3rd Conference on Retroviruses and Opportunistic Infections Washington, DC - January 28-February 1, 1996

Conf Retroviruses Opportunistic Infect 1996 Jan 28-Feb 1; 3rd:57 (abstract no. 24)

Felber BK¹, Valentin A², von Gegerfelt A¹, Zolotukhin AS¹, Tabernero C¹, Bear J¹, Pavlakis GN²
Human Retrovirus Pathogenesis Group, Human Retrovirus Section, ABL-BRP, NCI-FCRDC Frederick MD.

Molecular clones of HIV-1 and SlVmac239 were constructed containing multiple point mutations in rev and the RRE that do not affect the overlapping tat and env reading frames. Insertion of the cis-acting posttranscriptional control element of simian retrovirus type 1 (SRV-1CTE) into these c!ones resulted in efficient virus expression independent of the Rev/RRE regulatory system. The SRV-mediated expression is thought to involve cellular factor(s) functionally analogous to Rev that can activate expression at the posttranscriptional level. Computer predictions indicated that the SRV-1 RNA element forms a strong stem-bulge structure. Detailed structure/function analysis has revealed the importance of both primary and secondary structure of the element. Virus stocks produced after transfections of the HIV-1 hybrid molecular clone in human cells were infectious upon cell-free transmission, replicated about 5-10 times less efficiently than wild-type virus, and were propogated continuously in human PBMCS for more than 14 months as well as in human macrophages. Growth characteristics and sequence analysis after long-term culture demonstrated that no RRE(+) or Rev(+) revertants developed, and that the SRV-1 CTE remained intact. High titer Rev/RRE independent virus stock was produced and used to infect chimpanzee and pigtail macaque PBMCS. PBMCS from both species support the SRV-mediated HIV-1 virus production. Infection of a panel of human cell lines showed that the hybrid virus has a distinct cell type range. To further explore the role of Rev in the Ientiviral life cycle, we have also generated Rev-independent clones of SIVmac239 containing the SRV-1 CTE. The SIV hybrid virus has attenuated growth properties in vitro in rhesus macaque PBMCS and the CEMX174 T-cell line. Our data demonstrate that the Rev regulation can be replaced by other posttranscriptional mechanisms, and that Rev is not essential for virus replication in vitro. The availability of the SIV hybrid virus will allow us to investigate the role of Rev in virus propagation in vivo. The replacement of the Rev/RRE regulatory axis may generate viruses with altered biological properties in vivo and it may allow the production of attenuated nonpathogenic viral strains.

Research sponsored by the National Cancer Institute, DHHS, under contract with ABL.

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Copyright © 1996 - Foundation for Retrovirology and Human Health . Reproduction of this abstract (other than one copy for personal reference) must be cleared through the Foundation for Retrovirology and Human Health.