In human immunodeficiency virus (HIV) infection, there is one essential virulence protein, the Nef protein, which is identified as a negative factor due to it having a negative effect on viral replication (1). It performs critical functions in the host during the viral life cycle. The Nef protein is only active during the early stages of the HIV infection (2). It primarily enhances viral replication, infectivity and pathogenesis.
Therefore, Nef has a crucial role in HIV infection as it is based mainly on the Nef performance to downregulate CD4, from the cell surface, and major histocompatibility complex class I (MHC-I), the important surface molecules in T-cell activation (3). In one study they demonstrate that if the cells susceptible to HIV infection, the products of accessory gene can change severity of viral infection (4). Rhesus monkey were tested and demonstrated that a Nef gene is crucial for reaching high virus loads and development of AIDS-like disease in animals (5).
Nef is a 206 amino acid myristoylated peripheral membrane protein (6). It consists of a core domain, an anchor domain and two flexible loops which are located at the N terminus and near the C terminus (7). Nef-core domain has a type II polyproline for Scr family kinase to bind. Various host cell signaling pathways are influenced by Nef to promote HIV-1 replication in AIDS pathogenesis (4). To enhance infectivity through signaling pathway, Nef needs to directly interact with the Scr family of protein tyrosine kinases such as Hck and Lyn, Lck and Fyn (8).
There is a PxxP binding motif of Src homology region 3 (SH3) domain which is present in Nef, that plays an important role in promoting viral replication (8). The best understood phenotypic effect of Nef is the downregulation of CD4 receptor. The viral Nef protein targets the CD4 receptor for endocytosis and is followed by degradation through endosome and lysosome pathway (6). Nef firstly binds to dileucine motif of cytoplasmic tail of CD4 to which the protein tyrosine kinase Lck binds (8). As Nef dileucine motif is required for CD4 internalization, it also binds to
clathrin-coated pits (CCPs) or adaptor protein as it is one of Nef downstream partners. Adaptor protein complexes are crucial components of clathrincoated vesicles. The significant adaptor protein 2 (AP2) enhances endocytosis from plasma membrane while AP1, 3, and 4 contribute in the protein sorting from trans-Golgi network and endosomes to lysosomes (9). Another Nef downstream partner, COP-I Coatomer is a macromolecular structure involved in protein sorting. The COP-I coatomer has a well known role in the Endoplasmic Reticulum-Golgi transport. Nef particularly interacts with the ?
subunit of COP-I coatomer (9). Together, the combination of the functional adaptor protein and COP-I coatomer is mediated by the C-terminal flexible loop of the Nef protein (9). First, Nef is attached to plasma membrane by its myristoyl group and cytoplasmic tail of CD4 with AP2 to facilitate the formation of a clathrin coated pit that leads to the internalization of CD4 (7). Second, degradation of internalized CD4 molecules with enhanced by ? -COPI coatomer in endosome and routing to the lysosomal targeting and degradation of Nef-associated CD4 molecules.
Studies in transgenic mouse have evidently demonstrated that functional Nef expression in CD4+ cells caused an AIDS-like illness and it is required for better infectivity, higher viral load and faster disease development (10). MHC-I downregulation induced by Nef is less well understood than the downregulation of CD4. Cytotoxic T lymphocytes (CTL) in adaptive immune response recognize infected cells before it is eliminated (7). It requires the presentation of viral peptides by MHC-I molecule on the infected cell surface. However, Nef permits HIV-infected cells to escape from CTL.
This also permits infected cells to evade natural killer cells. In the presence of Nef, MHC-I molecules transport into the early Golgi apparatus (7). This is initiated by Nef acidic cluster bind to the complex of PACS-1, required for Nef-mediate downregulation of MHC-1 to TGN, and AP-1 and it will lead the MHC-I molecule to accumulate in the trans-Golgi network (9). Nef also trigger ADP ribosylation factor 6 (ARF6) GTPase through PI3K, thus activating the clathrinindependent transport of MHC-I molecule from cell surface to the TGN through an ARF6 compartment (11).
The infected cells are protected from CTL-mediated killing by downregulation of MHC-1 molecules. To keep the levels of MHC on the cell surface low, Nef must also block the recycling of MHC-I molecules back to the plasma membrane. Therefore, this enhances viral persistence and replication. Nef protein plays an important role by internalizing the several cell surface molecules such as the CD4, CD28, MHC-I and MHC-II molecules (3). Downregulation of CD4 in infected T cells makes sure that infected cells cannot provide effective help during T cell receptor (TCR) engagement.
Whereas downregulation of MHC-I reduces the presentation of HIV peptides to TCR on helper T cells and cytotoxic T cells. The overall effect of the Nef functions is to decrease T cells that facilitate to generate virus-specific antibody and cytotoxic responses. Nef protein presents to enhance viral survival, replication and virion infectivity. These Nef features could allow the development of drugs and treatment that would inhibit the process of HIV infection.
The deletion of Nef protein due to the deletion of Nef gene in HIV infection this leads to have non development in HIV infection (1). Reference 1) Joseph A. M. , Kumar M. , Mitra D. (2005) Nef: “Necessary and Enforcing Factor” in HIV Infection, Current HIV Research. 3, 87-94. 2) Ndolo T. , Dhillon N. K. , Dandekar S. (2002) Simian Immunodeficiency Virus Nef Protein Delays the Progression of CD4+ T Cells through G1/S Phase of the Cell Cycle, Journal of Virology. 76 (8), 3587-3595. 3) Gray L. R. , Gabuzda D. , Cowley D. , Ellett A. , Chiavaroli L. , Wesselingh S.
L. , Churchill M. J. , Gorry P. R. (2011) CD4 and MHC class 1 down-modulation activities of Nef alleles from brain- and lymphoid tissue-derived primary HIV-1 isolates, Journal of NeuroVirology. 17 (1), 82-91. 4) Bimber B. N. , Chugh P. , Lee H. Y. (2009) Nef gene evolution from a single transmitted strain in acute SIV infection, Retrovirology. 6, 57 5) Kestler H. W. , Ringler D. J. , Mori K. (1991) Important of the Nef gene for maintenance of high virus loads and for development of AIDS, Cell. 65, 651-62. 6) O’Neill E. , Kuo L. S. , Foster J. L.
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L. (2008) HIV-1 Nef Targets MHC-I and CD4 for Degradation Via a Final Common ? -COP–Dependent Pathway in T Cells, PLoS Pathogens. 4 (8), e1000131 10) Hanna Z. , Kay D. G. , Cool M. , Jothy S. , Rebai N. , Jolicoeur P. (1998) Transgenic mice expressing human immunodeficiency virus type 1 in immune cells develop a severe AIDS-like disease, J Virol. 72, 121-32. 11) Larsen J. E. , Massol R. H. , Kirchhausen T. (2004) HIV Nef-mediated Major Histocompatibility Complex Class I Down-Modulation Is Independent of Arf6 Activity, Molecular Biology of the cell. 15 (1), 323-331.