Subcellular localization of feline immunodeficiency virus integrase and mapping of its karyophilic determinant.
Feline immunodeficiency virus (FIV), like other members of the lentivirus subfamily, such as human immunodeficiency virus type 1 (HIV-1), can infect nondividing and terminally differentiated cells. The transport of the preintegration complex into the nucleus is cell cycle-independent, but the mechanism is not well understood. Integrase is a key component of the complex and has been suggested to play a role in nuclear import during HIV-1 replication. To determine its karyophilic property, FIV integrase fused with glutathione S-transferase and enhanced green fluorescent protein was expressed in various feline and human cells and the subcellular localization was visualized by fluorescence microscopy. Wild-type FIV integrase was karyophilic in all cell lines tested and capable of targeting the fusion protein to the nuclei of transfected cells. Analysis of deletion and point mutation variants of FIV integrase failed to reveal any canonical nuclear localization signal, and the karyophilic determinant was mapped to the highly conserved N-terminal zinc-binding HHCC motif. A region near the C-terminal domain enriched with basic amino acid residues also affected the nuclear import of integrase. However, the role of this region is only modulatory in comparison to that of the zinc-binding domain. The N-terminal zinc-binding domain does not bind DNA and instead is essential in integrase multimerization. We therefore postulate that the karyophilic property of FIV integrase requires subunit multimerization promoted by the HHCC motif. Alternatively, the HHCC motif may directly promote interaction between FIV integrase and cellular proteins involved in nuclear import.
Woodward
Cora L
CL
Department of Molecular and Medical Pharmacology, Molecular Biology Institute and AIDS Institute, School of Medicine, University of California, Los Angeles, California 90095, USA.
Wang
Yao
Y
Dixon
Wendy J
WJ
Htun
Han
H
Chow
Samson A
SA
D000818:Animals; D002415:Cats; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D016089:Immunodeficiency Virus, Feline / Q000201:enzymology / Q000472:pathogenicity*; D019426:Integrases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions / Q000201:enzymology*; D014162:Transfection; D016662:Virus Integration
D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D005982:Glutathione Transferase; D019426:Integrases