MOLECULAR DETERMINANTS OF L-TYPE CA2- SEGMENT EXCHANGE ANALYSIS OF THE CARBOXYL-TERMINAL CYTOPLASMIC MOTIF ENCODED BY EXON-40-42 OF THE HUMAN ALPHA(1C) SUBUNIT GENE( CHANNEL INACTIVATION )

Recently we have described a splice variant of the L-type Ca2+ channel (alpha(1C,86)) in which 80 amino acids (1572-1651) of the conventiona l alpha(1C,77) were substituted by another 81 amino acids due to alter native splicing of exons 40-42, Ba2+ current (I-Ba) through alpha(1C,8 6) exhibited faster inactivation kinetics, was strongly voltage-depend ent, and had no Ca2+-dependent inactivation. An oligonucleotide-direct ed segment substitution and expression of the mutated channels in Xeno pus oocytes were used to study the molecular determinants for gating o f the channel within the 80-amino acid domain. Replacement of segments 1572-1598 or 1595-1652 of the ''slow'' alpha(1C,77) channel with the respective segments of the ''fast'' alpha(1C,86) gave rise to rapidly inactivating alpha(1C,86)-like channel isoforms, We found that replace ment of either motifs (1572)IKTEG(1576) or (1600)LLDQV(1604) of alpha( 1C,77) with the respective sequences of alpha(1C,86) caused strong but partial acceleration of I-Ba inactivation. Replacement of both sequen ces produced an alpha(1C,86)-like fast channel which had no Ca2+-depen dent inactivation. These results support the hypothesis that motifs 15 72-1576 and 1600-1604 of alpha(1C,77) contribute cooperatively to inac tivation kinetics of alpha(1C) and are critical for Ca2+-dependent ina ctivation of the channel.