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Mutational analysis of the intramembranous H10 loop of yeast Nhx1 reveals a critical role in ion homoeostasis and vesicle trafficking
- Source :
- Biochemical Journal. 398:97-105
- Publication Year :
- 2006
- Publisher :
- Portland Press Ltd., 2006.
-
Abstract
- Yeast Nhx1 [Na+(K+)/H+ exchanger 1] is an intracellular Na+(K+)/H+ exchanger, localizing to the late endosome where it is important for ion homoeostasis and vesicle trafficking. Phylogenetic analysis of NHE (Na+/H+ exchanger) sequences has ident-ified orthologous proteins, including HsNHE6 (human NHE6), HsNHE7 and HsNHE9 of unknown physiological role. These appear distinct from well-studied mammalian plasma membrane isoforms (NHE1–NHE5). To explore the differences between plasma membrane and intracellular NHEs and understand the link between ion homoeostasis and vesicle trafficking, we examined the consequence of replacing residues in the intramembranous H10 loop of Nhx1 between transmembrane segments 9 and 10. The critical role for the carboxy group of Glu355 in ion transport is consistent with the invariance of this residue in all NHEs. Surprisingly, residues specifically conserved in the intracellular isoforms (such as Phe357 and Tyr361) could not be replaced with closely similar residues (leucine and phenylalanine) found in the plasma membrane isoforms without loss of function, revealing unexpected side chain specificity. The trafficking phenotypes of all Nhx1 mutants, including hygromycin-sensitivity and missorting of carboxypeptidase Y, were found to directly correlate with pH homoeostasis defects and could be proportionately corrected by titration with weak base. The present study demonstrates the importance of the H10 loop of the NHE family, highlights the differences between plasma membrane and intracellular isoforms and shows that trafficking defects are tightly coupled with pH homoeostasis.
- Subjects :
- Gene isoform
Saccharomyces cerevisiae Proteins
Sodium-Hydrogen Exchangers
Amino Acids, Acidic
DNA Mutational Analysis
Molecular Sequence Data
Mutant
Cathepsin A
Microbial Sensitivity Tests
Saccharomyces cerevisiae
Biology
Biochemistry
Homeostasis
Amino Acid Sequence
Transport Vesicles
Cation Transport Proteins
Molecular Biology
Late endosome
Ion transporter
Ion Transport
Dose-Response Relationship, Drug
Vesicle
Cell Membrane
Cell Biology
Hydrogen-Ion Concentration
Transmembrane protein
Cell biology
Protein Transport
Phenotype
Amino Acid Substitution
Mutation
Vacuoles
Hygromycin B
Intracellular
Research Article
Subjects
Details
- ISSN :
- 14708728 and 02646021
- Volume :
- 398
- Database :
- OpenAIRE
- Journal :
- Biochemical Journal
- Accession number :
- edsair.doi.dedup.....db5583ce08169f7da3d965f2a7a2b1cf