The block to transcriptional elongation within the human c-myc gene is determined in the promoter-proximal region.
A conditional block to transcriptional elongation is an important mechanism for regulating c-myc gene expression. This elongation block within the first c-myc exon was defined originally in mammalian cells by nuclear run-on transcription analyses. Subsequent oocyte injection and in vitro transcription analyses suggested that sequences near the end of the first c-myc exon are sites of attenuation and/or premature termination. We report here that the mapping of single stranded DNA in vivo with potassium permanganate (KMnO4) and nuclear run-on transcription assays reveal that polymerase is paused near position +30 relative to the major c-myc transcription initiation site. Deletion of 350 bp, including the sites of 3'-end formation and intrinsic termination defined in oocyte injection and in vitro transcription assays does not affect-the pausing of polymerase in the promoter-proximal region. In addition, sequences upstream of +47 are sufficient to confer the promoter-proximal pausing of polymerases and to generate the polarity of transcription farther downstream. Thus, the promoter-proximal pausing of RNA polymerase II complexes accounts for the block to elongation within the c-myc gene in mammalian cells. We speculate that modification of polymerase complexes at the promoter-proximal pause site may determine whether polymerases can read through intrinsic sites of termination farther downstream.
Krumm
A
A
Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, Washington 98104.
Meulia
T
T
Brunvand
M
M
Groudine
M
M
D001483:Base Sequence; D002467:Cell Nucleus / Q000502:physiology; D004273:DNA, Neoplasm / Q000187:drug effects / Q000235:genetics; D005091:Exons; D005809:Genes, Regulator; D016259:Genes, myc; D006801:Humans; D007438:Introns; D015473:Leukemia, Promyelocytic, Acute; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D011196:Potassium Permanganate / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D009838:Oligodeoxyribonucleotides; D011196:Potassium Permanganate; D012319:RNA Polymerase II