Finding new components of the target of rapamycin (TOR) signaling network through chemical genetics and proteome chips

doi:10.1073/pnas.0407117101

Finding new components of the target of rapamycin (TOR) signaling network through chemical genetics and proteome chips

^*Howard Hughes Medical Institute, Harvard Institute of Chemistry and Cell Biology, and Departments of Chemistry and Chemical Biology and ^**Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138; ^§Department of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, CT 06520; and ^‡‡Department of Molecular and Medical Pharmacology, University of California, Los Angeles, CA 90095

Contributed by Stuart L. Schreiber, October 11, 2004

Abstract

The TOR (target of rapamycin) proteins play important roles in nutrient signaling in eukaryotic cells. Rapamycin treatment induces a state reminiscent of the nutrient starvation response, often resulting in growth inhibition. Using a chemical genetic modifier screen, we identified two classes of small molecules, small-molecule inhibitors of rapamycin (SMIRs) and small-molecule enhancers of rapamycin (SMERs), that suppress and augment, respectively, rapamycin's effect in the yeast Saccharomyces cerevisiae. Probing proteome chips with biotinylated SMIRs revealed putative intracellular target proteins, including Tep1p, a homolog of the mammalian PTEN (phosphatase and tensin homologue deleted on chromosome 10) tumor suppressor, and Ybr077cp (Nir1p), a protein of previously unknown function that we show to be a component of the TOR signaling network. Both SMIR target proteins are associated with PI(3,4)P₂, suggesting a mechanism of regulation of the TOR pathway involving phosphatidylinositides. Our results illustrate the combined use of chemical genetics and proteomics in biological discovery and map a path for creating useful therapeutics for treating human diseases involving the TOR pathway, such as diabetes and cancer.

Footnotes

↵ ‡ To whom correspondence may be addressed. E-mail: stuart_schreiber{at}harvard.edu, jinghuang{at}mednet.ucla.edu, or michael.snyder{at}yale.edu.
↵ † Present address: Department of Molecular and Medical Pharmacology, University of California, Los Angeles, CA 90095.
↵ ¶ H.Z., S.J.H., and D.R.S. contributed equally to this work.
↵ ∥ Present address: Department of Pharmacology & HiT Center, The Johns Hopkins University School of Medicine, Baltimore, MD 21205.
↵ †† Present address: Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, United Kingdom.
Abbreviations: TOR, target of rapamycin; SMIR, small-molecule inhibitor of rapamycin; YPDA, yeast extract/peptone/dextrose/adenine; PI, phosphoinositide.
Freely available online through the PNAS open access option.