title: A chemical genetic screen for mTOR pathway inhibitors based on 4E-BP-dependent nuclear accumulation of eIF4E.
abstract: The signal transduction pathway wherein mTOR regulates cellular growth and proliferation is an active target for drug discovery. The search for new mTOR inhibitors has recently yielded a handful of promising compounds that hold therapeutic potential. This search has been limited by the lack of a high-throughput assay to monitor the phosphorylation of a direct rapamycin-sensitive mTOR substrate in cells. Here we describe a novel cell-based chemical genetic screen useful for efficiently monitoring mTOR signaling to 4E-BPs in response to stimuli. The screen is based on the nuclear accumulation of eIF4E, which occurs in a 4E-BP-dependent manner specifically upon inhibition of mTOR signaling. Using this assay in a small-scale screen, we have identified several compounds not previously known to inhibit mTOR signaling, demonstrating that this method can be adapted to larger screens.
authors.affiliation: Department of Biochemistry and McGill Cancer Centre, McGill University, Montreal, QC H3A 1A3, Canada.
authors.forename: Mark
authors.initials: M
authors.lastname: Livingstone
authors.forename: Ola
authors.initials: O
authors.lastname: Larsson
authors.forename: Rami
authors.initials: R
authors.lastname: Sukarieh
authors.forename: Jerry
authors.initials: J
authors.lastname: Pelletier
authors.forename: Nahum
authors.initials: N
authors.lastname: Sonenberg
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002867:Chromones / Q000494:pharmacology; D055808:Drug Discovery / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D039561:Eukaryotic Initiation Factor-4E / Q000235:genetics / Q000378:metabolism*; D039642:Eukaryotic Initiation Factors / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D009025:Morpholines / Q000494:pharmacology; D010750:Phosphoproteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002867:Chromones; C499527:Eif4ebp1 protein, mouse; C491255:Eif4ebp2 protein, mouse; D004791:Enzyme Inhibitors; D039561:Eukaryotic Initiation Factor-4E; D039642:Eukaryotic Initiation Factors; D047908:Intracellular Signaling Peptides and Proteins; D009025:Morpholines; D010750:Phosphoproteins; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C546842:MTOR protein, human; C546843:mTOR protein, mouse; D017346:Protein Serine-Threonine Kinases; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus