Javier Muñoz (Presenting Author) | Cell Signaling and Clinical Proteomics Group | Plaza Cruces s/n, 48903, Barakaldo, Spain
Abstract
Extracellular vesicles (EVs) are key mediators of intercellular communication and hold potential as biomarkers. However, the precise characterization of their molecular cargo is challenged by the complex spectrum of EV subtypes and co-isolated nanoparticles. Here, leveraging protein correlation profiling along density gradients, we systematically analyze over 9,000 proteins in human cancer cell lines and biofluids, providing a rigorous reassessment of virtually all protein constituents associated with small EVs (sEVs) and non-vesicular entities. We reveal that sEVs primarily incorporate plasma membrane proteins via selective cargo-loading mechanisms, with minimal inclusion of intraluminal soluble proteins. In contrast, the abundant cytosolic proteins frequently detected in sEVs preparations are bound to the outer surface of sEVs, stemming from extracellular contamination by cell debris and intracellular interactions with autophagic material. Our work provides a reference resource for understanding biogenesis, molecular determinants of cargo selection, and the functional roles of sEVs.
