Both Voltage- and Ca2+-Dependent Inactivation of Cav1.2 Ca2+ Channels are Suppresed in Myotubes, Likely Because of Triad Junction Proteins other than RyR1

Abstract

To investigate the relationship between calcium channel and EC coupling functions of CaV1.1 in skeletal muscle, we have compared its behavior with that of the cardiac L-type channel, CaV1.2. Using expression in tsA-201 cells, we showed that substituting CaV1.1 IQ motif residues into CaV1.2 IQ caused a loss of calmodulin binding and calcium-dependent inactivation (JBC 283:29301-11, 2008), raising the possibility that inactivation is maladaptive for CaV1.1 function in muscle. We thus compared inactivation of CaV1.2 in tsA-201 cells and dysgenic myotubes (null for CaV1.1). For CaV1.2 in tsA-201 cells (co-expressed with α2δ1 and β2a), I50 ms/Ipeak (R50) at +20 mV was .69 in 10 Ca and .87 in 10 Ba, whereas R50 for CaV1.2 in dysgenic myotubes was .95 at +20 mV in 10 Ca, indicating that both voltage- and calcium-dependent inactivation were suppressed. In tsA-201 cells, co-expression of β1a (the predominant skeletal muscle isoform) did not significantly alter R50 values from those with β2a. Furthermore, R50 values were similar for CaV1.2 expressed in dysgenic myotubes and myotubes null for both CaV1.1 and RyR1, suggesting that some component of muscle triad junctions other than RyR1 is responsible for the suppression of inactivation. We thus expressed CaV2.1 in dysgenic myotubes because this neuronal channel, unlike CaV1.2, is not targeted to triad junctions (PNAS 95:1903-1908, 1998). Consistent with the hypothesis CaV2.1 channels showed significant voltage- and calcium-dependent inactivation in dysgenic myotubes which were similar to those of the channel expressed in tsA-201 cells. We are currently seeking to identify the components involved in suppressing inactivation of CaV1.2 in myotubes. Supported by AHA0190016G to JDO and NIH/NIAMS (AR055104) and MDA4319 to KGB.