Glutamine plays an important biosynthetic role in all mammalian cells. Glutamine is used by the liver during diabetes, the kidney during metabolic acidosis and the mammary gland during lactation, it is also a major respiratory fuel for many rapidly dividing cells including cancer cells. The major enzymes of glutamine catabolism in mammalian tissues are two isozymes of phosphate activated glutaminase. The liver type is found exclusively in adult liver while the other enzyme (kidney-type) is found in all other glutamine utilizing tissues, including fetal liver and hepatoma cells.
The aim of the work described is to characterize and understand the role and regulation of phosphate activated glutaminase in rat liver. This work stems from the observation that hepatic glutaminase activity is increased 4-fold during streptozotocin diabetes. The enzyme has been purified, specific antibodies raised against it and used to identify a putative cDNA. The identify of the cDNA will be confirmed by comparison of the DNA and amino acid sequences, hybrid selection of mRNA and analysis of fusion peptide. The cDNA will be used to determine the relative abundance of mRNA and the rate of transcription hepatic glutaminase gene in liver from control and diabetic rats. Extracellular stimuli involved in the regulation of hepatic glutaminase gene expression will be identified using isolated hepatocytes and hepatoma cells. The cDNA will be used to identify genomic sequences encoding liver-type glutaminase and other glutamine utilizing enzymes. The distribution of liver- and kidney-type glutaminase (activity, protein and mRNA) will be determined in different hepatoma cells and in rat liver at different stages of development. Chromatin structure will be analyzed in cells showing differential expression of the two genes. These systems offer a unique model for the study of tissue-specific, hormonal and developmental regulation of glutaminase gene expression. The results will provide valuable insights into why there are two enzymes and what their functions are in the different cell types.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK037301-03
Application #
3236142
Study Section
Biochemistry Study Section (BIO)
Project Start
1989-07-01
Project End
1992-12-31
Budget Start
1991-05-15
Budget End
1991-12-31
Support Year
3
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Rutgers University
Department
Type
Schools of Earth Sciences/Natur
DUNS #
038633251
City
New Brunswick
State
NJ
Country
United States
Zip Code
08901
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Chung-Bok, M I; Vincent, N; Jhala, U et al. (1997) Rat hepatic glutaminase: identification of the full coding sequence and characterization of a functional promoter. Biochem J 324 ( Pt 1):193-200
Chung-Bok, M I; Watford, M (1997) Characterization of the hepatic glutaminase promoter. Contrib Nephrol 121:43-7
Curthoys, N P; Watford, M (1995) Regulation of glutaminase activity and glutamine metabolism. Annu Rev Nutr 15:133-59
Watford, M; Vincent, N; Zhan, Z et al. (1994) Transcriptional control of rat hepatic glutaminase expression by dietary protein level and starvation. J Nutr 124:493-9
Zhan, Z; Vincent, N C; Watford, M (1994) Transcriptional regulation of the hepatic glutaminase gene in the streptozotocin-diabetic rat. Int J Biochem 26:263-8
Watford, M (1993) Hepatic glutaminase expression: relationship to kidney-type glutaminase and to the urea cycle. FASEB J 7:1468-74
Watford, M (1991) Regulation of expression of the genes for glutaminase and glutamine synthetase in the acidotic rat. Contrib Nephrol 92:211-7
Dodgson, S J; Watford, M (1990) Differential regulation of hepatic carbonic anhydrase isozymes in the streptozotocin-diabetic rat. Arch Biochem Biophys 277:410-4
Smith, E M; Watford, M (1990) Molecular cloning of a cDNA for rat hepatic glutaminase. Sequence similarity to kidney-type glutaminase. J Biol Chem 265:10631-6

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