Isolation of novel animal cell lines defective in glycerolipid biosynthesis reveals mutations in glucose-6-phosphate isomerase

Jorge F. Haller, Conor Smith, Dailan Liu, Hongying Zheng, Keith Tornheim, Gil Soo Han, George M. Carman, Raphael A. Zoeller

Research output: Contribution to journalArticlepeer-review

4 Scopus citations

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.

Original languageEnglish (US)
Pages (from-to)866-877
Number of pages12
JournalJournal of Biological Chemistry
Volume285
Issue number2
DOIs
StatePublished - 2010

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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