We have long known that our cells undergo constant adaptation to changes in their environment, and new technology allows unprecedented tracking of these dynamic cellular events, as reported by Dr. Anne-Claude Gingras and her colleagues. Their paper was published online in Nature Methods on October 27, 2013.
Dr. Gingras' research program has focused on monitoring changes in protein-to-protein interactions that characterize disease processes. “In tracking these changes, we are striving for an approach that provides a holistic view of critical cellular events,” she says.
The team built on previous innovative road-maps of protein interactions developed in the Gringras lab, other labs at the Lunenfeld-Tanenbaum Research Institute, and internationally. Such maps, termed interactomes, encompass thousands of touch-points among proteins that allow scientists to assemble clear pictures of how proteins are organized as well as to identify how disease-related malfunctions affect these protein networks.
The Gingras paper introduces a research platform that couples affinity purification (AP) with a technology termed SWATH quantification. SWATH analysis (Sequential Window Acquisition of all THeoretical Spectra) can scan the target mass range in a timeframe compatible to that of liquid chromatography in a quantitative fashion, but without needing to predefine targets of interest. Through the use of high-level computational analysis, data can be rapidly extracted and statistical analysis performed to score modulated interactions.
“One critical benefit of SWATH is that it does not require specific assays to be developed for individual peptides,” says Dr. Jean-Philippe Lambert, the lead author and a post-doctoral fellow in the Gingras laboratory. “That’s important because it means that this system is efficient and cost-effective. As such, it is scalable for large-scale biological experiments, such as the ones taking place in the Lunenfeld-Tanenbaum Research Institute.”
For its proof of principle, the team examined protein-protein interactions that were triggered by melanoma-associated mutations in the kinase CDK4 as well as their response to NVP-AUY922, an HSP90 inhibitor currently in clinical trials.
“Our ability to understand the molecular causes of diseases is largely limited by the precision of our tools,” notes Jim Woodgett, Director of the Lunenfeld-Tanenbaum, “hence, this important advance will improve knowledge of many diseases and accelerate new ideas for therapies.”
The paper is titled “Mapping differential interactomes by affinity purification coupled with data independent mass spectrometry acquisition,” and was co-led by Lambert and Gingras at the Lunenfeld-Tanenbaum and their industrial partners, Gordana Ivosev and Stephen Tate at AB Sciex. Gingras is also an associate professor in the Molecular Genetics department at the University of Toronto. The twelve coauthors also represent the Whitehead Institute for Biomedical Research and the Massachusetts Institute of Technology; the Dana-Farber Cancer Institute at Harvard Medical School, and the ETH Zurich and University of Zurich.
Seed funding to the Lunenfeld-Tanenbaum team was provided by Venture Sinai. Other funds to the Lunenfeld-Tanenbaum leveraged from the Canadian Institute of Health Research, the National Institutes of Health, the Ontario Research Fund, and the Canada Research Chair program.