Recombinant plasmids of the human c-myc promoter-leader region and the bacterial chloramphenicol acetyltransferase (cat) gene were constructed. After transfection into different rodent and human cells, the 862-base-pair (bp) PvuII fragment carrying both c-myc promoters and 350 bp of the untranslated leader conferred 1/15 to 1/30 of the CAT activity mediated by the simian virus 40 promoter. The presence of additional sequences upstream of the PvuII fragment had an overall negative effect on c-myc promoter activity detectable by titration analysis with small amounts of transfected plasmid DNA. The analysis of numerous deletion constructs in the c-myc promoter-leader region as well as S1 mapping experiments demonstrated that the high CAT activity depended largely on the presence of the second promoter. By cotransfection of c-myc-cat constructs with plasmids carrying different parts of the c-myc promoter locus, targets for positively acting cellular factors were identified. Two positive regulatory elements were mapped within the 862-bp PvuII fragment. One was localized within the 248-bp PvuII-SmaI fragment -101 to -349 bp upstream of the first cap site and the other within the 142-pb XhoI-NaeI fragment of the first exon, comprising positions -95 to +47 relative to the second cap site. We conclude that the dual promotor of the human c-myc gene represents a strong eucaryotic promotor regulated by cooperation of positively and negatively acting cellular transcription factors.
Segments of simian virus 40 DNA spanning most of the leader sequence of the major late viral messenger RNA are dispensable.