Nucleic Acids Research, 1993, Vol. 21, No. 8 1735-1742
© 1993
MOLECULAR BIOLOGY |
High level activity of the mouse CCAAT/enhancer binding protein (C/EBP
) gene promoter involves autoregulation and several ubiquitous transcription factors
Center for Biotechnology, Karolinska Institute Novum, S141 57 Huddinge, Sweden 1Department of Medical Nutrition, Karolinska Institute Novum, S141 57 Huddinge, Sweden
* To whom correspondence should be addressed
Received February 18, 1993. Accepted February 26, 1993.
The promoter region of the mouse CCAAT-Enhancer Binding Protein (C/EBP
) gene Is capable of directing high levels of expression of reporter constructs In various cell lines, albeit even In cells that do not express their endogenous C/EBP gene. To understand the molecular mechanisms underlying this ubiquitous expression, we have characterized the promoter region of the mouse C/EBP gene by a variety of in vitro and in vivo methods. We show that three sites related in sequence to USF, BTE and C/EBP binding sites and present In promoter region –350/+ 3, are recognized by proteins from rat liver nuclear extracts. The sequence of the C/EBP promoter that Includes the USF binding site Is also capable of forming stable complexes with purified Myc + Max heterodlmers and mutation of this site drastically reduces transcription of C/EBP promoter luclferase constructs both in liver and non liver cell lines. In addition, we Identify three novel protein-binding sites two of which display similarity to NF–1 and a NFxB binding sites. The region located between nucleotides –197 and –178 forms several heat-stable complexes with liver nuclear proteins in vitro which are recognized mainly by antibodies specific for C/EBP. Furthermore, transient expression of C/EBP and to a lesser extent C/EBPß expression vectors, results in transactlvatlon of a co-transfected C/EBP promoter-luclferase reporter construct. These experiments support the notion that the C/EBP gene Is regulated by C/EBP but other C/EBP-related proteins may also be Involved.
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