Angiotensin II activates the GFAT promoter in mesangial cells.
Expression of glutamine:fructose-6-phosphate amidotransferase (GFAT), the rate-limiting enzyme for glucose entry into the hexosamine pathway, is transcriptionally regulated. Immunohistochemical studies of human kidney biopsies demonstrate increased GFAT expression in diabetic glomeruli, but the mechanism responsible for this overexpression is unknown. Given the role of ANG II in diabetic kidney disease, we chose to study the effect of ANG II on GFAT promoter activity in mesangial cells (MC). Exposure of MC to ANG II (10(-7) M) increased GFAT promoter activity (2.5-fold), mRNA (3-fold), and protein (1.6-fold). ANG II-mediated GFAT promoter activation was inhibited by the ANG II type I receptor antagonist candesartan (10(-8) M) but was unaffected by the ANG II type II receptor antagonist PD-123319 (10(-8) M). The intracellular calcium chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (10(-6) M), protein kinase C (PKC) inhibitors bisindoylmaleimide-4 (10(-6) M) and calphostin C (10(-7) M), protein tyrosine kinase (PTK) inhibitor genistein (10(-4) M), Src family kinase inhibitor PP2 (2.5 x 10(-7) M), p42/44 mitogen-activated protein kinase (MAPK) inhibitor PD-98059 (10(-5) M), and the epidermal growth factor (EGF) inhibitor AG-1478 all attenuated GFAT promoter activation by ANG II. We conclude that the GFAT promoter is activated by ANG II via the AT1 receptor. Promoter activation is calcium dependent and PKC dependent but also involves PTK signaling pathways including Src, the EGF receptor, and p42/44 MAPK.
James
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LR
Department of Medicine, University of Toronto, Toronto, Ontario, Canada. leighton_james@med.unc.edu
Ingram
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Thai
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Cai
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Scholey
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D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D004789:Enzyme Activation; D005920:Glomerular Mesangium / Q000187:drug effects* / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing) / Q000235:genetics / Q000378:metabolism*; D008297:Male; D011401:Promoter Regions, Genetic; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000235:genetics; D053773:Transforming Growth Factor beta1; D019010:Vascular Cell Adhesion Molecule-1 / Q000235:genetics
D012333:RNA, Messenger; C508960:TGFB1 protein, human; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D019010:Vascular Cell Adhesion Molecule-1; D000804:Angiotensin II; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing); D011505:Protein-Tyrosine Kinases; D011493:Protein Kinase C; D005947:Glucose; D002118:Calcium