EC Accepts, published online ahead of print on 6 November 2009

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Eukaryotic Cell doi:10.1128/EC.00184-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

A chemical genetic screen for modulators of exocytic transport identifies inhibitors of a transport mechanism linked to GTR2 function

Lisha Zhang, Min Huang, and Edina Harsay^*

Department of Molecular Biosciences and High Throughput Screening Facility, University of Kansas, Lawrence KS 66045

^* To whom correspondence should be addressed. Email: harsay{at}ku.edu.

Membrane and protein traffic to the cell surface is mediated by partially redundant pathways that are difficult to perturb in ways that yield a strong phenotype. Such robustness is expected in a fine-tuned process, regulated by environmental cues, that is required for controlled cell surface growth and cell proliferation. Synthetic genetic interaction screens are especially valuable for investigating complex processes involving partially redundant pathways or mechanisms. In a previous study, we used a triple-synthetic-lethal yeast mutant screen to identify a novel component of the late exocytic transport machinery, Avl9. In a chemical-genetic version of the successful mutant screen, we have now identified small molecules that cause a rapid (within 15 min) accumulation of secretory cargo and Golgi-like membranes at low (<2 μM) concentration, indicating that the compounds likely target the exocytic transport machinery at the Golgi. We screened for genes that, when overexpressed, suppress the drug effects, and found that the Ras-like small GTPase, Gtr2, but not its homolog and binding partner, Gtr1, efficiently suppresses the toxic effects of the compounds. Furthermore, assays for suppression of the secretory defect caused by the compounds suggest that Gtr proteins can regulate a pathway that is perturbed by the compounds. Because avl9 and gtr mutants share some phenotypes, our results indicate that the small molecules identified by our chemical-genetic strategy are promising tools for understanding Avl9 function and the mechanisms that control late exocytic transport.