Grant: Discovery Grants Program - Individual

Most cellular responses to signals have a common component of regulation arising from the coordinated recruitment of protein complexes to membranes. The coordination of this process is essential to almost every aspect of a cell's life and death. Studying how proteins interact with lipid membranes is extremely challenging by standard biophysical approaches, and has been a major hurdle in understanding signalling at the molecular level. In the last six years of NSERC support (2014-2019), I have made tremendous progress in defining how lipid signalling systems are regulated. The program I am proposing will continue and expand on this strong track record of success to examine the regulation of lipid signalling proteins. The long-term vision of this research program is to understand the molecular details of how large macromolecular assemblies are recruited, assembled, and regulated on lipid membranes. The specific objectives of the research program are as follows: Objectives  1. Define the molecular mechanism of regulation of guanine nucleotide exchange factors (GEFs) on membranes 2. Define how the Phospholipase C (PLC) family of enzymes is regulated on membranes Approach The biological systems under study in this program will be all be approached using a unique synergy of state of the art structural biology approaches, including HDX-MS, Electron Microscopy (EM), and XRC. This approach is unique within Canada, and provides trainees with an exceptional training opportunity. Many of the complexes under study are extremely large (>500 kDa), and are composed of multiple different proteins. The tandem application of these approaches is essential to the study of these complicated systems, and importantly these tools will be applied in a synergistic way, so that the different approaches can inform novel strategies to optimise the likelihood of success in obtaining high resolution structural information. Specifically, this approach allows for the study of high resolution structural information on how these large protein complexes are assembled, together with dynamic studies using HDX-MS to define conformational changes that occur on membrane surfaces. This approach will allow for critical insight into the molecular mechanisms regulating lipid signalling. Impact and significance This research program describes a long term vision of our goals to understand lipid signalling at the molecular level, as well as a set of clear and achievable medium and short term objectives to study lipid signalling systems of extreme complexity. This research will play a vital role in understanding the mechanisms by which lipid signalling complexes are regulated on membranes, which touches on almost all aspects of cell biology. The immediate impact of this proposal on Canada is the training of highly qualified personnel in advanced techniques including molecular biology, HDX-MS, EM, and XRC, which are all in high demand in the biotechnology sector.