Expression of GFAT1 and OGT in podocytes: transport of glucosamine and the implications for glucose uptake into these cells.
Glutamine:fructose-6-phosphate amidotransferase (GFAT) and N-acetylglucosaminyltransferase (OGT) participate in glucosamine (GlcN) production and its utilization in O-glycosylation, one of key post-translational modifications of nuclear and cytoplasmic proteins. For this purpose, cells require a high rate of intracellular production of GlcN and/or significant GlcN delivery. We studied the expression of GFAT1 and OGT and measured uptake of glucose and GlcN in cultured rat podocytes, the main cellular component of glomerular filtration barrier. RT-PCR revealed the presence of both GFAT1 and OGT mRNA. Immunofluorescence of GFAT1 has shown staining signal diffused within the cytoplasm of the cell body and processes. However, OGT was distinctly visible around the nucleus and, in diffuse form, within the cytoplasm of cell bodies and processes. Glucose was transported (1.3 +/- 0.2 nmol/min/mg protein) mainly by facilitative transporter systems whilst GlcN uptake (1.1 +/- 0.2 nmol/min/mg protein) in a significant part, involved a sodium-dependent transporter. There was interplay between glucose and GlcN uptake. In the presence of GlcN (50 microM), the rate of glucose uptake decreased by about 50%. The rate of GlcN uptake decreased by 28% in the presence of 5.6 mM glucose. Our results suggest that cultured podocytes possess limited ability to synthesize GlcN internally and therefore may need to receive GlcN from the extracellular environment.
Rogacka
Dorota
D
Department of Clinical Immunology and Transplantology, Medical University of Gdańsk, Gdańsk, Poland. rogacka@gumed.edu.pl
Piwkowska
Agnieszka
A
Jankowski
Maciej
M
Kocbuch
Katarzyna
K
Dominiczak
Marek H
MH
Stepiński
Jan K
JK
Angielski
Stefan
S
D000818:Animals; D001692:Biological Transport; D005260:Female; D005786:Gene Expression Regulation; D005944:Glucosamine / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism*; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing) / Q000235:genetics / Q000378:metabolism*; D017351:N-Acetylglucosaminyltransferases / Q000235:genetics / Q000378:metabolism*; D050199:Podocytes / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar
D012333:RNA, Messenger; D017351:N-Acetylglucosaminyltransferases; C037649:N-acetyllactosaminide beta-1,6-N-acetylglucosaminyltransferase; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing); D005947:Glucose; D005944:Glucosamine