A Truncated Form of Nef Selected during Pathogenic Reversion of Simian Immunodeficiency Virus SIVmac239[Delta]nef Increases Viral Replication

• Published in: Journal of virology 2003-01, Vol.77 (2), p.1245-1256
• Main Authors: Chakrabarti, LA, Metzner, K J, Ivanovic, T, Cheng, H, Louis-Virelizier, J, Connor, R I, Cheng-Mayer, C
• Format: Article
• Language: English
• ISSN: 0022-538X
• DOI: 10.1128/JVI.77.2.1245-1256.2003

The live, attenuated vaccine simian immunodeficiency virus SIVmac239[Delta]nef efficiently protects rhesus macaques against infection with wild-type SIVmac but occasionally causes CD4 super(+) T-cell depletion and progression to simian AIDS (SAIDS). Virus recovered from a vaccinated macaque (Rh1490) that progressed to SAIDS had acquired an additional deletion in the nef gene, resulting in a frameshift that restored the original nef open reading frame (R. I. Connor, D. C. Montefiori, J. M. Binley, J. P. Moore, S. Bonhoeffer, A. Gettie, E. A. Fenamore, K. E. Sheridan, D. D. Ho, P. J. Dailey, and P. A. Marx, J. Virol. 72:7501-7509, 1998). Intravenous inoculation of the Rh1490 viral isolate into four naive rhesus macaques induced CD4 super(+) T-cell depletion and disease in three out of four animals within 2 years, indicating a restoration of virulence. A DNA fragment encompassing the truncated nef gene amplified from the Rh1490 isolate was inserted into the genetic backbone of SIVmac239. The resulting clone, SIVmac239-[Delta]2nef, expressed a Nef protein of approximately 23 kDa, while the original SIVmac239[Delta]nef clone expressed a shorter protein of 8 kDa. The revertant form of Nef did not cause downregulation of CD4, CD3, or major histocompatibility complex class I. The infectivity of SIVmac239-[Delta]2nef was similar to that of SIVmac239[Delta]nef in single-cycle assays using indicator cell lines. In contrast, SIVmac239- [Delta]2nef replicated more efficiently than SIVmac239[Delta]nef in peripheral blood mononuclear cell (PBMC) cultures infected under unstimulated conditions. The p27 Gag antigen levels in SIVmac239-[Delta]2nef-infected cultures were still lower than those obtained with wild-type SIVmac239, consistent with a partial recovery of Nef function. The transcriptional activity of long terminal repeat (LTR)-luciferase constructs containing the nef deletions did not differ markedly from that of wild-type LTR. Introduction of a premature stop codon within Nef-[Delta]2 abolished the replicative advantage in PBMCs, demonstrating that the Nef-[Delta]2 protein, rather than the structure of the U3 region of the LTR, was responsible for the increase in viral replication. Taken together, these results show that SIV with a deletion in the nef gene can revert to virulence and that expression of a form of nef with multiple deletions may contribute to this process by increasing viral replication.