New ligands of membrane-associated HIV-1 Nef

HIV-1 Nef protein contributes essentially to the pathology of AIDS by a variety of protein-protein-interactions within the host cell. The versatile functionality of Nef is increased through different conformational states and post-translational modifications such as myristoylation of the N-terminal glycine residue. Up to now many interaction partners like βCOP, thioesterase II, Eed and TCRζ have been identified using classical yeast two-hybrid screens. Such screens rely on transcriptional activation of reporter genes in the nucleus to detect interactions. Thus, the detection of interaction partners that are integral membrane proteins, membrane-associated proteins or other proteins that do not translocate into the nucleus is hindered.

In the present study a split-ubiquitin based yeast two-hybrid screen was established. To identify novel membrane-localized interaction partners of Nef its myristoyl-anchor was replaced with the small membrane anchor Ost4p. Over 80% of the identified interaction partners of Ost4p-Nef are transmembrane proteins, which are localized in the plasma membrane or the ER/Golgi membrane. The numerous novel putative interaction partners may help to elucidate the molecular basis of many HIV-associated pathological observations in HIV-positive patients.