HIV Nef- and Notch1-dependent Endocytosis of ADAM17 Induces Vesicular TNF Secretion in Chronic HIV Infection

Tumor necrosis factor (TNF) is a key cytokine in HIV replication and pathogenesis. For reasons that are not entirely clear, the cytokine remains upregulated despite anti-retroviral therapy (ART). Here we demonstrate that HIV Nef induces an alternative TNF secretion mechanism that remains active in chronic infection. Ingestion of Nef-containing plasma extracellular vesicles (pEV) from ART patients by primary immune cells, but also Nef expression, induced intracellular proTNF cleavage and secretion of vesicular TNF endosomes. Key event was the Nef-mediated routing of the TNF-converting enzyme ADAM17 into Rab4+ early endosomes and the Rab27+ secretory pathway. Analysis of lymph-node tissue by multi-epitope-ligand-cartography (MELC) confirmed a vesicular TNF secretion phenotype that co-localized with persistent Nef expression, and implicated Notch1 as an essential co-factor. Surprisingly Notch1 had no transcriptional effect but was required for the endosomal trafficking of ADAM17. We conclude that Nef expression and Nef-containing pEV mobilize TNF from endosomal compartments in acute and chronic infection. •Nef/ADAM17 containing extracellular vesicles induce an endosomal TNF secretion type in primary target cells.•The mechanism required the shuttling of ADAM17 into Rab4+ endosomal compartments in a Notch1-dependent manner.•The mechanism could be demonstrated in tissue by multi-epitope-ligand-cartography (MELC) technology. Despite antiviral therapy, plasma levels of TNF remain upregulated and likely play a role in many comorbidities seen in chronic HIV infection. We found that this is due to high levels of HIV-induced plasma extracellular vesicles (pEV) containing the TNF processing ADAM17 protease. Interestingly these vesicles induced a different TNF secretion type. Whereas TNF is usually shed from the plasma membrane, pEV mobilized intracellular TNF storage compartments, secreting endosomal vesicles. We could confirm this mechanism analyzing lymph node tissue sections by a novel immunostaining technology. Our report supports our previous publication implying ongoing viral activity despite successful antiretroviral therapy.