Among the accessory proteins encoded by HIV-1, the 27 kDal membrane associated myristoylated Nef has proved to be enigmatic. The lack of consensus for the results of current in vitro assays measuring Nef effect on virus replication and transcription has required a shift in the focus from the virological to the cellular physiological roles of Nef. There is reasonable agreement in the literature on the potential roles of Nef in modulating certain cellular receptors. In this report, we have tried to address specific properties of Nef in this area by mechanistic studies of Nef effects on CD4 and major histocompatibility complex Class I receptor (MHC-I) and chemokine signalling pathways. We have observed in the past that Nef downregulated CD4 by a bi- modal mechanism. Unquestionably, Nef enhanced CD4 endocytosis was the only demonstrable effect when Nef was expressed acutely in T cell lines that express CD4 constitutively. However, when Nef and CD4 were co-expressed in cells lacking endogenous CD4, we demonstrated a Nef induced defect in the early steps of CD4 synthesis and transport in addition to the accelerated endocytosis of the CD4 receptor. The cytoplasmic CD4 defect may have been due to interference with the intracellular vesicular traffic of certain nascent proteins in Nef expressing cells. The ensuing delay in the recruitment of selected membrane proteins such as CD4 into these vesicles may lead to their premature degradation. MHC-I expression levels and usage status are important in AIDS pathogenesis since perturbation of antigen presentation by infected cells may allow them to escape T killer cell surveillance that is a vital immune mechanism against virus infection. We undertook an exhaustive analysis of potential Nef effects on MHC-I. We found that the Nef effect on MHC-I was quite variable with different MHC-I alleles and different cells. Whereas it is likely that Nef may have enhanced endocytosis and degradation of the susceptible receptor, we could not find consistent evidence for this. Two distinct sets of chemokine co-receptors have been shown to be required for HIV entry in the CD4 positive cells. They are the CCR5 and CXCR4 receptors that are used by the macrophage (M)- tropic and T cell (T)-tropic strains of HIV respectively. Both the CD4 and the chemokine receptor have to be present on the cell to allow HIV entry. Although HIV entry takes place in cells that express mutant coreceptors that have lost the ability to signal through chemokines, it is not clear whether the magnitude of early virus replication events are influenced by the chemokine signaling pathways. We found that Nef proteins of HIV and SIV differentially modulated CCR5 and CXCR4 with potential implications for tropism switching during natural infection.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Intramural Research (Z01)
Project #
1Z01AI000528-11
Application #
6098957
Study Section
Special Emphasis Panel (LMM)
Project Start
Project End
Budget Start
Budget End
Support Year
11
Fiscal Year
1998
Total Cost
Indirect Cost
City
State
Country
United States
Zip Code
Rose, Jeremy J; Janvier, Katy; Chandrasekhar, Soundararajulu et al. (2005) CD4 down-regulation by HIV-1 and simian immunodeficiency virus (SIV) Nef proteins involves both internalization and intracellular retention mechanisms. J Biol Chem 280:7413-26
Rose, Jeremy J; Foley, John F; Murphy, Philip M et al. (2004) On the mechanism and significance of ligand-induced internalization of human neutrophil chemokine receptors CXCR1 and CXCR2. J Biol Chem 279:24372-86
Janvier, Katy; Kato, Yukio; Boehm, Markus et al. (2003) Recognition of dileucine-based sorting signals from HIV-1 Nef and LIMP-II by the AP-1 gamma-sigma1 and AP-3 delta-sigma3 hemicomplexes. J Cell Biol 163:1281-90
Venkatesan, Sundararajan; Rose, Jeremy J; Lodge, Robert et al. (2003) Distinct mechanisms of agonist-induced endocytosis for human chemokine receptors CCR5 and CXCR4. Mol Biol Cell 14:3305-24
Shaheduzzaman, Syed; Krishnan, Vyjayanthi; Petrovic, Ana et al. (2002) Effects of HIV-1 Nef on cellular gene expression profiles. J Biomed Sci 9:82-96
Nam, Y S; Petrovic, A; Jeong, K S et al. (2001) Exchange of the basic domain of human immunodeficiency virus type 1 Rev for a polyarginine stretch expands the RNA binding specificity, and a minimal arginine cluster is required for optimal RRE RNA binding affinity, nuclear accumulation, and trans-activa J Virol 75:2957-71
Venkatesan, S; Petrovic, A; Locati, M et al. (2001) A membrane-proximal basic domain and cysteine cluster in the C-terminal tail of CCR5 constitute a bipartite motif critical for cell surface expression. J Biol Chem 276:40133-45
Jeong, K S; Nam, Y S; Venkatesan, S (2000) Deletions near the N-terminus of HIV-1 Rev reduce RNA binding affinity and dominantly interfere with Rev function irrespective of the RNA target. Arch Virol 145:2443-67
Van Ryk, D I; Venkatesan, S (1999) Real-time kinetics of HIV-1 Rev-Rev response element interactions. Definition of minimal binding sites on RNA and protein and stoichiometric analysis. J Biol Chem 274:17452-63