Nucleic Acids Research, 1989, Vol. 17, No. 1 253-270
© 1989
MOLECULAR BIOLOGY |
5' flanking and first intron sequences of the human ß-actin gene required for efficient promoter activity
Division of Biological Research, Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto 500 Sherbourne Street, Toronto, Ontario M4X 1K9, Canada 1Department of Bacteriology, University of Tokyo 7-3-1 Hongo, Bunkyo-ku, Tokyo 113, Japan
*To whom correspondence should be addressed
Received September 1, 1988. Revised December 1, 1988. Accepted December 1, 1988.
We have identified a CCAAT box element that is required for the efficient transcription of the human ß-actin gene. Both in vivo transient transfection assays in cultured HeLa cells and in vitro run-off transcription assays in HeLa whole cell extracts demonstrated the requirement of this element for efficient promoter activity. A gel mobility shift assay revealed a Hela nuclear factor that specifically interacted with the ß-actin CCAAT element in vitro; mutation of the first three base pairs of the CCAAT pentanucleotide abolished binding of this factor. Competition gel shift experiments revealed that three sequence elements located within the ß-actin promoter, each containing a CC(A/T)6GG motif similar to that contained within the c-fos serum response element, were able to bind a different nuclear factor, serum response factor (SRF). One of these CC(A/T)6GG motifs is contained within a first intron fragment that enhanced transcription from a heterologous promoter in vivo.
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