Identification of a c-myb attenuator-binding factor.
Expression of the c-myb proto-oncogene is developmentally regulated at the level of transcription elongation. In pre-B cells, complete c-myb transcripts are produced, whereas transcripts are attenuated near or within a 300-base pair (bp) interval of the first c-myb intron in mature cells. Hypothesizing that transcription attenuation results from a protein complex that physically impedes the progress of RNA polymerase II through the intron, we used electrophoretic mobility shift assays (EMSA) to search for DNA-binding activities that correlated with downregulation of c-myb transcription. We identified a stage-specific DNA binding activity, termed ABF, present in mature B cells but not in pre-B cells. ABF binds to a 15-bp DNA element located within a 300-bp BstEII-XbaI fragment. DMSO-treatment of murine erythroleukemia cells results in rapid downregulation of c-myb transcription and upregulation of ABF DNA binding activity. Thus, ABF binding activity correlates with downregulation of c-myb transcription in two systems. Preliminary biochemical characterization of ABF from mature B cells demonstrates that its primary DNA-binding component is a 64-kDa-protein. We hypothesize that this factor may represent a member of the transcriptional attenuation complex.
Perkel
Jeffrey M
JM
Abramson Family Cancer Research Institute, University of Pennsylvania, Room 456, BRB 11/111, 421 Curie Boulevard, Philadelphia, PA 19104, USA.
Simon
M Celeste
MC
Rao
Anjana
A
D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism*; D015152:Blotting, Northern; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism / Q000502:physiology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D024202:Electrophoretic Mobility Shift Assay; D018507:Gene Expression Regulation, Developmental; D015973:Gene Expression Regulation, Leukemic; D020570:Genes, myb; D007438:Introns; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D008970:Molecular Weight; D016328:NF-kappa B / Q000378:metabolism; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics; D009876:Operon; D020598:Proto-Oncogene Proteins c-myb / Q000096:biosynthesis / Q000235:genetics; D012319:RNA Polymerase II / Q000378:metabolism; D011993:Recombinant Fusion Proteins; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D004268:DNA-Binding Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009363:Neoplasm Proteins; D020598:Proto-Oncogene Proteins c-myb; D011993:Recombinant Fusion Proteins; C450200:c-myb attenuator-binding factor; D004247:DNA; D012319:RNA Polymerase II; D004121:Dimethyl Sulfoxide