Ets factors regulate the polycystic kidney disease-1 promoter
- Department of Biochemistry and Molecular Biology, and Kidney Institute, University of Kansas Medical Center, Kansas City, KS 66160 (United States)
The Ets family of transcription factors consists of a group of highly conserved sequence-specific DNA binding proteins that functionally cooperate with other transcription factors to regulate a number of diverse cellular processes including proliferation, differentiation, and apoptosis. We have analyzed a 3.3 kb 5'-upstream region of the human PKD1 promoter, using transient transfection in HEK293T cells and Drosophila SL2 cells, to demonstrate that the PKD1 promoter is a target of Ets family transcription factors. Our studies showed that PKD1 promoter-luciferase reporter gene expression is downregulated by cotransfected Fli-1 and is upregulated by cotransfected Ets-1. Using deletion constructs, we demonstrated that the sequences responding to Fli-1 and Ets-1 lie within the -200 to +33 bp proximal promoter. This region was found to contain two putative Ets response elements (EREs): an upstream (Ets-A) sequence 5'-CGGAA-3' (-181 to -185) and a downstream (Ets-B) sequence 5'-CGGAT-3' (-129 to -133). Site-directed mutagenesis indicated that both EREs are functional. A Fli-1 DNA binding domain mutant construct (W321R), which is incapable of binding DNA, was unable to inhibit basal promoter activity. In contrast, a Fli-1 DNA binding domain truncation mutant construct, which only contains the DNA binding domain and lacks the transactivation domain, was able to inhibit. These results suggest that the effect of Fli-1 is through direct binding to these EREs. Direct binding of Fli-1 and Ets-1 to the Ets-A and Ets-B sites was supported by electrophoretic mobility shift assays. Lastly, competition between Fli-1 and Ets-1 for the two EREs was demonstrated by showing that increasing amounts of Ets-1 could overcome Fli-1 repression of promoter activity. Taken together, these experiments define the proximal PKD1 promoter region as a potential target of Ets family transcription factors.
- OSTI ID:
- 20798901
- Journal Information:
- Biochemical and Biophysical Research Communications, Vol. 342, Issue 4; Other Information: DOI: 10.1016/j.bbrc.2006.02.045; PII: S0006-291X(06)00346-9; Copyright (c) 2006 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA); ISSN 0006-291X
- Country of Publication:
- United States
- Language:
- English
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