Extracellular Nef Protein Targets CD4⁺T Cells for Apoptosis by Interacting with CXCR4 Surface Receptors

Abstract

The effects of soluble Nef protein on CD4؉ T cells were examined. CD4 ؉ -T-cell cultures exposed to soluble Nef were analyzed for apoptosis by terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling and hallmarks of apoptosis including cytoplasmic shrinkage, nuclear fragmentation, DNA laddering, and caspase activation. We observed dose-and time-dependent inductions of apoptosis. DNA laddering and activated caspase 3 were also evident. Cells treated with Nef/protein kinase inhibitor complexes were protected from Nef-induced apoptosis, suggesting possible roles for protein kinases in the apoptosis pathway. Similarly, cells treated with Nef/anti-Nef antibody complexes were protected from Nef-induced apoptosis. The cellular receptor responsible for Nef-induced apoptosis was identified through antibody-and ligand-blocking experiments as a receptor commonly involved in viral entry. CXCR4 antibodies, as well as the endogenous ligand SDF-1␣, were effective in blocking Nef-induced apoptosis, while CCR5 and CD4 antibodies were ineffective. Moreover, a CXCR4-deficient cell line, MDA-MB-468, which was resistant to Nef-induced apoptosis, became sensitive upon transfection with a CXCR4-expressing vector. This study suggests that extracellular Nef protein could contribute to the decline of CD4 counts prior to and during the onset of AIDS in patients with human immunodeficiency virus type 1 infections.

Pathology of Pulmonary Hypertension

“…In human monocyte-derived macrophages (MDMs), Nef activates the STAT1 pathway and the secretion of MIP-1, IL-1-α, IL-6, and TNF-α [102]. Extracellular Nef found in HIV patients (approximately 10 ng/ml) enters the vascular endothelium in vivo via CXCR4 [103]. Finally, Nef can be proapoptotic or prosurvival, depending on the context of expression and the particular cell type [104].…”

HIV Type 1 Nef Is Released from Infected Cells in CD45+Microvesicles and Is Present in the Plasma of HIV-Infected Individuals

“…19,20 We have also shown that cellfree tissue culture supernatant contains Nef, which can also induce apoptosis in T cells. To confirm the apoptotic activity of the microvesicular Nef isolated in this study we exposed Jurkat and U937 cells to Nef-GFP microvesicles for 3, 6, or 18 h (Fig.…”

“…[16][17][18] More specifically, early reports demonstrate that soluble extracellular Nef induces apoptosis in T cells via a CXCR4-dependent interaction. 19,20 Given this result, we have considered whether soluble Nef is released from infected cells and could play a significant role in mediating bystander apoptosis in CD4 þ T cells. Nef has no known intrinsic enzymatic activities and yet it promotes pleiotropic effects such as enhancement of infectivity, immune evasion, and cellular activation.…”

Risk of Echocardiographic Pulmonary Hypertension in Individuals with Human Immunodeficiency Virus–Hepatitis C Virus Coinfection

“…Although rare, PH complicating HIV infection has not decreased appreciably with available antiretroviral therapy (ART), and the mechanistic link between PH and HIV has not been clearly established (4). Although HIV ribonucleic acid (RNA) has not been isolated in the pulmonary vasculature, HIV proteins such as Tat and Nef cause endothelial dysfunction (6)(7)(8)(9)(10)(11)(12). It stands to reason that other viral infections may cause downstream fibroproliferation and remodeling in the pulmonary circulation.…”