Determinants in HIV-1 Nef for enhancement of virus replication and depletion of CD4+ T lymphocytes in human lymphoid tissue ex vivo

Background: HIV-1 Nef critically contributes to AIDS in part by augmenting virus titers in infected individuals. Analyzing which of Nef's activities contribute to HIV pathogenesis has been hampered by the lack of a cell culture model in which Nef exerts pronounced effects on HIV replication. The human lymphoid aggregate culture (HLAC) from tonsil maintains the cell populations and cytokine milieu found in vivo, supports a productive infection without exogenous stimulation, and Nef contributes to efficient HIV-1 replication as well as CD4(+) T cell depletion in this experimental ex vivo-model. Results: To identify determinants in Nef that mediate these activities, we infected HLAC with a panel of isogenic HIV-1(NL4-3) strains that encode for well-characterized mutants of HIV-1(SF2) Nef. Determination of HIV-1 replication revealed that enhancement of the virus spread by Nef is governed by a complex set of protein interaction surfaces. In contrast, increased CD4(+) T lymphocyte depletion depended on only two protein interaction surfaces in Nef that mediate either downregulation of cell surface CD4 or interaction with the NAKC signalosome. Consistently, in HLAC from 9 out of 14 donors, Nef enhanced CD4(+) T cell depletion in the absence of a significant effect on virus replication. Moreover, our results suggest that this Nef-dependent enhancement in depletion occurred predominately in uninfected bystander CD4(+) T cells. Conclusion: Our findings suggest that Nef facilitates depletion of CD4(+) T lymphocytes in HIV-1-infected lymphoid tissue ex vivo by increasing the pool of productively infected cells and by sensitizing bystander cells for killing. This ability might contribute to Nef's pathogenic potential in vivo.