Replication kinetics of N- and B-tropic murine leukemia viruses on permissive and nonpermissive cells in vitro

Development of a mouse model for human immunodeficiency virus type 1 (HIV-1) infection has advanced through the progressive identification of host cell factors required for HIV-1 replication. Murine cells lack HIV-1 receptor molecules, do not support efficient viral gene expression, and lack factors necessary for the assembly and release of virions. Many of these blocks have been described using mouse fibroblast cell lines. Here we identify a postentry block to HIV-1 infection in mouse T-cell lines that has not been detected in mouse fibroblasts. While murine fibroblastic lines are comparable to human T-cell lines in permissivity to HIV-1 transduction, infection of murine T cells is 100-fold less efficient. Virus entry occurs efficiently in murine T cells. However, reduced efficiency of the completion of reverse transcription and nuclear transfer of the viral preintegration complex are observed. Although this block has similarities to the restriction of murine retroviruses by Fv1, there is no correlation of HIV-1 susceptibility with cellular Fv1 genotypes. In addition, the block to HIV-1 infection in murine T-cell lines cannot be saturated by a high virus dose. Further studies of this newly identified block may lend insight into the early events of retroviral replication and reveal new targets for antiretroviral interventions.Murine cells are refractory to human immunodeficiency virus type 1 (HIV-1) replication at multiple stages of the viral life cycle. While this has allowed a finer inspection of the role of various host factors in HIV-1 replication, it has been an impediment to the development of a genetically modified mouse permissive to HIV-1 infection. A number of host factors have been identified that are indispensable for replication of HIV-1. Necessary factors absent in murine T cells include the human forms of the HIV-1 receptor and coreceptor molecules, CD4 and CCR5, whose murine orthologs do not support HIV-1 entry (10). In contrast, the murine form of CXCR4 can be utilized as a coreceptor by HIV-1 (7, 69). Postentry, human cyclin T1 is necessary for efficient Tat-mediated transactivation of the HIV-1 long terminal repeat (LTR) (22). Additional blocks in later steps of the HIV-1 life cycle in murine cells include excessive splicing of HIV-1 genomic RNA (49) and defects in Rev function (72). Aberrant splicing of HIV-1 mRNA appears to be partially corrected by the human splice inhibitor p32 (80). HIV-1 particle assembly is also impaired in murine cells (6,53). This restriction can be overcome by fusion of murine cells with human cells, suggesting that murine cells lack a factor (or factors) necessary for HIV-1 Gag assembly and release (52). Substitution of the HIV-1 matrix (MA) region with that of murine leukemia virus (MLV) also circumvents this block, supporting the notion that species-specific cofactors are critical for virion assembly and release (13,63).Not only do mouse cells lack some of the factors necessary for HIV-1 replication, they also express factors that actively interfere with retrovira...

mentioning

confidence: 99%

Murine T Cells Potently Restrict Human Immunodeficiency Virus Infection

Baumann

Unutmaz

Miller

et al. 2004

“…The determinants of N-versus B-tropism have been mapped to a single amino acid residue within the MLV capsid (CA) protein (23). Fv1-mediated restriction of MLV infection can be overcome at high multiplicities of infection, and infection of restricting cells can give rise to multihit titration curves, implying that infection by one virus particle is facilitated by the presence of others (1,7,10,13,31,34). This inhibition of restriction is highly specific in that unrestricted viral particles do not inhibit restriction, presumably because Fv1 does not recognize and therefore cannot be saturated by unrestricted capsids.…”

mentioning

confidence: 99%

Capsid-Dependent and -Independent Postentry Restriction of Primate Lentivirus Tropism in Rodent Cells

Hatziioannou

Cowan

Bieniasz

2004

Retrovirus tropism can be restricted by cellular factors such as Fv1, Ref1, and Lv1 that inhibit infection by targeting the incoming viral capsid. Here, we show that rodent cells exhibit differential sensitivity to infection by vesicular stomatitis virus G-pseudotyped lentiviruses and that differences between human immunodeficiency virus type 1 and simian immunodeficiency virus (SIVmac) infectivity are sometimes, but not always, governed by determinants in capsid-p2. In at least one case, resistance to SIVmac infection could be eliminated by saturation of target cells with noninfectious SIVmac particles. However, cross-saturation experiments and analysis of Fv1-null cells engineered to express natural or artificial Fv1 proteins revealed that lentivirus restriction in mouse cells is independent of Fv1. Overall, these findings indicate that novel restriction factors in rodents can modulate sensitivity to specific primate lentiviruses.

“…Although the mechanisms by which Fv1, Ref1, and Lv1 inhibit infection are unknown, certain characteristics are common to the resistance phenotype conferred by each of these factors. Most notably, the viral determinant of restriction is the capsid protein (CA) (10,12,13,23,30,34,41), and high levels of incoming restricted virions can saturate Fv1, Ref1, or Lv1, thereby abrogating infection resistance (2,3,6,10,11,14,33,35,40,42). Recently, we have shown that saturation of human or African green monkey cells with one retrovirus is capable of abolishing resistance to another retrovirus, provided that both viruses are restricted but irrespective of whether they are closely or distantly related (23).…”

mentioning

confidence: 99%

Species-Specific Tropism Determinants in the Human Immunodeficiency Virus Type 1 Capsid

Hatziioannou

Cowan

Schwedler

et al. 2004

118

109

Retroviral tropism is determined in part by cellular restriction factors that block infection by targeting the incoming viral capsid. Indeed, human immunodeficiency virus type 1 (HIV-1) infection of many nonhuman primate cells is inhibited by one such factor, termed Lv1. In contrast, a restriction factor in humans, termed Ref1, does not inhibit HIV-1 infection unless nonnatural mutations are introduced into the HIV-1 capsid protein (CA). Here, we examined the infectivity of a panel of mutant HIV-1 strains carrying substitutions in the N-terminal CA domain in cells that exhibit restriction attributable to Lv1 or Ref1. Manipulation of HIV-1 CA could alter HIV-1 tropism, and several mutations were identified that increased or decreased HIV-1 infectivity in a target-cell-specific manner. Many residues that affected HIV-1 tropism were located in the three variable loops that lie on the outer surface of the modeled HIV-1 conical capsid. Some tropism determinants, including the CypA binding site, coincided with residues whose mutation conferred on HIV-1 CA the ability to saturate Ref1 in human cells. Notably, a mutation that reverses the infectivity defect in human cells induced by CypA binding site mutation inhibits recognition by Ref1. Overall, these findings demonstrate that exposed variable loops in CA and a partial CypA "coat" can modulate restriction and HIV-1 tropism and suggest a model in which the exposed surface of the incoming retroviral capsid is the target for inhibition by host cell-specific restriction factors.