Chloride channel function in the yeast TRK-potassium transporters

J Membr Biol. 2004 Apr 1;198(3):177-92. doi: 10.1007/s00232-004-0671-1.

Abstract

The TRK proteins-Trk1p and Trk2p- are the main agents responsible for "active" accumulation of potassium by the yeast Saccharomyces cerevisiae. In previous studies, inward currents measured through those proteins by whole-cell patch-clamping proved very unresponsive to changes of extracellular potassium concentration, although they did increase with extracellular proton concentration-qualitatively as expected for H(+) coupling to K(+) uptake. These puzzling observations have now been explored in greater detail, with the following major findings: a) the large inward TRK currents are not carried by influx of either K(+) or H(+), but rather by an efflux of chloride ions; b) with normal expression levels for Trk1p and Trk2p in potassium-replete cells, the inward TRK currents are contributed approximately half by Trk1p and half by Trk2p; but c) strain background strongly influences the absolute magnitude of these currents, which are nearly twice as large in W303-derived spheroplasts as in S288c-derived cells (same cell-size and identical recording conditions); d) incorporation of mutations that increase cell size (deletion of the Golgi calcium pump, Pmr1p) or that upregulate the TRK2 promoter, can further substantially increase the TRK currents; e) removal of intracellular chloride (e.g., replacement by sulfate or gluconate) reveals small inward currents that are K(+)-dependent and can be enhanced by K(+) starvation; and f) finally, the latter currents display two saturating kinetic components, with preliminary estimates of K(0.5) at 46 micro M [K(+)](out) and 6.8 m M [K(+)](out), and saturating fluxes of approximately 5 m M/min and approximately 10 m M/min (referred to intracellular water). These numbers are compatible with the normal K(+)-transport properties of Trk1p and Trk2p, respectively.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Biological Transport, Active / physiology
  • Cation Transport Proteins / physiology*
  • Chloride Channels / physiology*
  • Chlorine / metabolism*
  • Ion Channel Gating / physiology
  • Membrane Potentials / physiology*
  • Mutagenesis, Site-Directed
  • Potassium / metabolism*
  • Recombinant Proteins / metabolism
  • Saccharomyces cerevisiae / classification
  • Saccharomyces cerevisiae / physiology*
  • Saccharomyces cerevisiae Proteins / physiology*
  • Species Specificity
  • Structure-Activity Relationship

Substances

  • Cation Transport Proteins
  • Chloride Channels
  • Recombinant Proteins
  • Saccharomyces cerevisiae Proteins
  • TRK2 protein, S cerevisiae
  • TRK1 protein, S cerevisiae
  • Chlorine
  • Potassium