An adaptable genomic-proteomic approach to characterize cell-surface Vimentin's membrane topology and interactome

When found on the cell surface, Vimentin, a cytoskeletal protein, engages in multiple pathophysiological processes. Here, in human fibroblasts and endothelial cells, we used CRISPR-Cas9 to insert a hemagglutinin tag at six different loci within the vim gene. We then utilized tag immunodetection to define cell surface Vimentin's (csVim) membrane topology, followed by proximity biotinylation to identify its extracellular interactome; validating a subset of hits with a quantitative in vitro binding assay.