Protein interactions targeting the latency-associated nuclear antigen of Kaposi's sarcoma-associated herpesvirus to cell chromosomes.
Maintenance of Kaposi's sarcoma-associated herpesvirus (KSHV) latent infection depends on the viral episomes in the nucleus being distributed to daughter cells following cell division. The latency-associated nuclear antigen (LANA) is constitutively expressed in all KSHV-infected cells. LANA binds sequences in the terminal repeat regions of the KSHV genome and tethers the viral episomes to chromosomes. To better understand the mechanism of chromosomal tethering, we performed glutathione S-transferase (GST) affinity and yeast two-hybrid assays to identify LANA-interacting proteins with known chromosomal association. Two of the interactors were the methyl CpG binding protein MeCP2 and the 43-kDa protein DEK. The interactions of MeCP2 and DEK with LANA were confirmed by coimmunoprecipitation. The MeCP2-interacting domain was mapped to the previously described chromatin binding site in the N terminus of LANA, while the DEK-interacting domain mapped to LANA amino acids 986 to 1043 in the C terminus. LANA was unable to associate with mouse chromosomes in chromosome spreads of transfected NIH 3T3 cells. However, LANA was capable of targeting to mouse chromosomes in the presence of human MeCP2 or DEK. The data indicate that LANA is tethered to chromosomes through two independent chromatin binding domains that interact with different protein partners.
Krithivas
Anita
A
Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland 21231, USA.
Fujimuro
Masahiro
M
Weidner
Magdalena
M
Young
David B
DB
Hayward
S Diane
SD
D000595:Amino Acid Sequence; D000818:Animals; D000956:Antigens, Viral; D002460:Cell Line; D002868:Chromosomal Proteins, Non-Histone; D002875:Chromosomes / Q000378:metabolism / Q000821:virology*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D019288:Herpesvirus 8, Human / Q000235:genetics* / Q000502:physiology; D006801:Humans; D051783:Methyl-CpG-Binding Protein 2; D051379:Mice; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015513:Oncogene Proteins / Q000378:metabolism*; D000075223:Poly-ADP-Ribose Binding Proteins; D011233:Precipitin Tests; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins; D020798:Two-Hybrid System Techniques
D000956:Antigens, Viral; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; C084738:DEK protein, human; C497392:MECP2 protein, human; C497393:Mecp2 protein, mouse; D051783:Methyl-CpG-Binding Protein 2; D009687:Nuclear Proteins; D015513:Oncogene Proteins; D000075223:Poly-ADP-Ribose Binding Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C109540:latency-associated nuclear antigen; D005982:Glutathione Transferase