Bestrophin-1 modulates currents through voltage-dependent L-type Ca(2+) channels by physically interacting with the ?-subunits of Ca(2+) channels. The main function of ?-subunits is to regulate the number of pore-forming Ca(V)-subunits in the cell membrane and modulate Ca(2+) channel currents. To understand the influence of full-length bestrophin-1 on ?-subunit function, we studied binding and localization of bestrophin-1 and Ca(2+) channel subunits, together with modulation of Ca(V)1.3 Ca(2+) channels currents. In heterologeous expression, bestrophin-1 showed co-immunoprecipitation with either, ?3-, or ?4-subunits. We identified a new highly conserved cluster of proline-rich motifs on the bestrophin-1 C-terminus between amino acid position 468 and 486, which enables possible binding to SH3-domains of ?-subunits. A bestrophin-1 that lacks these proline-rich motifs (?CT-PxxP bestrophin-1) showed reduced efficiency to co-immunoprecipitate with ?3 and ?4-subunits. In the presence of ?CT-PxxP bestrophin-1, ?4-subunits and Ca(V)1.3 subunits partly lost membrane localization. Currents from Ca(V)1.3 subunits were modified in the presence of ?4-subunit and wild-type bestrophin-1: accelerated time-dependent activation and reduced current density. With ?CTPxxP bestrophin-1, currents showed the same time-dependent activation as with wild-type bestrophin-1, but the current density was further reduced due to decreased number of Ca(2+) channels proteins in the cell membrane. In summary, we described new proline-rich motifs on bestrophin-1 C-terminus, which help to maintain the ability of ?-subunits to regulate surface expression of pore-forming Ca(V) Ca(2+)-channel subunits.
