Genetic screens in yeast
have identified over 50 vacuolar protein sorting (vps)genes,
involved in membrane trafficking from the late Golgi to the
vacuole. We show that VPS44is identical to NHX1,a gene
that encodes a sodium/ proton exchanger. The Saccharomyces cerevisiae
protein Nhx1p shows high homology to mammalian sodium/ proton
exchangers of the NHE family. Nhx1p is thought to transport sodium
ions into the prevacuole compartment in exchange for protons. We have
cloned the NHX1gene and introduced a C-terminal triple HA
tag. We have also deleted NHX1in a haploid yeast strain
(nhx1delta). Pulse-chase experiments show that approximately
35% of the newly synthesized soluble vacuolar protein carboxypeptidase
Y is missorted in the nhx1delta cells, and is secreted from the
cell. nhx1delta cells accumulate late Golgi, prevacuole and
lysosome markers in an aberrant structure next to the vacuole, and
late Golgi proteins are proteolytically cleaved more rapidly than in
wild type cells. Our results show that efficient transport out of the
prevacuolar compartment requires Nhx1p, and that nhx1delta
cells exhibit phenotypes characteristic of the 'class E' group of
vpsmutants. Our analysis of Nhx1p provides the first evidence
that a sodium/ proton exchange protein is important for correct
protein sorting, and that intraorganellar ion balance may be important
for endosomal function in yeast.
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