Cloning and characterization of voltage-gated sodium and calcium channel homologs from the single-cell choanoflagellate, Salpingoeca rosetta

Abstract

We have isolated a complement of gene homologs from the simplest extant eukaryotic species to possess voltage-gated sodium (Na+) and calcium (Ca2+) channels, Salpingoeca rosetta. The isolated channels share the same 4 Domain, 6 Transmembrane helices (4x6TM) architecture characteristic of voltage-gated ion channels and do not exhibit any alternative splicing. Transfection and expression of both channels in Human Embryonic Kidney-293T (HEK-293T) cells generate voltage-dependent ionic currents. SroNav2 codes for an 1831 amino-acid transmembrane protein of four repeat domains with a selectivity filter ring, DEEA, resembling Ca2+-selective sodium channel genes found exclusively in non-vertebrate animals. The structurally similar to Nav1, SroNav2 shows non-selective ion permeability and allows the passage of both divalent and monovalent ions. The SroNav2 selectivity filter mutant, DEKA, produced to mimic neuronal vertebrate Nav1 channels, produces a highly selective channel that exclusively passes monovalent ions. SroCav1 codes for a 1664 amino-acid transmembrane protein, requires co-expression of a native β auxiliary subunit, and is highly selective and does not allow the passage of sodium. SroCav1 lacks obvious calcium-dependent inactivation and does not exhibit the same long-lasting currents characteristic of L-type channels with Ba2+ as a charge carrier. We envisage that these homologs of voltage-gated Ca2+ and Na+ channels found in single cell choanoflagellate Salpingoeca rosetta may generate Ca2+-dependent action potentials that signal between cells of choanoflagellate colonies, regulate intra-cellular events, or control movement of it’s single flagellum or cilia.