Agradecimentos: We thank Juliano Coelho da Silveira and Fausto Bruno dos Reis Almeida for providing NTA equipment available at the Faculty of Zootechnics and Food Engineering - FZEA/USP (Pirassununga, SP), and at Ribeirão Preto Medical School FMRP/USP (Ribeirão Preto, SP), respectively. We thank Roberta R. C. Rosales for excellent technical assistance. We acknowledge the Mass Spectrometry Laboratory at Brazilian Biosciences National Laboratory, CNPEM, Campinas, Brazil for their support with the analyses. Funding and additional information M. E. d. S.-J., C. S. d. C., Y. C. J., and R. L. R. thank the São Paulo Research Foundation (FAPESP) for doctoral fellowships. L. A. T. thank FAPESP for a postdoctoral fellowship. This work was supported by FAPESP grants no. 18/00297-7 and 22/15928-8 to L. L. P. D. S. Ver menos

Abstract: Extracellular vesicles (EVs) are biomolecule carriers for intercellular communication in health and disease. Nef is a HIV virulence factor that is released from cells within EVs and is present in plasma EVs of HIV-1 infected individuals. We performed a quantitative proteomic analysis to fully characterize the Nef-induced changes in protein composition of T cell-derived EVs and identify novel host targets of HIV. Several proteins with well-described roles in infection or not previously associated with HIV pathogenesis were specifically modulated by Nef in EVs. Among the downregulated proteins are the interferon-induced transmembrane 1, 2, and 3 (IFITM1-3) proteins, broad-spectrum antiviral factors known to be cell-to-cell transferable by EVs. We demonstrate that Nef depletes IFITM1-3 from EVs by excluding these proteins from the plasma membrane and lipid rafts, which are sites of EVs biogenesis in T cells. Our data establish Nef as a modulator of EVs' global protein content and as an HIV factor that antagonizes IFITMs Ver menos