Ca2+-sensitive inactivation of L-type Ca2+ channels depends on multiple cytoplasmic amino acid sequences of the alpha1C subunit.

Ca2+-dependent inactivation of Ca2+ currents is a physiological phenomenon widely associated with L-type Ca2+ channels. Although the pore-forming alpha1C subunit of the channel is the target for Ca2+ binding, the amino acid sequences involved in the binding and/or in the coordination of Ca2+-dependent inactivation are still unclear. Based on previous experiments, we have prepared truncation mutants of a human alpha1C subunit by systematically deleting an EF-hand motif and sequences in a segment of 80 amino acids in the carboxyl-terminal tail. We found that the rate as well as the Ca2+ dependence of inactivation of currents through these mutated channels were very different. We have identified three amino acid sequences, the presence of which is important for Ca2+-dependent inactivation: (i) a putative Ca2+-binding EF-hand motif, (ii) two hydrophilic residues (asparagine and glutamic acid) 77-78 amino acids downstream of the EF-hand motif, and (iii) a putative IQ calmodulin binding motif. We suggest that Ca2+-dependent inactivation is a cooperative process involving several amino acid sequences in cytoplasmic segments of the alpha1C subunit.