits substantially decreased the steady-state fraction of fast inactivating channels. Finally, the β2 variants also had distinctive effects on L-type channel steady-state activation gating, as revealed by differences in the waveforms of tail-activation (G-V) curves, and conferred differing degrees of prepulse facilitation to the channel. Our results predict important physiological consequences arising from subtle changes in Ca2+-channel β2-subunit structure due to alternative splicing and emphasize the utility of splice variants in probing structure-function mechanisms.
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