Our research programme aims at establishing mechanisms of regulation of an ancient family of giant kinases involved in cellular homeostasis. The phosphoinositide 3-kinase like family of enzymes includes unique lipid and protein kinases. Projects include understanding mechanisms of regulation of the lipid kinase VPS34 and the protein kinases mTOR, ATM and ATR. VPS34 and mTOR are present in large complexes that are activated at membrane interfaces, and they are essential responding to nutritional stress. ATM and ATR are present in large complexes that enable them to assume a fundamental role in sensing and responding to DNA damage. They are activated in response to DNA damage and are master kinases linking DNA damage to cell cycle progression and cell death. The enzymes are part of complexes that sense DNA breaks and phosphorylate a wide range of targets in the DNA damage response. Cancer cells rely on this pathway to survive genomic instability. We are characterising structural mechanisms regulation of these enzyme com-plexes as well as developing small molecule inhibitors and activators of them.
The project involves single-particle electron cryo-microscopy (cryo-EM) and cryo-electron tomography (cryo-ET) of VPS34- and mTOR-containing complexes on membranes and characterisation of the dynamics of the complexes by hydrogen/deuterium exchange mass spectrometry (HDX-MS) and enzyme kinetics. Work in the group has developed HDX-MS analysis of protein/protein and protein/membrane interactions to an unprecedented level. This powerful technique has been applied to precisely map interactions within even very large complexes, such as the 80S ribosome. An important dimension of the project is that you will become adept at cryo-EM, HDX-MS and X-ray crystallography. The structural work will be carried out in parallel with mammalian cell biology and in vitro enzymology.
Inglis AJ, Masson GR, Shao S, Perisic O, McLaughlin SH, Hegde RS, Williams RL. (2019) Activation of GCN2 by the ribosomal P-stalk. Proc Natl Acad Sci U S A. 116(11):4946-4954. PMID: 30804176.
Baretić D, Pollard HK, Fisher DI, Johnson CM, Santhanam B, Truman CM, Kouba T, Fersht AR, Phillips C, Williams RL (2017) Structures of closed and open conformations of dimeric human ATM. Sci Adv. 10;3(5):e1700933. PMID 28508083.
Rostislavleva, K., Soler, N., Ohashi, Y., Zhang, L., Pardon, E., Burke, J.E., Masson, G.R., Johnson, C., Steyaert, J., Ktistakis, N.T., et al. (2015). Structure and flexibility of the endosomal Vps34 complex reveals the basis of its function on membranes. Science 350, aac7365–aac7365.
Yang, H., Jiang, X., Li, B., Yang, H.J., Miller, M., Yang, A., Dhar, A., and Pavletich, N.P. (2017). Mechanisms of mTORC1 activation by RHEB and inhibition by PRAS40. Nature 552, 368–373.