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J. Biol. Chem., Vol. 267, Issue 28, 19938-19943, Oct, 1992
MA O'Reilly, AG Geiser, SJ Kim, LA Bruggeman, AX Luu, AB Roberts and MB Sporn
Transforming growth factor TGF-beta 2 is encoded by multiple mRNA
transcripts of 5.8, 5.1, 4.0, 3.8, and 2.8 kilobase pairs (kb) that are
expressed in various human and monkey cells. Northern blot analysis using
genomic fragments of DNA was used to demonstrate that some of this size
heterogeneity is due to differences in the length of the 5'- untranslated
region. Probes that were colinear with the first 600 nucleotides of the
5'-untranslated region detected only the 5.8-, 4.0-, and 3.8-kb
transcripts. In order to identify DNA elements that regulate the
transcription of these mRNA transcripts, deletion constructs of 5'-
flanking DNA were ligated to the coding region for chloramphenicol
acetyltransferase (CAT) and analyzed for promoter activity in several cell
lines. Sequences responsible for putative enhancer and silencer regions
were identified between -778 and -40 relative to the transcription
initiation site. Addition of a cyclic AMP-responsive element/activating
transcription factor-like element at -74 resulted in a 5-10-fold increase
in CAT activity over that expressed with a construct that contained only
the TATA box. This increase in CAT activity was suppressed by the addition
of DNA sequences between -257 and -187, whereas sequences between -778 and
-257 stimulated CAT activity. Point mutations within the ATF binding site
at -74 resulted in a marked decrease in CAT expression. Cotransfection with
ATF-1 or ATF-2 expression plasmids resulted in both dose-dependent
stimulatory and inhibitory activities that were cell type-dependent. These
studies identify multiple transcription initiation sites for TGF-beta 2 and
demonstrate that transcription from one of these promoters is dependent
upon an ATF binding site located 5' of the TATA box.
Identification of an activating transcription factor (ATF) binding site in the human transforming growth factor-beta 2 promoter
Laboratory of Chemoprevention, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
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