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A sequence element in the GLUT4 gene that mediates repression by insulin
1Department of Pediatrics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21287-3311, USA. dcooke@welchlink.welch.jhu.edu
The Journal of Biological Chemistry
|April 16, 1998
Summary
Insulin treatment reduces glucose transporter type 4 (GLUT4) expression in fat cells. A specific DNA region in the GLUT4 gene promoter is identified as crucial for this insulin-induced gene repression.
Area of Science:
- Molecular Biology
- Cell Biology
- Metabolic Regulation
Background:
- Insulin is a key regulator of glucose metabolism.
- GLUT4 (glucose transporter type 4) is essential for insulin-stimulated glucose uptake in adipocytes.
- Understanding GLUT4 gene regulation by insulin is critical for metabolic research.
Purpose of the Study:
- To investigate the molecular mechanisms by which insulin down-regulates GLUT4 gene expression in 3T3-L1 adipocytes.
- To identify specific DNA sequences and protein factors involved in insulin-mediated repression of the GLUT4 promoter.
Main Methods:
- Utilized promoter-reporter gene constructs with varying lengths of the GLUT4 5'-flanking region.
- Employed DNase I footprinting assays to identify DNA-protein interactions.
- Performed gel mobility shift assays and Southwestern analysis to characterize binding proteins.
- Generated deletion mutants to confirm the functional role of identified regulatory elements.
Main Results:
- Insulin significantly down-regulated reporter gene expression driven by the full-length GLUT4 promoter (2900 and 785 bp).
- A specific region (-707 to -681 bp) of the GLUT4 5'-flanking region was protected from DNase I digestion, indicating protein binding.
- Gel mobility shift and Southwestern analyses identified specific protein-DNA interactions within this region.
- Deletion of the -706 to -676 bp sequence abolished insulin-induced repression, confirming its necessity.
Conclusions:
- A novel insulin-responsive element in the GLUT4 promoter, located between -706 and -676 bp, mediates insulin-induced gene repression.
- This element and its associated binding proteins represent a distinct mechanism of GLUT4 gene regulation by insulin, independent of known insulin response elements.