Abstract
Extracellular vesicles (EVs) influence cell phenotypes and functions via protein, nucleic acid and lipid cargoes. EVs are heterogeneous, due to diverse biogenesis mechanisms that remain poorly understood. Our previous study revealed that the endoplasmic reticulum (ER) membrane contact site (MCS) linker protein VAP-A drives biogenesis of a subset of RNA-enriched EVs. Here, we examine the protein content of VAP-A-regulated EVs. Using label-free proteomics, we identified down- and up-regulated proteins in sEVs purified from VAP-A knockdown (KD) colon cancer cells. Gene set enrichment analysis (GSEA) of the data revealed protein classes that are differentially sorted to SEVs dependent on VAP-A. STRING protein-protein interaction network analysis of the RNA-binding protein (RBP) gene set identified several RNA functional machineries that are downregulated in VAP-A KD EVs, including ribosome, spliceosome, mRNA surveillance, and RNA transport proteins. We also observed downregulation of other functionally interacting protein networks, including cadherin-binding, unfolded protein binding, and ATP-dependent proteins.