Ca v 2.3 Ca 2+ channel interacts with the G1-subunit of V-ATPase

Abstract

Background: Calcium channels are essential in coupling action potential to signal transduction in cells. There are several types of calcium channels, which can be pharmacologically classified as L-, N-, P/Q-, R- and T-type. But molecular basis of R-type channels is less clearly understood compared the other channel types. Therefore the current study aims at understanding the molecular functions of R-type calcium channels by identifying interaction partners of the channel. Methods: In order to do so, a yeast two hybrid (Y2H) screen, with carboxy terminus of α1 subunit of the channel, as the bait, was performed. G1 subunit of v-ATPase was identified as a putative interaction partner of human Ca v 2.3 by using the Y2H screening. The interaction was confirmed by immunoprecipitation. To study the functional importance of the interaction, bafilomycin A 1 , a potent and specific inhibitor of v-ATPase was used in patch-clamp recordings in Ca v 2.3 stably-transfected HEK-293 cells (2C6) as well as in electroretinography of the isolated bovine retina expressing R-type Ca 2+ channels. Results: G1 subunit of v-ATPase interacts with C-terminal tail of Ca v 2.3 and bafilomycin A 1 reduces Ca v 2.3 mediated calcium currents. Additionally peak I Ca is inhibited in retinal signal transduction when recorded as ERG b-wave. Conclusions: The results suggest that v-ATPase interacts physically and also functionally with Ca v 2.3. This is the first demonstration of association of Ca v 2.3 C-terminus with a protein complex which is involved in transmembrane signalling. © 2011 S. Karger AG, Basel.