title: The HIV-1 integrase mutant R263A/K264A is 2-fold defective for TRN-SR2 binding and viral nuclear import.
abstract: Transportin-SR2 (Tnpo3, TRN-SR2), a human karyopherin encoded by the TNPO3 gene, has been identified as a cellular cofactor of HIV-1 replication, specifically interacting with HIV-1 integrase (IN). Whether this interaction mediates the nuclear import of HIV remains controversial. We previously characterized the TRN-SR2 binding interface in IN and introduced mutations at these positions to corroborate the biological relevance of the interaction. The pleiotropic nature of IN mutations complicated the interpretation. Indeed, all previously tested IN interaction mutants also affected RT. Here we report on a virus with a pair of IN mutations, IN(R263A/K264A), that significantly reduce interaction with TRN-SR2. The virus retains wild-type reverse transcription activity but displays a block in nuclear import and integration, as measured by quantitative PCR. The defect in integration of this mutant resulted in a smaller increase in the number of two-long terminal repeat circles than for virus specifically blocked at integration by raltegravir or catalytic site mutations (IN(D64N/D116N/E152Q)). Finally, using an eGFP-IN-labeled HIV fluorescence-based import assay, the defect in nuclear import was corroborated. These data altogether underscore the importance of the HIV-IN TRN-SR2 protein-protein interaction for HIV nuclear import and validate the IN/TRN-SR2 interaction interface as a promising target for future antiviral therapy.
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium.
authors: Stéphanie
authors: S
authors: De Houwer
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium.
authors: Jonas
authors: J
authors: Demeulemeester
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium.
authors: Wannes
authors: W
authors: Thys
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium.
authors: Susana
authors: S
authors: Rocha
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium.
authors: Lieve
authors: L
authors: Dirix
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium.
authors: Rik
authors: R
authors: Gijsbers
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium.
authors: Frauke
authors: F
authors: Christ
authors: From the Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, Katholieke Universiteit Leuven, 3000, Leuven, Belgium zeger.debyser@med.kuleuven.be.
authors: Zeger
authors: Z
authors: Debyser
mesh_terms: D021581:Active Transport, Cell Nucleus; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D015153:Blotting, Western; D002467:Cell Nucleus / Q000821:virology; D019427:HIV Integrase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D014779:Virus Replication / Q000235:genetics; D028961:beta Karyopherins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D050505:Mutant Proteins; D028961:beta Karyopherins; C424165:transportin SR2; D019427:HIV Integrase; C514801:p31 integrase protein, Human immunodeficiency virus 1
keywords: HIV Integrase; HIV-1 Protease; Human Immunodeficiency Virus (HIV); Nuclear Import; Nuclear Transport; TNPO3; TRN-SR2; Transportin-3; Viral Replication