HIV-1 replication in cell lines harboring INI1/hSNF5 mutations

BACKGROUND: INI1/hSNF5 is a cellular protein that directly interacts with HIV-1 integrase (IN). It is specifically incorporated into HIV-1 virions. A dominant negative mutant derived from INI1 inhibits HIV-1 replication. Recent studies indicate that INI1 is associated with pre-integration and reverse transcription complexes that are formed upon viral entry into the target cells. INI1 also is a tumor suppressor, biallelically deleted/mutated in malignant rhabdoid tumors. We have utilized cell lines derived from the rhabdoid tumors, MON and STA-WT1, that harbor either null or truncating mutations of INI1 respectively, to assess the effect of INI1 on HIV-1 replication. RESULTS: We found that while HIV-1 virions produced in 293T cells efficiently transduced MON and STA-WT1 cells, HIV-1 particle production was severely reduced in both of these cells. Reintroduction of INI1 into MON and STA-WT1 significantly enhanced the particle production in both cell lines. HIV-1 particles produced in MON cells were reduced for infectivity, while those produced in STA-WT1 were not. Further analysis indicated the presence of INI1 in those virions produced from STA-WT1 but not from those produced from MON cells. HIV-1 produced in MON cells were defective for synthesis of early and late reverse transcription products in the target cells. Furthermore, virions produced in MON cells were defective for exogenous reverse transcriptase activity carried out using exogenous template, primer and substrate. CONCLUSION: Our results suggest that INI1-deficient cells exhibit reduced particle production that can be partly enhanced by re-introduction of INI1. Infectivity of HIV-1 produced in some but not all INI1 defective cells, is affected and this defect may correlate to the lack of INI1 and/or some other proteins in these virions. The block in early events of virion produced from MON cells appears to be at the stage of reverse transcription. These studies suggest that presence of INI1 or some other host factor in virions and reverse transcription complexes may be important for early events of HIV-1 replication

HIV-1 Dynamics in the Host Cell: A Review of Viral- and Host- Protein Interactions and Potential Therapeutic Targets for HIV-1 Infection

HIV-1, the causative agent of AIDS, is a sophisticated retrovirus that has both evolved to invade the complex human immune system and adapted to utilize the host machinery for its own propagation. A dynamic interaction between the virus and host systems can be observed at every step of the HIV-1 lifecycle. Host factors are involved not only in mounting antiviral responses, but are also hijacked by the virus to enhance viral replication. Host factors are necessary for viral replication during entry, reverse transcription, nuclear import, integration, transcription, nuclear export, translation, assembly, and budding. Recently, a new class of host factors, called “host restriction factors,” has been identified that prevent retroviral replication in a specific host cell environment and constitute an important part of intracellular innate immunity against the virus. These restriction factors act as barriers to retroviral replication at various stages within the infected cell. Nevertheless, the HIV-1 virus has learned to subvert these antiviral responses and successfully propagate within the permissive host environment. This review article describes the identification and mechanism of action of several pro- and anti-HIV-1 host factors. It is likely that we are only beginning to get a glimpse of an ongoing complex battle between HIV-1 and the host, the understanding of which should provide valuable information for the development of novel therapeutic strategies against HIV-1.

Recruitment of a SAP18-HDAC1 Complex into HIV-1 Virions and Its Requirement for Viral Replication

HIV-1 integrase (IN) is a virally encoded protein required for integration of viral cDNA into host chromosomes. INI1/hSNF5 is a component of the SWI/SNF complex that interacts with HIV-1 IN, is selectively incorporated into HIV-1 (but not other retroviral) virions, and modulates multiple steps, including particle production and infectivity. To gain further insight into the role of INI1 in HIV-1 replication, we screened for INI1-interacting proteins using the yeast two-hybrid system. We found that SAP18 (Sin3a associated protein 18 kD), a component of the Sin3a-HDAC1 complex, directly binds to INI1 in yeast, in vitro and in vivo. Interestingly, we found that IN also binds to SAP18 in vitro and in vivo. SAP18 and components of a Sin3A-HDAC1 complex were specifically incorporated into HIV-1 (but not SIV and HTLV-1) virions in an HIV-1 IN–dependent manner. Using a fluorescence-based assay, we found that HIV-1 (but not SIV) virion preparations harbour significant deacetylase activity, indicating the specific recruitment of catalytically active HDAC into the virions. To determine the requirement of virion-associated HDAC1 to HIV-1 replication, an inactive, transdominant negative mutant of HDAC1 (HDAC1H141A) was utilized. Incorporation of HDAC1H141A decreased the virion-associated histone deacetylase activity. Furthermore, incorporation of HDAC1H141A decreased the infectivity of HIV-1 (but not SIV) virions. The block in infectivity due to virion-associated HDAC1H141A occurred specifically at the early reverse transcription stage, while entry of the virions was unaffected. RNA-interference mediated knock-down of HDAC1 in producer cells resulted in decreased virion-associated HDAC1 activity and a reduction in infectivity of these virions. These studies indicate that HIV-1 IN and INI1/hSNF5 bind SAP18 and selectively recruit components of Sin3a-HDAC1 complex into HIV-1 virions. Furthermore, HIV-1 virion-associated HDAC1 is required for efficient early post-entry events, indicating a novel role for HDAC1 during HIV-1 replication

Specificity of Interaction of INI1/hSNF5 with Retroviral Integrases and Its Functional Significance

Integrase interactor 1 (INI1)/hSNF5 is a host factor that directly interacts with human immunodeficiency virus type 1 (HIV-1) integrase and is incorporated into HIV-1 virions. Here, we show that while INI1/hSNF5 is completely absent from purified microvesicular fractions, it is specifically incorporated into HIV-1 virions with an integrase-to-INI1/hSNF5 stoichiometry of approximately 2:1 (molar ratio). In addition, we show that INI1/hSNF5 is not incorporated into related primate lentiviral and murine retroviral particles despite the abundance of the protein in producer cells. We have found that the specificity in incorporation of INI1/hSNF5 into HIV-1 virions is directly correlated with its ability to exclusively interact with HIV-1 integrase but not with other retroviral integrases. This specificity is also reflected in our finding that the transdominant mutant S6, harboring the minimal integrase interaction domain of INI1/hSNF5, blocks HIV-1 particle production but not that of the other retroviruses in 293T cells. Taken together, these results suggest that INI1/hNSF5 is a host factor restricted for HIV-1 and that S6 acts as a highly specific and potent inhibitor of HIV-1 replication