Primary T-cells from human CD4/CCR5-transgenic rats support all early steps of HIV-1 replication including integration, but display impaired viral gene expression

Abstract

Background: In vivo studies on HIV-1 pathogenesis and testing of antiviral strategies have been hampered by the lack of an immunocompetent small animal model that is highly susceptible to HIV-1 infection. Since native rodents are nonpermissive, we developed transgenic rats that selectively express the HIV-1 receptor complex, hCD4 and hCCR5, on relevant target cells. These animals display a transient low-level plasma viremia after HIV-1 YU-2 infection, demonstrating HIV-1 susceptibility in vivo. However, unlike macrophages, primary CD4 T-cells from double-transgenic animals fail to support viral spread ex vivo. To identify quantitative limitations or absolute blocks in this rodent species, we quantitatively assessed the efficiency of key steps in the early phase of the viral replication cycle in a side-by-side comparison in infected cell lines and primary T-cells from hCD4/hCCR5-transgenic rats and human donors.

Endogenous CD317/Tetherin Limits Replication of HIV-1 and Murine Leukemia Virus in Rodent Cells and Is Resistant to Antagonists from Primate Viruses

“…3A and B, flow cytometric quantification of cell surface-exposed mCD317 demonstrated potent (shRNA1, mean fluorescence intensity [MFI] of CD317 surface expression of 66) or intermediate (shRNA3, MFI of CD317 surface expression of 223; shRNA4, MFI of CD317 surface expression of 196) depletion of the restriction factor relative to that obtained with a control shRNA (MFI of CD317 surface expression of 832) in neomycin-selected S1A.TB cultures. To monitor the effect of variable mCD317 levels on S1A.TB cells on MLV replication, we performed infection studies with MLV-GFP, a replication-competent MoMLV which encodes an egfp gene driven from an IRES in the untranslated region between the env gene and the 3Ј LTR (14). Following infection with MLV-GFP at a low multiplicity of infection (MOI), moderate viral spread was detected in control shRNA-expressing (Fig.…”

“…pMoMLV-GFP was constructed by introducing the egfp gene driven from an internal ribosomal entry site (IRES) into the untranslated region between the env gene and the 3Ј long terminal repeat (LTR) of MoMLV (GenBank accession no. AF033811.1) at unique NotI/ MluI sites, and viral stocks (MLV-GFP) were generated as previously reported (14). Both the virus titers and reverse transcriptase activities of MLV-GFP and the wild-type virus produced in BOSC cells were comparable (V. Hermann and H.-G. Kräusslich, unpublished data).…”

“…Cells from rats and mice do not support or only inefficiently support various steps of the HIV-1 replication cycle (2,3,14,28,48,54,59). Molecular characterization of some of these species-specific barriers has revealed the inability of rodent orthologs of cellular factors, essential for HIV-1 replication in human cells, to support distinct steps of the viral life cycle.…”

“…Molecular characterization of some of these species-specific barriers has revealed the inability of rodent orthologs of cellular factors, essential for HIV-1 replication in human cells, to support distinct steps of the viral life cycle. Specifically, expression of the HIV-1 receptor complex, as well as of the HIV-1 Tat-interacting protein hCyclin T1, has overcome barriers in the early phase of the HIV-1 replication cycle at the levels of entry and viral transcription, respectively (3,14,27,50,54). Corresponding transgenesis in laboratory rats has resulted in considerable permissivity for HIV-1 in vivo, characterized by high proviral loads in lymphatic organs (27).…”

Pharmacovirological Impact of an Integrase Inhibitor on Human Immunodeficiency Virus Type 1 cDNA Species In Vivo

“…Specifically, the in vivo relationship of pharmacokinetics and the virological impact of raltegravir is still poorly defined, as dose-ranging studies of raltegravir showed equal efficacies of viral-load reduction across a range of doses (20,40,42). Animal models, such as HIV-susceptible transgenic Sprague-Dawley rats expressing the HIV-1 receptor complex (17,19,33,41), can support the preclinical validation process of antiviral compound candidates, allowing the quantification of their virological impacts and serum concentrations. In these in vivo models, an antiviral compound can be titrated from no effect to full effect, and mechanistic hypotheses can be probed under well-controlled experimental conditions.…”