title: Artificial zinc finger fusions targeting Sp1-binding sites and the trans-activator-responsive element potently repress transcription and replication of HIV-1.
abstract: Tat activates transcription by interacting with Sp1, NF-kappaB, positive transcription elongation factor b, and trans-activator-responsive element (TAR). Tat and Sp1 play major roles in transcription by protein-protein interactions at human immunodeficiency virus, type 1 (HIV-1) long terminal repeat. Sp1 activates transcription by interacting with cyclin T1 in the absence of Tat. To disrupt the transcription activation by Tat and Sp1, we fused Sp1-inhibiting polypeptides, zinc finger polypeptide, and the TAR-binding mutant Tat (TatdMt) together. A designed or natural zinc finger and Tat mutant fusion was used to target the fusion to the key regulatory sites (GC box and TAR) on the long terminal repeat and nascent short transcripts to disrupt the molecular interaction that normally result in robust transcription. The designed zinc finger and TatdMt fusions were targeted to the TAR, and they potently repressed both transcription and replication of HIV-1. The Sp1-inhibiting POZ domain, TatdMt, and zinc fingers are key functional domains important in repression of transcription and replication. The designed artificial zinc fingers were targeted to the high affinity Sp1-binding site, and by being fused with TatdMt and POZ domain, they strongly block both Sp1-cyclin T1-dependent transcription and Tat-dependent transcription, even in the presence of excess expressed Tat.
authors.affiliation: Laboratory of Retroviruses and Gene Therapy, Indang Institute of Molecular Biology & Department of Medical Laboratory Science, Inje University, 64 Jeodong 2nd Street, Indang Building, Jung-Ku, Seoul 100-032, Korea.
authors.forename: Yeon-Soo
authors.initials: YS
authors.lastname: Kim
authors.forename: Jung-Min
authors.initials: JM
authors.lastname: Kim
authors.forename: Deug-Lim
authors.initials: DL
authors.lastname: Jung
authors.forename: Jae-Eun
authors.initials: JE
authors.lastname: Kang
authors.forename: Sukyung
authors.initials: S
authors.lastname: Lee
authors.forename: Jin Soo
authors.initials: JS
authors.lastname: Kim
authors.forename: Wongi
authors.initials: W
authors.lastname: Seol
authors.forename: Hyun-Chul
authors.initials: HC
authors.lastname: Shin
authors.forename: Heung Sun
authors.initials: HS
authors.lastname: Kwon
authors.forename: Carine
authors.initials: C
authors.lastname: Van Lint
authors.forename: Nouria
authors.initials: N
authors.lastname: Hernandez
authors.forename: Man-Wook
authors.initials: MW
authors.lastname: Hur
mesh_terms: D001665:Binding Sites; D002460:Cell Line; D002474:Cell-Free System; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D056906:Cyclin T; D016213:Cyclins / Q000737:chemistry; D017353:Gene Deletion; D015696:Gene Products, tat / Q000378:metabolism; D006678:HIV; D015497:HIV-1 / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008954:Models, Biological; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012319:RNA Polymerase II / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D015032:Zinc / Q000737:chemistry; D016335:Zinc Fingers; D054322:tat Gene Products, Human Immunodeficiency Virus
chemical_list: C503684:CCNT1 protein, human; D056906:Cyclin T; D016213:Cyclins; D015696:Gene Products, tat; D010455:Peptides; D011993:Recombinant Fusion Proteins; D016329:Sp1 Transcription Factor; D054322:tat Gene Products, Human Immunodeficiency Virus; D015500:Chloramphenicol O-Acetyltransferase; D012319:RNA Polymerase II; D015032:Zinc