Human immunodeficiency virus type 1 Nef: adapting to intracellular trafficking pathways

Abstract

The Nef protein of primate lentiviruses is a unique protein that has evolved in several ways to manipulate the biology of an infected cell to support viral replication, immune evasion, pathogenesis, and viral spread. Nef is a small (25- to 34-kDa), myristoylated protein that binds to a collection of cellular factors and acts as an adaptor to generate novel protein interactions to accomplish specific functions. Of the many biological activities attributed to Nef, the reduction of surface levels of the viral receptor (CD4) and antigen-presenting molecules (major histocompatibility complex class I) has been intensely examined; recent evidence demonstrates that Nef utilizes multiple, distinct pathways to affect these proteins. To accomplish this, Nef promotes the formation of multiprotein complexes, recruiting host adaptor proteins to commandeer intracellular vesicular trafficking routes. The altered trafficking of several other host molecules has also been reported, and an emerging theory suggests that Nef generates pleiotrophic effects in the secretory and endocytic pathways that reprogram intracellular protein trafficking and may ultimately provide an efficient platform for viral assembly. This review critically discusses some of the major findings regarding the impact of human immunodeficiency virus type 1 Nef on host protein transport and addresses some emerging directions in this area of human immunodeficiency virus biology.

FIG. 1.

Sequence alignment and protein-protein interaction domains in HIV-1 and SIV Nef. The HIV-1_NL4-3 Nef and SIV_mac239 Nef sequences are aligned. Boxes indicate protein motifs and their cellular binding partners (in parentheses). Residues important for protein interactions are underlined. The asterisks indicate residues in SIV_mac239 Nef that are homologous to important residues in HIV_NL4-3 Nef that have not been tested for functional homology.

FIG. 2.

HIV-1 Nef can associate with components of intracellular vesicular transport pathways. The cartoon outlines the major known trafficking pathways between intracellular organelles. Vesicle coat proteins that bind to HIV-1 Nef are indicated at their proposed sites of cargo selection.

FIG. 3.

Nef-associated protein complexes. (A to C) Schematic diagrams of protein-protein interactions that have been reported for Nef-mediated disruption of MHC-I trafficking (A), CD4 downregulation by Nef (B), and AP binding mediated by the Nef dileucine motif (C). (D) Summary of the experimental evidence supporting the numbered protein-protein interactions depicted in A to C.

FIG. 4.

Differential effects on MHC-I allotypes enable HIV-1 Nef to escape both CTL and NK cell surveillance. (A) A normal cell that does not harbor any viral antigens is not affected by cell-mediated immune surveillance. (B) Virally infected cells are recognized and lysed due to CTL recognition of the antigen-MHC-I complex. (C) Virally infected cells that have reduced levels of all MHC-I allotypes (nonselective disruption) are lysed by NK cells due to lack of inhibitory MHC-I allotypes (HLA-C and HLA-E). (D) The selective removal of surface HLA-A and HLA-B allotypes prevents CTL recognition, while the maintenance of HLA-C and HLA-E expression protects the infected cell from NK cell lysis.

FIG. 5.

MHC-I and AP-1 colocalize in Nef-expressing T cells. CEM T cells that stably expressed yellow fluorescent protein-tagged HLA-A2 (green) were transduced with a control adenovirus (nef⁻) or an adenovirus that expressed HIV-1 Nef (nef⁺). Cells were adhered to glass slides, and the gamma subunit of AP-1 (γ-adaptin, red) was detected by indirect immunofluorescence. Individual Z sections through the middle of the cell were collected using a Zeiss LSM 510 confocal microscope. Scale bar = 5 μm.

FIG. 6.

Model for MHC-I transport in Nef-expressing cells. (A) Nef binds to MHC-I early in the secretory pathway. (B) Nef inhibits the transport of newly synthesized MHC-I to the cell surface. (C) MHC-I that escapes the transport block can be rerouted from the cell surface. (D) Nef and MHC-I accumulate in the TGN at steady state, where they cooperate to recruit AP-1. (E) MHC-I is delivered to acidic compartments for degradation.

FIG. 7.

Summary of the proposed mechanisms for Nef-induced CD4 downregulation. (A) Nef targets CD4 molecules from the cell surface, resulting in their enhanced internalization. (B) Nef causes the degradation of CD4 in acidic compartments.

FIG. 8.

Proposed mechanism for differential effects of Nef on CD4 and MHC-I. In this model Nef preferentially binds to hypophosphorylated MHC-I molecules early in the secretory pathway (83) and recruits AP-1 at its normal localization in the Golgi. In contrast, Nef may preferentially associate with plasma membrane forms of CD4 and recruits AP-2 at this location.