Nucleic Acids Research, 1994, Vol. 22, No. 8 1463-1469
© 1994
MOLECULAR BIOLOGY |
Different binding specificities and transactivation of variant CRE's by CREB complexes
Department of Obstetrics and Gynecology, University of Oklahoma Health Sciences Center Oklahoma City, OK 73190, USA 1Gene Regulation Laboratory, Imperial Cancer Research Fund, Lincolns Inn Fields London WC2A 3PX, UK
*To whom correspondence should be addressed
Received December 6, 1993. Revised March 8, 1994. Accepted March 8, 1994.
The DNA binding specificities of CREB1 and CREB2 homodlmers and the CREB2/cJun heterodimer were analyzed with a CASTing technique. All but one of the selected sequences varied from the consensus CRE (TGACGTCA) by three nucleotldes or less. The profile of variations selected and the binding affinity for these sequences were unique for each CREB complex. The affinities were not effected by the palindromic nature of the sequences, but were strongly effected by flanking sequences. The strength of DNA binding in vitro correlated with the degree of transactivation observed in JEG-3 cells transfected with reporter plasmids harboring CRE variants, when hybrid CREB proteins fused to the VP16 activation domain were expressed. When native CREB proteins were expressed, the correlation was attenuated by the nature of the variant sequence. A CRE variant (TGACATCA) found in several natural promoters, exhibited the lowest basal transcription rate of the variants and a lower level of induction than expected when compared with the in vitro binding data. These results indicate that transactivation of DNA sequence elements is strongly effected by the strength of transcription factor binding, and that individual sequences can attenuate the level of induction.
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