Interaction between cell cycle regulator, E2F-1, and NF-kappaB mediates repression of HIV-1 gene transcription.
The NF-kappaB/Rel family of transcription factors is one of the main targets of cytokines and other agents that induce HIV-1 gene expression. Some of these extracellular stimuli arrest cells in the G1 phase of the mitotic division cycle and modulate the activity of the tumor suppressor protein Rb and its partner E2F-1. Earlier studies indicated that E2F-1, a transcription factor that stimulates expression of S-phase-specific genes, is able to repress transcription directed by the human immunodeficiency virus (HIV-1) type-1 promoter in a variety of cells, including those of glial and lymphocytic origin. Here, we demonstrate that E2F-1 may regulate the activity of the HIV-1 long terminal repeat through its ability to bind sequences in the NF-kappaB enhancer region and to interact with the NF-kappaB subunit, p50. Gel retardation and methylation interference assays show that E2F-1 is able to bind specifically to a site embedded within the two NF-kappaB elements. Gel retardation/immunoblot analysis using purified E2F-1 and p50 homodimers reveals the presence of complexes containing both proteins. Affinity chromatography and co-immunoprecipitation assays provide evidence for direct interaction of E2F-1 and p50 in the absence of their DNA target sequences. In vitro transcription assay demonstrates that E2F-1 represses NF-kappaB mediated transcription in a cell-free system. Functional studies in Jurkat T lymphocytic cells point to the importance of both the E2F and NF-kappaB binding sites in E2F-1 mediated repression of HIV-1 promoter, in vivo. The results of this study suggest that NF-kappaB activity may be regulated by its interaction with the cell cycle regulatory protein, E2F-1.
Kundu
M
M
Center for NeuroVirology and NeuroOncology, and Department of Neurology, Allegheny University of the Health Sciences, Philadelphia, Pennsylvania 19102, USA.
Guermah
M
M
Roeder
R G
RG
Amini
S
S
Khalili
K
K
D001665:Binding Sites; D002352:Carrier Proteins; D018797:Cell Cycle Proteins / Q000378:metabolism*; D004279:DNA, Viral / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D015180:Electrophoresis, Gel, Two-Dimensional; D015967:Gene Expression Regulation, Viral; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D019169:Jurkat Cells; D016297:Mutagenesis, Site-Directed; D016328:NF-kappa B / Q000378:metabolism*; D052002:NF-kappa B p50 Subunit; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004279:DNA, Viral; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D016328:NF-kappa B; D052002:NF-kappa B p50 Subunit; D011994:Recombinant Proteins; D056465:Retinoblastoma-Binding Protein 1; D050680:Transcription Factor DP1; D014157:Transcription Factors