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Novel sequence determinants in peroxisome proliferator signaling
C N Palmer1, M H Hsu, H J Griffin
1Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, California 92037, USA.
The Journal of Biological Chemistry
|July 7, 1995
Summary
Peroxisome proliferators activate CYP4A6 gene transcription via PPAR alpha and RXR alpha binding to enhancer elements. Specific sequences flanking the DR1 motif are crucial for this interaction, not just the DR1 alone.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- The cytochrome P-450 4A6 (CYP4A6) gene is activated by peroxisome proliferators.
- This activation involves enhancer elements (Z, X, -27) and nuclear receptor binding.
- Peroxisome proliferator response elements (PPREs) typically contain direct repeat sequences (DR1).
Purpose of the Study:
- To investigate the role of the DR1 motif and flanking sequences in the Z enhancer element for CYP4A6 gene activation.
- To determine the binding requirements of peroxisome proliferator-activated receptor alpha (PPAR alpha) and retinoid X receptor alpha (RXR alpha) heterodimers.
Main Methods:
- Mutational analysis of the Z enhancer element.
- Gel retardation assays to assess binding of nuclear receptors (PPAR alpha, RXR alpha, ARP-1) to modified oligonucleotides.
Main Results:
- The DR1 motif is necessary but not sufficient for transcriptional activation by peroxisome proliferators.
- Sequences immediately upstream (5') of the DR1 motif are critical for PPAR alpha.RXR alpha heterodimer binding.
- Mutations in the 5' flanking region reduced PPAR alpha.RXR alpha binding but not homodimer binding of RXR alpha or ARP-1.
Conclusions:
- The DR1 motif alone does not define a functional PPRE.
- Specific upstream sequences are essential for high-affinity binding of PPAR alpha.RXR alpha heterodimers to the CYP4A6 Z element.
- These upstream sequences confer specificity for PPAR alpha.RXR alpha binding over other nuclear receptors like ARP-1.