Functional expression of the cDNA encoded by the human ATP1AL1 gene

Academic Article

Abstract

  • The human ATP1AL1 gene encodes a protein expressed in brain, kidney, and skin and that is highly homologous to the recently cloned nongastric isoforms of H-K-adenosinetriphosphatase (H-K-ATPase). We have generated polyclonal antibodies against the protein encoded by ATP1AL1 and used them to monitor the protein's expression and distribution in transfection studies. The protein was retained in the endplasmic reticulum when it was transiently expressed alone in COS cells. In COS cells cotransfected with ATP1AL1 plus gastric H-K-ATPase β-subunit cDNAs (ATPlALl-gH-Kβ), both proteins reached the surface. Stably transfected lines of HEK 293 cells expressing both of these proteins demonstrate a 86Rb+ uptake activity sensitive to both 2-methyl, 8-(phenylmethoxy)imidazo(1, 2-a)pyridine 3-acetonitrile (SCH-28080) and ouabain (inhibitory constants of -131 and 42 μM, respectively). Outward proton fluxes were measured in the same cells as the spontaneous intracellular pH (pH;) recovery in cells loaded with a pH-sensitive dye [2′, 7′-bis(carboxyethy)-5(6)-carboxyfluorescein] and subjected to acid loading through an NH C1 pulse. The cells expressing both the ATP1AL1-encoded protein and the gastric H-K-ATPase β-subunit possess a net acid extrusion activity that can be inhibited by 1 mM ouabain. Comparison of the Rb influx and proton efflux, however, does not support equal H /Rb exchange mediated by this pump under the conditions of pH -monitoring experiments. Moreover, whereas the acid extrusion activity mediated by the pump shows a marked pH dependence, the 86Rb uptake activity present in the cells expressing the ATPlALl-gH-Kβ complex cannot be stimulated by acute lowering of pHi. These data suggest that the ATP1AL1-encoded protein is the catalytic a-subunit of a human K -dependent ATPase. The possible implications of the discrepancy between Rb uptake and pH monitoring data are discussed. Copyright © 1996 the American Physiological Society. 4 i i 86 + + + + + 86 +
  • Authors

    Published In

    Digital Object Identifier (doi)

    Pubmed Id

  • 15890718
  • Author List

  • Grishin AV; Bevensee MO; Modyanov NN; Rajendran V; Boron WF; Caplan MJ
  • Volume

  • 271
  • Issue

  • 3 PART 2