title: Adenovirus E1A N-terminal amino acid sequence requirements for repression of transcription in vitro and in vivo correlate with those required for E1A interference with TBP-TATA complex formation.
abstract: The adenovirus (Ad) E1A 243R oncoprotein encodes an N-terminal transcription repression domain that is essential for early viral functions, cell immortalization, and cell transformation. The transcription repression function requires sequences within amino acids 1 to 30 and 48 to 60. To elucidate the roles of the TATA-binding protein (TBP), p300, and the CREB-binding protein (CBP) in the mechanism(s) of E1A repression, we have constructed 29 amino acid substitution mutants and 5 deletion mutants spanning the first 30 amino acids within the E1A 1-80 polypeptide backbone. These mutant E1A polypeptides were characterized with regard to six parameters: the ability to repress transcription in vitro and in vivo, to disrupt TBP-TATA box interaction, and to bind TBP, p300, and CBP. Two regions within E1A residues 1 to 30, amino acids 2 to 6 and amino acid 20, are critical for E1A transcription repression in vitro and in vivo and for the ability to interfere with TBP-TATA interaction. Replacement of 6Cys with Ala in the first region yields the most defective mutant. Replacement of 20Leu with Ala, but not substitutions in flanking residues, yields a substantially defective phenotype. Protein binding assays demonstrate that replacement of 6Cys with Ala yields a mutant completely defective in interaction with TBP, p300, and CBP. Our findings are consistent with a model in which the E1A repression function involves interaction of E1A with p300/CBP and interference with the formation of a TBP-TATA box complex.
authors.affiliation: Institute for Molecular Virology, Saint Louis University School of Medicine, St. Louis, Missouri 63110, USA.
authors.forename: Janice M
authors.initials: JM
authors.lastname: Boyd
authors.forename: Paul M
authors.initials: PM
authors.lastname: Loewenstein
authors.forename: Qing-quan
authors.initials: QQ
authors.lastname: Tang Qq
authors.forename: Li
authors.initials: L
authors.lastname: Yu
authors.forename: Maurice
authors.initials: M
authors.lastname: Green
mesh_terms: D017500:Adenovirus E1A Proteins / Q000235:genetics / Q000378:metabolism*; D000260:Adenoviruses, Human / Q000235:genetics / Q000378:metabolism*; D000596:Amino Acids; D001665:Binding Sites; D050882:CREB-Binding Protein; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship; D016385:TATA Box; D035181:TATA-Box Binding Protein; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
chemical_list: D017500:Adenovirus E1A Proteins; D000596:Amino Acids; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D010455:Peptides; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D035181:TATA-Box Binding Protein; D015534:Trans-Activators; D014157:Transcription Factors; D050882:CREB-Binding Protein; C494363:CREBBP protein, human