Proton channels in molluscs: A new bivalvian-specific minimal H_V 4 channel

Recently, three proton channels (H_V ) have been identified and characterized in Aplysia californica (AcH_V 1-3). Focusing on AcH_V 1 and AcH_V 2, analysis of Transcriptome Shotgun Assembly and genomic databases of 91 molluscs identified H_V homologous channels in other molluscs: channels homologous to AcH_V 1 and to AcH_V 2 were found in 90 species (56 full-length sequences) and in 33 species (18 full-length sequences), respectively. Here, we report the discovery of a fourth distinct proton channel family, H_V 4. This new family has high homology to AcH_V 1 and AcH_V 2 and was identified only in bivalvian molluscs (13 species, 12 full-length sequences). Typically, these channels possess an extracellular S1-S2 loop of intermediate size (~ 20 amino acids) compared to the shorter loops of molluscan H_V 1 channels (~ 13 amino acids) and the much larger loops of molluscan H_V 2 channels (> 65 amino acids). The characteristic voltage-sensor motif in S4 possesses only two arginine residues with the common third arginine being replaced by a lysine. Moreover, H_V 4 channels are much smaller with only around 200 amino acids in total length. The smallest functional channel found so far in nature (189 amino acids) is expressed in the pacific oyster Crassostrea gigas (CgH_V 4) and might be considered an archetypical minimal proton channel. Functional expression and electrophysiological characterization demonstrated that CgH_V 4 shares distinctive hallmarks of other investigated proton channels as high proton selectivity, slow activation, and pH- and voltage-regulated gating. This work is the first description of a H_V 4 type channel, adding a new member to the recently expanded family of proton channels.