The cystic fibrosis transmembrane regulator is present and functional in endosomes. Role as a determinant of endosomal pH

G. L. Lukacs, Xiu-Bao D Chang, N. Kartner, O. D. Rotstein, J. R. Riordan, S. Grinstein

Research output: Contribution to journalArticle

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Abstract

Cystic fibrosis is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR), which lead to defective Cl- conductance in epithelial cells. While the CFTR gene product has been detected in the plasma membrane, its presence and functional role in the membranes of intracellular compartments remain to be established. The purpose of the present experiments was to functionally localize CFTR in the endosomal membrane and to test the role of the associated Cl- conductance in the regulation of endosomal pH (pH(en)). When using conductive protonophores, the net H+ flux across the endosomal membrane of Chinese hamster ovary (CHO) cells is limited by the movement of counterions. Thus, ionic permeability could be estimated indirectly, from the changes in pH(en) determined fluorimetrically. Measurements in situ and in a cell-free microsomal preparation indicate the presence of a protein kinase A (PKA)-activated anion conductance in endosomes from CHO cells transfected with CFTR, but not in endosomes from wild-type or mock-transfected cells. In endosomes isolated from CFTR-expressing cells, the stimulatory effect of PKA was diminished by a specific peptide inhibitor of PKA, by alkaline phosphatase treatment or by a monoclonal antibody against the second nucleotide binding fold of CFTR. Increasing counterion permeability by phosphorylation of CFTR or by addition of valinomycin failed to alter the rate or extent of endosomal acidification in situ. Our observations indicate that functional CFTR, susceptible to activation by PKA, is present in endosomes of transfected CHO cells. More importantly, the data suggest that factors other than counterion permeability are the major determinants of pH(en).

Original languageEnglish (US)
Pages (from-to)14568-14572
Number of pages5
JournalJournal of Biological Chemistry
Volume267
Issue number21
StatePublished - 1992
Externally publishedYes

Fingerprint

Cystic Fibrosis Transmembrane Conductance Regulator
Endosomes
Cystic Fibrosis
Cyclic AMP-Dependent Protein Kinases
Cricetulus
Ovary
Permeability
Membranes
Valinomycin
Intracellular Membranes
Phosphorylation
Gene encoding
Acidification
Cell membranes
Regulator Genes
Peptides
Anions
Alkaline Phosphatase
Nucleotides
Genes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Lukacs, G. L., Chang, X-B. D., Kartner, N., Rotstein, O. D., Riordan, J. R., & Grinstein, S. (1992). The cystic fibrosis transmembrane regulator is present and functional in endosomes. Role as a determinant of endosomal pH. Journal of Biological Chemistry, 267(21), 14568-14572.

The cystic fibrosis transmembrane regulator is present and functional in endosomes. Role as a determinant of endosomal pH. / Lukacs, G. L.; Chang, Xiu-Bao D; Kartner, N.; Rotstein, O. D.; Riordan, J. R.; Grinstein, S.

In: Journal of Biological Chemistry, Vol. 267, No. 21, 1992, p. 14568-14572.

Research output: Contribution to journalArticle

Lukacs, GL, Chang, X-BD, Kartner, N, Rotstein, OD, Riordan, JR & Grinstein, S 1992, 'The cystic fibrosis transmembrane regulator is present and functional in endosomes. Role as a determinant of endosomal pH', Journal of Biological Chemistry, vol. 267, no. 21, pp. 14568-14572.
Lukacs, G. L. ; Chang, Xiu-Bao D ; Kartner, N. ; Rotstein, O. D. ; Riordan, J. R. ; Grinstein, S. / The cystic fibrosis transmembrane regulator is present and functional in endosomes. Role as a determinant of endosomal pH. In: Journal of Biological Chemistry. 1992 ; Vol. 267, No. 21. pp. 14568-14572.
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