Abstract | AIMS/HYPOTHESIS: METHODS: Mutated human GFAT expression vectors were prepared by PCR-site directed mutagenesis. Wild-type and mutated vectors were transfected into human embryonic kidney 293 cells and mesangial cells and GFAT enzyme activity was assessed by formation of glucosamine-6-phosphate. Production of TGF-beta1 and fibronectin protein was examined by ELISA. RESULTS: Mutation of histidine 577 or lysine 676 to alanine led to a complete loss of GFAT enzyme activity. An increased concentration of wild-type GFAT in mesangial cells enhanced both TGF-beta1 and fibronectin production 1.5-fold, while mesangial cells transfected with the mutated GFAT constructs showed no effect. CONCLUSION/INTERPRETATION: The data indicate that the hexosamine pathway-mediated induction of TGF-beta1 synthesis in mesangial cells is dependent on GFAT enzyme activity. Our results add to previous observations showing that the hexosamine pathway could increase the transcriptional activity of nuclear proteins leading to enhanced cytokine synthesis.
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Authors | C Weigert, U Friess, K Brodbeck, H U Häring, E D Schleicher |
Journal | Diabetologia
(Diabetologia)
Vol. 46
Issue 6
Pg. 852-5
(Jun 2003)
ISSN: 0012-186X [Print] Germany |
PMID | 12802498
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
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Chemical References |
- Fibronectins
- Recombinant Proteins
- VEGFB protein, human
- Vascular Endothelial Growth Factor B
- Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing)
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Topics |
- Amino Acid Substitution
- Animals
- Cell Line
- Fibronectins
(genetics)
- Genetic Vectors
- Glomerular Mesangium
(physiology)
- Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing)
(genetics, metabolism)
- Humans
- Mice
- Mutagenesis, Site-Directed
- Polymerase Chain Reaction
- Recombinant Proteins
(metabolism)
- Transfection
- Vascular Endothelial Growth Factor B
(genetics)
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