Coiled-coil structure-mediated dimerization of template activating factor-I is critical for its chromatin remodeling activity.
Template activating factor-I (TAF-I)alpha and TAF-Ibeta have been identified as the host factors that activate DNA replication of the adenovirus genome complexed with viral basic core proteins (Ad core). TAF-I causes a structural change of the Ad core, thereby stimulating not only replication but also transcription from the Ad core DNA in vitro. TAF-I also activates transcription from the reconstituted chromatin consisting of DNA fragments and purified histones through chromatin remodeling. Although the carboxyl-terminal region, which is highly rich in acidic amino acids, is essential for the TAF-I activity, it remains unclear how other parts are involved in its activity. The native TAF-I isolated from HeLa cells exists as either hetero- or homo-oligomer. Here, we have demonstrated by cross-linking assays that most of TAF-I exists as a dimer. Analyses using deletion mutant TAF-I proteins revealed that the amino-terminal region of TAF-I common to both alpha and beta is essential for dimerization. This region is predicted to form a coiled-coil structure. Indeed, mutations disrupting this putative structure abolished the dimerization capability and reduced the TAF-I activity in the Ad core DNA replication assay. Furthermore, we found that TAF-I mutants lacking the acidic tail act in a dominant-negative manner in this assay. These observations strongly suggest that the dimerization of TAF-I is important for its activity.
Miyaji-Yamaguchi
M
M
Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, 226-8501, Japan.
Okuwaki
M
M
Nagata
K
K
D000256:Adenoviridae / Q000235:genetics; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D002843:Chromatin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone; D003432:Cross-Linking Reagents; D004261:DNA Replication; D004268:DNA-Binding Proteins; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D019281:Dimerization; D005799:Genes, Dominant / Q000235:genetics; D016679:Genome, Viral; D006367:HeLa Cells; D056488:Histone Chaperones; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D017384:Sequence Deletion; D014157:Transcription Factors; D014758:Viral Core Proteins / Q000378:metabolism
D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D056488:Histone Chaperones; D009687:Nuclear Proteins; C513473:SET protein, human; D014157:Transcription Factors; D014758:Viral Core Proteins; C025630:CAGCTG-specific type II deoxyribonucleases; D015252:Deoxyribonucleases, Type II Site-Specific