Identification of a downstream sequence and binding protein that regulate adenovirus major late promoter transcription in vitro.
Gel electrophoresis mobility shift and DNase I footprint assays detect a cellular nuclear protein in extracts made from uninfected human cells which binds to a downstream promoter sequence (DPS) in the human adenovirus 2 major late promoter. By DNase I footprint and mutation analyses, we have determined that this new regulatory element extends from positions +146 to +165 (relative to the cap site at position +1). We show by UV cross-linking that a 40-kDa polypeptide specifically binds to this region. Mutations within the DPS which decrease protein binding by 80-90% also cause a 2.5-3-fold decrease in in vitro major late promoter transcription efficiency. Alteration of the template in the 5'-flanking region of the DPS does not affect nuclear protein binding or transcription efficiency. Interestingly, a T----G transversion at position +160 which increases protein binding also impairs promoter activity.
Cohen
R B
RB
Section on RNA and Protein Biosynthesis, National Heart, Lung, and Blood Institute, Bethesda, Maryland 20892.
Yang
L
L
Thompson
J A
JA
Safer
B
B
D000260:Adenoviruses, Human / Q000235:genetics*; D001483:Base Sequence; D002472:Cell Transformation, Viral; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D005786:Gene Expression Regulation; D005796:Genes; D005814:Genes, Viral; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic; D014764:Viral Proteins
D004268:DNA-Binding Proteins; C051096:MLTF protein, adenovirus; D009687:Nuclear Proteins; D014157:Transcription Factors; D014764:Viral Proteins