title: Cd(2+) sensitivity and permeability of a low voltage-activated Ca(2+) channel with CatSper-like selectivity filter.
abstract: CatSper is a sperm-specific Ca(2+) channel that plays an essential role in the male fertility. However, its biophysical properties have been poorly characterized mainly due to its deficient heterologous expression. As other voltage-gated Ca(2+) channels (CaVs), CatSper possesses a conserved Ca(2+)-selective filter motif ([T/S]x[D/E]xW) in the pore region. Interestingly, CatSper conserves four aspartic acids (DDDD) as the negatively charged residues in this motif while high voltage-activated CaVs have four glutamic acids (EEEE) and low voltage-activated CaVs possess two glutamic acids and two aspartic acids (EEDD). Previous studies based on site-directed mutagenesis of L- and T-type channels showed that the number of D seems to have a negative correlation with their cadmium (Cd(2+)) sensitivity. These results suggest that CatSper (DDDD) would have low sensitivity to Cd(2+). To explore Cd(2+)-sensitivity and -permeability of CatSper, we performed two types of experiments: 1) Electrophysiological analysis of heterologously expressed human CaV3.1 channel and three pore mutants (DEDD, EDDD and DDDD), 2) Cd(2+) imaging of human spermatozoa with FluoZin-1. Electrophysiological studies showed a significant increase in Cd(2+) and manganese (Mn(2+)) currents through the CaV3.1 mutants as well as a reduction in the inhibitory effect of Cd(2+) on the Ca(2+) current. In fluorescence imaging with human sperm, we observed an increase in Cd(2+) influx potentiated by progesterone, a potent activator of CatSper. These results support our hypothesis, namely that Cd(2+)-sensitivity and -permeability are related to the absolute number of D in the Ca(2+)-selective filter independently to the type of the Cav channels.
authors.affiliation: Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto of Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos 62210, Mexico. Electronic address: egarza@ibt.unam.mx.
authors.forename: Edgar
authors.initials: E
authors.lastname: Garza-López
authors.affiliation: Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto of Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos 62210, Mexico. Electronic address: jcchavez@ibt.unam.mx.
authors.forename: Julio César
authors.initials: JC
authors.lastname: Chávez
authors.affiliation: Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto of Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos 62210, Mexico. Electronic address: csantana@ibt.unam.mx.
authors.forename: Carmen
authors.initials: C
authors.lastname: Santana-Calvo
authors.affiliation: Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto of Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos 62210, Mexico. Electronic address: iglogo@ibt.unam.mx.
authors.forename: Ignacio
authors.initials: I
authors.lastname: López-González
authors.affiliation: Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto of Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos 62210, Mexico. Electronic address: takuya@ibt.unam.mx.
authors.forename: Takuya
authors.initials: T
authors.lastname: Nishigaki
mesh_terms: D001224:Aspartic Acid / Q000378:metabolism; D002104:Cadmium / Q000494:pharmacology*; D015220:Calcium Channels / Q000378:metabolism*; D020747:Calcium Channels, T-Type / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects*; D057809:HEK293 Cells; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects; D008297:Male; D008345:Manganese / Q000378:metabolism; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D011374:Progesterone / Q000494:pharmacology; D013094:Spermatozoa / Q000187:drug effects / Q000378:metabolism
chemical_list: C121855:CACNA1G protein, human; C438823:CATSPER1 protein, human; D015220:Calcium Channels; D020747:Calcium Channels, T-Type; D050505:Mutant Proteins; D002104:Cadmium; D001224:Aspartic Acid; D008345:Manganese; D011374:Progesterone
keywords: CatSper selectivity filter; Fluorescence imaging; Heterologous expression; Patch clamp; T-type calcium channel