Organelle assembly in yeast: characterization of yeast mutants defective in vacuolar biogenesis and protein sorting.
Open Access
- 1 October 1988
- journal article
- research article
- Published by Rockefeller University Press in The Journal of cell biology
- Vol. 107 (4) , 1369-1383
- https://doi.org/10.1083/jcb.107.4.1369
Abstract
Yeast vacuole protein targeting (vpt) mutants exhibit defects in the sorting and processing of multiple vacuolar hydrolases. To evaluate the impact these vpt mutations have on the biogenesis and functioning of the lysosome-like vacuole, we have used light and electron microscopic techniques to analyze the vacuolar morphology in the mutants. These observations have permitted us to assign the vpt mutants to three distinct classes. The class A vpt mutants (26 complementation groups) contain 1-3 large vacuoles that are morphologically indistinguishable from those in the parental strain, suggesting that only a subset of the proteins destined for delivery to this compartment is mislocalized. One class A mutant (vpt13) is very sensitive to low pH and exhibits a defect in vacuole acidification. Consistent with a potential role for vacuolar pH in protein sorting, we found that bafilomycin A1, a specific inhibitor of the vacuolar ATPase, as well as the weak base ammonium acetate and the proton ionophore carbonyl cyanide m-chlorophenylhydrazone, collapse the pH gradient across the vacuolar membrane and cause the missorting and secretion of two vacuolar hydrolases in wild-type cells. Mutants in the three class B vpt complementation groups exhibit a fragmented vacuole morphology. In these mutants, no large normal vacuoles are observed. Instead, many (20-40) smaller vacuole-like organelles accumulate. The class C vpt mutants, which constitute four complementation groups, exhibit extreme defects in vacuole biogenesis. The mutants lack any organelle resembling a normal vacuole but accumulate other organelles including vesicles, multilamellar membrane structures, and Golgi-related structures. Heterozygous class C zygotes reassemble normal vacuoles rapidly, indicating that some of the accumulated aberrant structures may be intermediates in vacuole formation. These class C mutants also exhibit sensitivity to osmotic stress, suggesting an osmoregulatory role for the vacuole. The vpt mutants should provide insights into the normal physiological role of the vacuole, as well as allowing identification of components required for vacuole protein sorting and/or vacuole assembly.This publication has 53 references indexed in Scilit:
- Constitutive and Regulated Secretion of ProteinsAnnual Review of Cell Biology, 1987
- Pleiotropic plasma membrane ATPase mutations of Saccharomyces cerevisiae.Molecular and Cellular Biology, 1987
- Protein sorting in yeast: The localization determinant of yeast vacuolar carboxypeptidase Y resides in the propeptideCell, 1987
- Gene dosage-dependent secretion of yeast vacuolar carboxypeptidase Y.The Journal of cell biology, 1986
- How mitochondria import proteinsBiochimica et Biophysica Acta (BBA) - Reviews on Biomembranes, 1984
- Early stages in the yeast secretory pathway are required for transport of carboxypeptidase Y to the vacuoleCell, 1982
- Properties of H+-translocating adenosine triphosphatase in vacuolar membranes of SAccharomyces cerevisiae.Journal of Biological Chemistry, 1981
- Effect of weak bases on the intralysosomal pH in mouse peritoneal macrophages.The Journal of cell biology, 1981
- Fluorescence probe measurement of the intralysosomal pH in living cells and the perturbation of pH by various agents.Proceedings of the National Academy of Sciences, 1978
- The Isolation and Properties of the Yeast Cell VacuoleJournal of General Microbiology, 1968