HIV-1 integrase forms stable tetramers and associates with LEDGF/p75 protein in human cells.
We studied human immunodeficiency virus, type 1 (HIV-1) integrase (IN) complexes derived from nuclei of human cells stably expressing the viral protein from a synthetic gene. We show that in the nuclear extracts IN exists as part of a large distinct complex with an apparent Stokes radius of 61 A, which dissociates upon dilution yielding a core molecule of 41 A. We isolated the IN complexes from cells expressing FLAG-tagged IN and demonstrated that the 41 A core is a tetramer of IN, whereas 61 A molecules are composed of IN tetramers associated with a cellular protein with an apparent molecular mass of 76 kDa. This novel integrase interacting protein was found to be identical to lens epithelium-derived growth factor (LEDGF/p75), a protein implicated in regulation of gene expression and cellular stress response. HIV-1 IN and LEDGF co-localized in the nuclei of human cells stably expressing IN. Furthermore, recombinant LEDGF robustly enhanced strand transfer activity of HIV-1 IN in vitro. Our findings indicate that the minimal IN molecule in human cells is a homotetramer, suggesting that at least an octamer of IN is required to accomplish coordinated integration of both retroviral long terminal repeats and that LEDGF is a cellular factor involved in this process.
Cherepanov
Peter
P
Rega Institute for Medical Research, Katholieke Universiteit Leuven, Minderbroedersstraat 10, B-3000, Leuven, Belgium. Peter.Cherepanov@uz.kuleuven.ac.be
Maertens
Goedele
G
Proost
Paul
P
Devreese
Bart
B
Van Beeumen
Jozef
J
Engelborghs
Yves
Y
De Clercq
Erik
E
Debyser
Zeger
Z
D000595:Amino Acid Sequence; D002458:Cell Fractionation; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000378:metabolism; D019427:HIV Integrase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D020836:Protein Structure, Quaternary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013388:Succinimides / Q000378:metabolism; D016662:Virus Integration / Q000502:physiology*
D003432:Cross-Linking Reagents; D036341:Intercellular Signaling Peptides and Proteins; D046911:Macromolecular Substances; D010455:Peptides; D011994:Recombinant Proteins; D013388:Succinimides; C119380:lens epithelium-derived growth factor; C035759:3,3'-dithiobis(sulfosuccinimidyl propionate); D019427:HIV Integrase