Nuclear protein binding to the 5' enhancer region of the intracisternal A particle long terminal repeat.
The interactions of nuclear proteins from embryonal carcinoma cells (PCC3) with the long terminal repeats (LTRs) of murine intracisternal A particle (IAP) genes were studied. Two protein-DNA complexes were detected between PCC3 nuclear extract and IAP LTRs in a gel mobility shift assay. An additional complex was observed when enriched fractions from a heparin-agarose column were used as the source of proteins. Two regions within the LTR of IAP 81 were identified as the sites of protein interaction by DNase I protection. One region encompasses 43 nucleotides within the U3 region at the 5' end of LTR 81. The other covers a 78 base pair region lying within 100 nucleotides upstream from the transcription initiation site. Studies using constructs containing intact or deleted versions of the LTR fused to the bacterial chloramphenicol acetyltransferase gene indicated that the absence of the 5' 47 base pairs reduced the level of chloramphenicol acetyltransferase transcription to 20% of that driven by the entire LTR. Southwestern analysis of PCC3 nuclear extracts and column fractions revealed that a 28,000- and a 46,000-dalton protein were the major species that interact with the 5' end of IAP LTR 81.
Zierler
M
M
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218.
Christy
R J
RJ
Huang
R C
RC
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism*; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D002846:Chromatography, Affinity; D002848:Chromatography, DEAE-Cellulose; D004273:DNA, Neoplasm / Q000235:genetics / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D003850:Deoxyribonuclease I; D004742:Enhancer Elements, Genetic; D007426:Genes, Intracisternal A-Particle; D015151:Immunoblotting; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins / Q000378:metabolism; D012091:Repetitive Sequences, Nucleic Acid; D013724:Teratoma; D014162:Transfection; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins; D015500:Chloramphenicol O-Acetyltransferase; D003850:Deoxyribonuclease I