MOLECULAR DETERMINANTS OF BETA(1) SUBUNIT-INDUCED GATING MODULATION IN VOLTAGE-DEPENDENT NA+ CHANNELS

Recombinant brain, skeletal muscle, and heart voltage-gated Na+ channe l alpha subunits differ in their functional responses to an accessory beta(1) subunit when coexpressed in Xenopus oocytes. We exploited the distinct beta(1) subunit responses observed for the human heart (hH1) and human skeletal muscle (hSkM1) isoforms to identify determinants of this response, Chimeric a subunits were constructed by exchanging the S5-S6 interhelical loops of each domain between hH1 and hSkM1 and the n examined for effects on inactivation induced by coexpressed beta(1) subunit in oocytes, Substitution of single S5-S6 loops in either domai n 1 (D1/S5-S6) or domain 4 (D4/S5-S6) of hSkM1 by the corresponding se gments of hH1 produced channels that exhibited an attenuated response to coexpressed beta(1) subunit. Substitutions of both D1/S5-S6 and D4/ S5-S6 in hSkM1 by the corresponding loops from hH1 completely abolishe d the effects of the beta(1) subunit on inactivation. The reciprocal c himera in which both D1/S5-S6 and D4/S5-S6 from hSkM1 were transplante d into hH1 exhibited significant beta(1) responsiveness (accelerated i nactivation). The region within D4/S5-S6 that conferred beta(1) respon siveness was determined to reside primarily within an extracellular lo op between the putative pore-forming segment SS2 and D4/S6. Gating mod ulation was also demonstrated using a chimeric beta subunit consisting of the extracellular domains of beta(1) and the transmembrane and C-t erminal domains of the rat brain beta(2) subunit. These results sugges t that the D1/S5-S6 and D4/S5-S6 loops in the alpha subunit and the ex tracellular domain of the beta(1) subunit are important determinants o f the beta(1) subunit-induced gating modulation in Na+ channels.