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| Unbiased screen to identify novel AMPK substrates in mammalian cells
Our identification of the energy-sensing AMPK as a critical component of lifespan extension by DR in invertebrates raised the question of which substrates of AMPK were key for lifespan extension. While some of the substrates of AMPK in energy metabolism have been well characterized, the ensemble of AMPK substrates is not known. We have recently undertook a major effort to identify AMPK substrates in an unbiased manner, by conducting a chemical genetic screen for direct AMPK substrates in mammalian cells in collaboration with Kevan Shokat's lab at UCSF. We generated an analog-specific version of AMPK that can use a bulky analog of ATP and transfer the phosphate group to specific substrates in vivo. Coupled with tandem mass spectrometry, this approach allowed us to identify about 30 novel AMPK substrates. The results from our screen indicate that AMPK phosphorylates many more substrates in mammalian cells than initially anticipated, including components of the mitotic apparatus and cytoskeleton, and provide an important link between energy sensing and the completion of mitosis (Banko et al, Molecular Cell, 2011). This chemical genetic methods allows to identify direct targets of a protein kinase in cells, and could be used for other protein kinases. These findings have important implications for the mechanisms by which energy-sensing pathways regulates mitotic completion of cells that still proliferate in adult organisms, such as stem cells and cancer cells. Publications Banko MR , Allen JJ, Schaffer BE, Wilker EW, Tsou P, White KL, Villén J, Wang B, Kims SR, Sakamoto K, Gygi SP, Cantley LC, Yaffe MB, Shokat KM, and Brunet A (2011). Chemical genetic screen for AMPKa2 substrates unvocvers a netowrk of proteins involved in mitosis. Molecular Cell, 44: 878-892. Abstract PDF Supplementary information
Specific projects Regulation of the FOXO transcription factor network Importance of pro-longevity genes in aging neural stem cells Role of FOXO transcription factors in cognitive function Mechanisms underlying longevity by dietary restriction in C. elegans: role of the energy sensor AMPK Unbiased screen to identify novel AMPK substrates in mammalian cells Epigenetic regulation of aging in C. elegans
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