Cd(2+) sensitivity and permeability of a low voltage-activated Ca(2+) channel with CatSper-like selectivity filter

Cell Calcium. 2016 Jul;60(1):41-50. doi: 10.1016/j.ceca.2016.03.011. Epub 2016 Mar 31.

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.

Keywords: CatSper selectivity filter; Fluorescence imaging; Heterologous expression; Patch clamp; T-type calcium channel.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aspartic Acid / metabolism
  • Cadmium / pharmacology*
  • Calcium Channels / metabolism*
  • Calcium Channels, T-Type / metabolism*
  • Cell Membrane / drug effects
  • Cell Membrane / metabolism
  • Cell Membrane Permeability / drug effects*
  • HEK293 Cells
  • Humans
  • Ion Channel Gating / drug effects
  • Male
  • Manganese / metabolism
  • Mutant Proteins / metabolism
  • Mutation / genetics
  • Progesterone / pharmacology
  • Spermatozoa / drug effects
  • Spermatozoa / metabolism

Substances

  • CACNA1G protein, human
  • CATSPER1 protein, human
  • Calcium Channels
  • Calcium Channels, T-Type
  • Mutant Proteins
  • Cadmium
  • Aspartic Acid
  • Manganese
  • Progesterone