ABSTRACT
INI1/SMARCB1 binds to HIV-1 integrase (IN) through its Rpt1 domain and exhibits multifaceted role in HIV-1 replication. Determining the NMR structure of INI1-Rpt1 and modeling its interaction with the IN-C-terminal domain (IN-CTD) reveal that INI1-Rpt1/IN-CTD interface residues overlap with those required for IN/RNA interaction. Mutational analyses validate our model and indicate that the same IN residues are involved in both INI1 and RNA binding. INI1-Rpt1 and TAR RNA compete with each other for IN binding with similar IC50 values. INI1-interaction-defective IN mutant viruses are impaired for incorporation of INI1 into virions and for particle morphogenesis. Computational modeling of IN-CTD/TAR complex indicates that the TAR interface phosphates overlap with negatively charged surface residues of INI1-Rpt1 in three-dimensional space, suggesting that INI1-Rpt1 domain structurally mimics TAR. This possible mimicry between INI1-Rpt1 and TAR explains the mechanism by which INI1/SMARCB1 influences HIV-1 late events and suggests additional strategies to inhibit HIV-1 replication.
Subject(s)
HIV Integrase/metabolism , HIV-1/physiology , RNA, Viral/metabolism , SMARCB1 Protein/metabolism , Virus Replication , Genome, Viral , HIV Integrase/chemistry , HIV Integrase/genetics , Host-Pathogen Interactions , Humans , Magnetic Resonance Spectroscopy , Models, Molecular , Molecular Docking Simulation , Protein Binding , Protein Domains , RNA, Viral/chemistry , SMARCB1 Protein/chemistry , SMARCB1 Protein/genetics , Virion/growth & development , Virion/metabolismABSTRACT
Import-Karyopherin or Importin proteins bind nuclear localization signals (NLSs) to mediate the import of proteins into the cell nucleus. Karyopherin ß2 or Kapß2, also known as Transportin, is a member of this transporter family responsible for the import of numerous RNA binding proteins. Kapß2 recognizes a targeting signal termed the PY-NLS that lies within its cargos to target them through the nuclear pore complex. The recognition of PY-NLS by Kapß2 is conserved throughout eukaryotes. Kap104, the Kapß2 homolog in Saccharomyces cerevisiae, recognizes PY-NLSs in cargos Nab2, Hrp1, and Tfg2. We have determined the crystal structure of Kapß2 bound to the PY-NLS of the mRNA processing protein Nab2 at 3.05-Å resolution. A seven-residue segment of the PY-NLS of Nab2 is observed to bind Kapß2 in an extended conformation and occupies the same PY-NLS binding site observed in other Kapß2·PY-NLS structures.
