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Mouse Slc4a11 Mediates Extracellular‐pH‐Sensitive H + ‐Conduction in Xenopus oocytes
Author(s) -
Myers Evan J,
Marshall Aniko,
Parker Mark D
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.971.6
Subject(s) - xenopus , extracellular , hyperpolarization (physics) , reversal potential , chemistry , intracellular ph , membrane potential , voltage clamp , conductance , biophysics , patch clamp , microbiology and biotechnology , biochemistry , biology , gene , stereochemistry , receptor , mathematics , combinatorics , nuclear magnetic resonance spectroscopy
The SLC4 gene family includes ten members, eight of which encode bicarbonate transporters involved in intracellular pH (pH i ) regulation. SLC4A11 , a gene mutated in corneal dystrophy, has been proposed to encode a H + ‐permeable pH i regulator although its roster of substrates and their mode of transport remain controversial. In this study we set out to determine the molecular action of mouse Slc4a11 (mSlc4a11) expressed in isolation in Xenopus laevis oocytes. Using two‐electrode voltage‐clamp circuitry, we assessed the membrane conductance ( G m ) and membrane potential ( V m ) of oocytes as they were exposed to nominally HCO 3 − ‐free solutions of pH 6.5, 7.5, and 8.5. In pH 6.5 and 7.5 solutions, G m of mSlc4a11‐expressing oocytes was not significantly different from G m of H 2 O‐injected oocytes although V m of these cells was slightly more depolarized than V m of H 2 O‐injected cells at pH 6.5 and 7.5. For mSlc4a11‐expressing oocytes, but not for H 2 O‐injected oocytes, a switch from pH 7.5 to 8.5 solution elicited a significant increase in G m (P<0.01, n=6) and a hyperpolarization of V m to −73 ± 1 mV (P<0.01, n=6), a value consistent with the estimated reversal potential ( E rev ) for H + ( E H ). The conductance persisted with unaltered E rev even in the absence of extracellular Na + , K + , and Cl − (n=3). During prolonged (25 min) exposure to pH 8.5 solution, G m continued to rise without any further change in V m , but both G m and V m were immediately reset when pH 7.5 solution was restored (n=4). Notably, oocytes expressing the mutant mSlc4a11‐Arg99His, a mimic of the R125H mutation linked to human corneal dystrophy, behaved like H 2 O‐injected oocytes upon exposure to pH 8.5 solution (n=3) despite apparently normal accumulation of the mutant in the oocyte plasma membrane, as determined by biotinylation. Using a H + ‐selective microelectrode, we monitored pH i (with extracellular pH at 8.5) as we clamped oocyte V m to values more positive or more negative than predicted E H . The pH i of H 2 O‐injected oocytes was not substantially influenced by these maneuvers. On the other hand, the pH i of oocytes expressing mSlc4a11 increased when V m was > E H and decreased when V m was < E H (n=3 or each). We find that the response of V m to changes in the transmembrane H + ‐gradient is close to Nernstian (52 ± 12 mV/pH unit, n=3) in mSlc4a11‐expressing cells, demonstrating H + ‐selectivity. Together these data are consistent with the hypothesis that mSlc4a11 mediates an uncoupled, extracellular‐pH‐sensitive H + ‐conduction that is related to its physiological role. Support or Funding Information Supported by Start‐up Funds from UB:SUNY (to MDP).