The CYFIP2 Arg87Cys Mutation Reshapes the Interactome and Alters Stress Fiber-Associated Proteins

The Arg87Cys mutation in CYFIP2 has been implicated in early-onset developmental and epileptic encephalopathy (DEE), yet its molecular consequences remain incompletely understood. CYFIP2 functions as part of the WAVE regulatory complex (WRC), where it plays a key role in actin filament polymerization and interacts with RNA-binding proteins (RBPs) to regulate mRNA translation. Previous studies have demonstrated that this mutation alters the structure of CYFIP2, resulting in a gain of function in the WRC. Additionally, the mutation appears to promote the formation of intracellular granules that colocalize with RBPs; however, the impacts of this mutation on these interactions remain unclear. Here, we combined cross-linking mass spectrometry (XL-MS) and label-free quantitative proteomics to investigate how this mutation alters the protein interactome and actin regulatory pathways. XL-MS data revealed exclusive cross-links between actin and tropomyosin isoforms in the Arg87Cys variant, suggesting altered cytoskeletal dynamics. Quantitative proteomic analysis of patient-derived neural progenitor cells corroborated these findings by revealing an increase in the abundance of stress-fiber-associated proteins. Altogether, our results point to a broader impact of CYFIP2 mutations on cytoskeletal organization and protein interaction networks with potential implications beyond the nervous system. This is the first study to our knowledge to suggest a link between the Arg87Cys mutation and dysregulation of stress-fiber-related proteins, indicating possible disruption of other actin-processing pathways.