Affinity-ranking BIN1 interactions underpinning centronuclear myopathy

Abstract

Deletion of the protein-protein interaction SH3 domain of the membrane remodeling amphiphysin 2 (BIN1) protein was found to lead to centronuclear myopathy in patients, yet only few interaction partners of BIN1 SH3 have been identified so far, precluding a better understanding of the pathomechanism. Here we used the holdup assay to proteome-wide measure steady-state affinity constants of BIN1 SH3 domain for thousands of full-length cellular proteins, as well as for hundreds of putative SH3-binding sites found within the identified BIN1 partners. Besides confirming known partners, such as dynamin 2 (DNM2), we also identified and affinity-characterized numerous others, like SMCHD1, which were previously implicated in different neuromuscular disorders. We also assessed the impact of a set of rare natural BIN1 SH3 domain variants on affinity interactomes and identified potentially harmful ones that exhibited perturbed affinity profiles, whose impacts were confirmed in a cellular assay for BIN1-mediated membrane remodeling, tentatively connecting them to neuromuscular disorders. In the study, we develop a new affinityinteractomic strategy, which can be generally applied to study the consequences of disease-associated genomic variants of any kind.