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Isolation of novel animal cell lines defective in glycerolipid biosynthesis reveals mutations in glucose-6-phosphate isomerase.
- Source :
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The Journal of biological chemistry [J Biol Chem] 2010 Jan 08; Vol. 285 (2), pp. 866-77. Date of Electronic Publication: 2009 Nov 10. - Publication Year :
- 2010
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Abstract
- Glycerolipids are structural components for membranes and serve in energy storage. We describe here the use of a photodynamic selection technique to generate a population of Chinese hamster ovary cells that display a global deficiency in glycerolipid biosynthesis. One isolate from this population, GroD1, displayed a profound reduction in the synthesis of phosphatidylcholine, phosphatidylethanolamine, and triglycerides but presented high levels of phosphatidic acid and normal levels of phosphatidylinositol synthesis. This was accompanied by a reduction in phosphatidate phosphatase 1 (PAP1) activity. Expression cloning and sequencing of the cDNA obtained from GroD1 revealed a point mutation, Gly-189 --> Glu, in glucose-6-phosphate isomerase (GPI), a glycolytic enzyme involved in an inherited disorder that results in anemia and neuromuscular symptoms in humans. GPI activity was reduced by 87% in GroD1. No significant differences were found in DNA synthesis, protein synthesis, and ATP levels, whereas glycerol 3-phosphate levels were increased in the mutant. Expression of wild-type hamster GPI restored GPI activity, glycerolipid biosynthesis, and PAP1 activity in GroD1. Two additional, independently isolated GPI-deficient mutants displayed similar phenotypes with respect to PAP1 activity and glycerolipid biosynthesis. These findings uncover a novel relationship between GPI, involved in carbohydrate metabolism, and PAP1, a lipogenic enzyme. These results may also help to explain neuromuscular symptoms associated with inherited GPI deficiency.
- Subjects :
- Anemia enzymology
Anemia genetics
Animals
Antigens, Neoplasm genetics
Antigens, Neoplasm metabolism
Biomarkers, Tumor genetics
Biomarkers, Tumor metabolism
CHO Cells
Cricetinae
Cricetulus
Glucose-6-Phosphate Isomerase genetics
Humans
Lectins, C-Type genetics
Lectins, C-Type metabolism
Metabolism, Inborn Errors enzymology
Metabolism, Inborn Errors genetics
Neuromuscular Diseases enzymology
Neuromuscular Diseases genetics
Pancreatitis-Associated Proteins
Phospholipids genetics
Point Mutation
Triglycerides genetics
Glucose-6-Phosphate Isomerase metabolism
Phospholipids biosynthesis
Triglycerides biosynthesis
Subjects
Details
- Language :
- English
- ISSN :
- 1083-351X
- Volume :
- 285
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- The Journal of biological chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 19903819
- Full Text :
- https://doi.org/10.1074/jbc.M109.068213