Stable incorporation versus dynamic exchange of ? subunits in a native Ca2+ channel complex.

ABSTRACT: Voltage-gated Ca(2+) channels are multi-subunit membrane proteins that transduce depolarization into cellular functions such as excitation-contraction coupling in muscle or neurotransmitter release in neurons. The auxiliary ? subunits function in membrane targeting of the channel and modulation of its gating properties. However, whether ? subunits can reversibly interact with, and thus differentially modulate, channels in the membrane is still unresolved. In the present study we applied fluorescence recovery after photobleaching (FRAP) of GFP-tagged ?1 and ? subunits expressed in dysgenic myotubes to study the relative dynamics of these Ca(2+) channel subunits for the first time in a native functional signaling complex. Identical fluorescence recovery rates of both subunits indicate stable interactions, distinct recovery rates indicate dynamic interactions. Whereas the skeletal muscle ?1a isoform formed stable complexes with CaV1.1 and CaV1.2, the non-skeletal muscle ?2a and ?4b isoforms dynamically interacted with both ?1 subunits. Neither replacing the I-II loop of CaV1.1 with that of CaV2.1, nor deletions in the proximal I-II loop, known to change the orientation of ? relative to the ?1 subunit, altered the specific dynamic properties of the ? subunits. In contrast, a single residue substitution in the ? interaction pocket of ?1aM293A increased the FRAP rate threefold. Taken together, these findings indicate that in skeletal muscle triads the homologous ?1a subunit forms a stable complex, whereas the heterologous ?2a and ?4b subunits form dynamic complexes with the Ca(2+) channel. The distinct binding properties are not determined by differences in the I-II loop sequences of the ?1 subunits, but are intrinsic properties of the ? subunit isoforms.

SUBMITTER: Campiglio M 

PROVIDER: S-EPMC4148589 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature