The human polycomb group EED protein interacts with the integrase of human immunodeficiency virus type 1.
Human EED, a member of the superfamily of WD-40 repeat proteins and of the Polycomb group proteins, has been identified as a cellular partner of the human immunodeficiency virus type 1 (HIV-1) matrix (MA) protein (R. Peytavi et al., J. Biol. Chem. 274:1635-1645, 1999). In the present study, EED was found to interact with HIV-1 integrase (IN) both in vitro and in vivo in yeast. In vitro, data from mutagenesis studies, pull-down assays, and phage biopanning suggested that EED-binding site(s) are located in the C-terminal domain of IN, between residues 212 and 264. In EED, two putative discrete IN-binding sites were mapped to its N-terminal moiety, at a distance from the MA-binding site, but EED-IN interaction also required the integrity of the EED last two WD repeats. EED showed an apparent positive effect on IN-mediated DNA integration reaction in vitro, in a dose-dependent manner. In situ analysis by immunoelectron microscopy (IEM) of cellular distribution of IN and EED in HIV-1-infected cells (HeLa CD4(+) cells or MT4 lymphoid cells) showed that IN and EED colocalized in the nucleus and near nuclear pores, with maximum colocalization events occurring at 6 h postinfection (p.i.). Triple colocalizations of IN, EED, and MA were also observed in the nucleoplasm of infected cells at 6 h p.i., suggesting the ocurrence of multiprotein complexes involving these three proteins at early steps of the HIV-1 virus life cycle. Such IEM patterns were not observed with a noninfectious, envelope deletion mutant of HIV-1.
Violot
Sébastien
S
Laboratoire de Virologie and Pathogénèse Virale, Faculté de Médecine RTH Laennec, CNRS UMR-5537, France.
Hong
Saw See
SS
Rakotobe
Dina
D
Petit
Caroline
C
Gay
Bernard
B
Moreau
Karen
K
Billaud
Geneviève
G
Priet
Stéphane
S
Sire
Joséphine
J
Schwartz
Olivier
O
Mouscadet
Jean-François
JF
Boulanger
Pierre
P
D000595:Amino Acid Sequence; D002460:Cell Line; D019427:HIV Integrase / Q000378:metabolism*; D015497:HIV-1 / Q000201:enzymology / Q000378:metabolism / Q000502:physiology; D006801:Humans; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D063151:Polycomb Repressive Complex 2; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques
C491488:EED protein, human; D011994:Recombinant Proteins; D012097:Repressor Proteins; D063151:Polycomb Repressive Complex 2; D019427:HIV Integrase