Systematic platform for the edgetic study of protein biophysical interactions
For almost half a century, geneticists have used gene deletions to understand the function of genes and how they pertain to human disease. However, deletions do not always recapitulate the phenotypic effects, particularly diseases that arise from to missense mutations in protein-coding genes. A recent study has shown that two-thirds of missense mutations in disease genes give rise to alleles that perturb protein-protein interactions, with half of them corresponding to ‘‘edgetic’’ alleles that affect only a subset of interactions while leaving most other interactions unperturbed (Sahni et al., 2015). With comprehensive reference interactome maps nearing completion, a systematic understanding of how different mutations affect interactome networks and how changes in interactome networks lead to phenotypic alteration becomes not only possible but is also urgently needed. Unfortunately, there is still no efficient way to systematically perturb or “knock-out” each individual interaction in the interactome and determine their biological consequences.
We are building a high-throughput platform that uses forward and reverse genetics approaches to systematically identify interaction-defective alleles as well as alleles that are defective for a particular function. Furthermore, our platform aims to find compensatory mutations in the interacting partner that restores a given interaction and consequently, the biological function in which the interacting proteins are involved.
We are applying this platform to the yeast Rpd3 histone deacetylase complex, which serves as a tractable model system for understanding the biophysical or molecular features that make a mutation edgetic or compensable, how a pair of mutations can compensate each other on 3D structures, the relationship that biophysical compensatory mutations have with their corresponding phenotypic outcome and conversely, how phenotypic mutations influence the underlying biophysical interaction. This platform can ultimately be adapted for interactome-wide edge perturbation screens in higher eukaryotes by using emerging CRISPR technologies to introduce compensatory mutations of known edgetic human disease mutations in order to develop a clearer understanding of the function of interactions in our human interactome.
Please contact Michael Calderwood with your questions and comments.
We are building a high-throughput platform that uses forward and reverse genetics approaches to systematically identify interaction-defective alleles as well as alleles that are defective for a particular function. Furthermore, our platform aims to find compensatory mutations in the interacting partner that restores a given interaction and consequently, the biological function in which the interacting proteins are involved.
We are applying this platform to the yeast Rpd3 histone deacetylase complex, which serves as a tractable model system for understanding the biophysical or molecular features that make a mutation edgetic or compensable, how a pair of mutations can compensate each other on 3D structures, the relationship that biophysical compensatory mutations have with their corresponding phenotypic outcome and conversely, how phenotypic mutations influence the underlying biophysical interaction. This platform can ultimately be adapted for interactome-wide edge perturbation screens in higher eukaryotes by using emerging CRISPR technologies to introduce compensatory mutations of known edgetic human disease mutations in order to develop a clearer understanding of the function of interactions in our human interactome.
Please contact Michael Calderwood with your questions and comments.