Makoto F Kuwabara, Joschua Klemptner, Julia Muth, Emilia De Martino, Dominik Oliver, Thomas K Berger
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引用次数: 0
Abstract
Voltage-gated ion channels allow ion flux across biological membranes in response to changes in the membrane potential. HCNL1 is a recently discovered voltage-gated ion channel that selectively conducts protons through its voltage-sensing domain (VSD), reminiscent of the well-studied depolarization-activated Hv1 proton channel. However, HCNL1 is activated by hyperpolarization, allowing the influx of protons, which leads to an intracellular acidification in zebrafish sperm. Zinc ions (Zn2+) are important cofactors in many proteins and essential for sperm physiology. Proton channels such as Hv1 and Otopetrin1 are inhibited by Zn2+. We investigated the effect of Zn2+ on heterologously expressed HCNL1 channels using electrophysiological and fluorometric techniques. Extracellular Zn2+ inhibits HCNL1 currents with an apparent half-maximal inhibition (IC50) of 26 μM. Zn2+ slows voltage-dependent current kinetics, shifts the voltage-dependent activation to more negative potentials, and alters hyperpolarization-induced conformational changes of the voltage sensor. Our data suggest that extracellular Zn2+ inhibits HCNL1 currents by multiple mechanisms, including modulation of channel gating. Two histidine residues located at the extracellular side of the VSD might weakly contribute to Zn2+ coordination: mutants with either histidine replaced with alanine show modest shifts of the IC50 values to higher concentrations. Interestingly, Zn2+ inhibits HCNL1 at even lower concentrations from the intracellular side (IC50 ≈ 0.5 μM). A histidine residue at the intracellular end of S1 (position 50) is important for Zn2+ binding: much higher Zn2+ concentrations are required to inhibit the mutant HCNL1-H50A (IC50 ≈ 106 μM). We anticipate that Zn2+ will be a useful ion to study the structure-function relationship of HCNL1 as well as the physiological role of HCNL1 in zebrafish sperm.
期刊介绍:
BJ publishes original articles, letters, and perspectives on important problems in modern biophysics. The papers should be written so as to be of interest to a broad community of biophysicists. BJ welcomes experimental studies that employ quantitative physical approaches for the study of biological systems, including or spanning scales from molecule to whole organism. Experimental studies of a purely descriptive or phenomenological nature, with no theoretical or mechanistic underpinning, are not appropriate for publication in BJ. Theoretical studies should offer new insights into the understanding ofexperimental results or suggest new experimentally testable hypotheses. Articles reporting significant methodological or technological advances, which have potential to open new areas of biophysical investigation, are also suitable for publication in BJ. Papers describing improvements in accuracy or speed of existing methods or extra detail within methods described previously are not suitable for BJ.