This Article Full Text Full Text (PDF) Supplemental material Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Wagner, M. C. Articles by Goebl, M. G. Search for Related Content PubMed PubMed Citation Articles by Wagner, M. C. Articles by Goebl, M. G.

 Previous Article  |  Next Article 

Antimicrobial Agents and Chemotherapy, February 2006, p. 587-595, Vol. 50, No. 2
0066-4804/06/$08.00+0     doi:10.1128/AAC.50.2.587-595.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Loss of the Homotypic Fusion and Vacuole Protein Sorting or Golgi-Associated Retrograde Protein Vesicle Tethering Complexes Results in Gentamicin Sensitivity in the Yeast Saccharomyces cerevisiae

Department of Medicine, Division of Nephrology, and the Indiana Center for Biological Microscopy,¹ Department of Biochemistry and Molecular Biology and the Walther Cancer Institute, Indiana University School of Medicine, Indianapolis, Indiana 46202²

Received 4 August 2005/ Returned for modification 10 October 2005/ Accepted 23 November 2005

Gentamicin continues to be a primary antibiotic against gram-negative infections. Unfortunately, associated nephro- and ototoxicity limit its use. Our previous mammalian studies showed that gentamicin is trafficked to the endoplasmic reticulum in a retrograde manner and subsequently released into the cytosol. To better dissect the mechanism through which gentamicin induces toxicity, we have chosen to study its toxicity using the simple eukaryote Saccharomyces cerevisiae. A recent screen of the yeast deletion library identified multiple gentamicin-sensitive strains, many of which participate in intracellular trafficking. Our approach was to evaluate gentamicin sensitivity under logarithmic growth conditions. By quantifying growth inhibition in the presence of gentamicin, we determined that several of the sensitive strains were part of the Golgi-associated retrograde protein (GARP) and homotypic fusion and vacuole protein sorting (HOPS) complexes. Further evaluation of their other components showed that the deletion of any GARP member resulted in gentamicin-hypersensitive strains, while the deletion of other HOPS members resulted in less gentamicin sensitivity. Other genes whose deletion resulted in gentamicin hypersensitivity included ZUO1, SAC1, and NHX1. Finally, we utilized a Texas Red gentamicin conjugate to characterize gentamicin uptake and localization in both gentamicin-sensitive and -insensitive strains. These studies were consistent with our mammalian studies, suggesting that gentamicin toxicity in yeast results from alterations to intracellular trafficking pathways. The identification of genes whose absence results in gentamicin toxicity will help target specific pathways and mechanisms that contribute to gentamicin toxicity.

Supplemental material for this article may be found at http://aac.asm.org/.