Phosphorylation of S955 at the protein kinase A consensus promotes maturation of the alpha subunit of the colonic H+,K+ -ATPase

J Am Soc Nephrol. 2006 Jul;17(7):1833-40. doi: 10.1681/ASN.2006010032. Epub 2006 May 31.

Abstract

All the alpha subunits of the Na+,K+ -ATPases and H+,K+ -ATPases have a protein kinase A (PKA) consensus sequence near or in the ninth transmembrane domain. The role of this domain in influencing alpha subunit synthesis/degradation, plasma membrane localization, and 86Rb+ uptake has not been established for the alpha subunit of the colonic H+,K+ -ATPase. This study examined the effect of mutating S955 (within the PKA consensus site of the alpha subunit of the colonic H+,K+ -ATPase [HKalpha2]) to alanine (S955/A) or aspartic acid (S955/D) on alpha subunit expression and function. The results demonstrate that a negatively charged amino acid at position 955 of HKalpha2 promotes higher expression levels of both whole-cell and plasma membrane-localized HKalpha2. Moreover, inhibition of PKA reduced expression of wild-type HKalpha2 and associated 86Rb+ uptake. Last, the activity of the HKalpha2 S955/A was rescued by treatment with 4-phenylbutyric acid, a compound that was shown previously to restore function to the cystic fibrosis transmembrane conductance regulator.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Alanine / genetics
  • Biotinylation
  • Cell Line
  • Cell Membrane / enzymology
  • Colon / metabolism
  • Consensus Sequence
  • Cyclic AMP-Dependent Protein Kinases / genetics
  • Cyclic AMP-Dependent Protein Kinases / metabolism*
  • H(+)-K(+)-Exchanging ATPase / chemistry
  • H(+)-K(+)-Exchanging ATPase / genetics
  • H(+)-K(+)-Exchanging ATPase / metabolism*
  • Humans
  • Mutation
  • Phosphorylation
  • Protein Subunits / metabolism
  • Rubidium Radioisotopes / pharmacokinetics
  • Transfection

Substances

  • Protein Subunits
  • Rubidium Radioisotopes
  • Cyclic AMP-Dependent Protein Kinases
  • H(+)-K(+)-Exchanging ATPase
  • Alanine