INI1 /hSNF5 in HIV-1 replication: Characterisation of a transdominant mutant
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Integration is a key event in the replication of all retroviruses including the human immunodeficiency virus type 1 (HIV-1). Integrase (IN), that catalyzes integration of viral cDNA into the host genome and the host factors associated with this enzyme are attractive new targets for developing anti-virals against HIV-1, for therapeutic interventions of AIDS. Integrase Interactor 1 (INI1)/hSNF5 is a human protein that specifically binds to HIV-1 IN. We carried out genetic analysis to determine the role of INI1/hSNF5 in HIV-1 replication and to develop novel inhibitors against this virus, by isolating and characterizing transdominant mutants of INI1/hSNF5.;Previously, we have demonstrated that a highly conserved 60 as Rpt 1 region of INI1 is necessary and sufficient to bind to HIV-1 IN. We expressed a small fragment of INI1/hSNF5 (termed S6) spanning this minimal-IN binding domain in mammalian producer cells, and found that it profoundly inhibited HIV-1 particle production (10,000 to 100,000 fold). Stable expression of S6 protected the T-cells from infection by full-length clones of HIV-1. Mutations in S6 or IN that disrupt IN-INI1 interaction abrogated the inhibitory effect. An IN-deficient HIV-1 containing Vpr-RT-IN in trans was not affected by S6 (Yung et al., 2001).;Studies in our laboratory (Craig et al., 2002) have demonstrated that the presence of a masked NES in INI1/hSNF5 (which is unmasked in S6) mediates the hCRM1-dependent nuclear export. We found that disrupting the NES of S6 resulted in the abrogation of the inhibition indicating the significance of nuclear export properties for this process. To determine the specificity of S6-mediated inhibition, we first tested the interaction of INI1/hSNF5 with integrases from other primate retroviruses using in vitro binding assays. We found that the interaction of INI1/hSNF5 is highly specific and is restricted to that of HIV-1 IN. Consistent with this observation, our results suggest that S6 does not significantly inhibit particle production of other lentiviruses. These studies underscore the significance of IN-interaction in S6-mediated inhibition of HIV-1 particle production.;Our results suggest that ectopic expression of S6 specifically inhibits the HIV-1 particle production in a transdominant manner via its specific interaction with IN within Gag-Pro-Pol, providing a novel strategy to control HIV-1 replication.
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