Insulin directly down-regulates the gene expression of the rat CYP2E1 by altering its mRNA stability (De Waziers, I., Garlatti, M., Bouguet, J., Beaune, P. H., and Barouki, R. (1995) Mol. Pharmacol. 47, 474 -479). Because the regulation of CYP mRNA stability was poorly understood, the molecular mechanisms involved in this regulation in the rat hepatoma H4IIEC3 cell line were studied. By using RNase T1 protection methods, the formation of a major CYP2E1 RNA-protein complex was observed. By competition experiments, the binding site of this complex was located on a 16-nucleotide sequence in the 5-proximal region of the CYP2E1-coding sequence. Insulin did not modify the binding pattern of proteins to this sequence. and transfections of expression vectors or antisense oligonucleotides were undertaken to demonstrate the actual functionality of the 16-mer sequence. The insertion of this sequence in a luciferase gene was sufficient to render the chimeric mRNA sensitive to insulin. Furthermore, transfection of H4IIEC3 cells with antisense oligonucleotide complementary to this sequence blocked the insulin effect on the CYP2E1 mRNA expression, i.e. its rapid degradation. All these results demonstrate that this 16-nucleotide sequence is implicated in the CYP2E1 post-transcriptional regulation by insulin.

Background:In plants, a key regulatory step in chlorophyll biosynthesis is catalyzed by two light-dependent isozymes. Results: The two isozymes operate via the same reaction mechanism but differ in their catalytic efficiencies. Conclusion: Different substrate affinities and conformational flexibilities modulate the catalytic reaction. Significance: Detailed understanding of light-driven reactions in nature has a big impact on the development of artificial energy conversion systems.

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