PCYT2 synthesizes CDP-glycerol in mammals and reduced PCYT2 enhances the expression of functionally glycosylated α-dystroglycan.

Abstract

α-Dystroglycan (α-DG) is a highly glycosylated cell-surface protein. Defective O-mannosyl glycan on α-DG is associated with muscular dystrophies and cancer. In the biosynthetic pathway of the O-mannosyl glycan, fukutin (FKTN) and fukutin-related protein (FKRP) transfer ribitol phosphate (RboP). Previously, we reported that FKTN and FKRP can also transfer glycerol phosphate (GroP) from CDP-glycerol (CDP-Gro) and showed the inhibitory effects of CDP-Gro on functional glycan synthesis by preventing glycan elongation in vitro. However, whether mammalian cells have CDP-Gro or associated synthetic machinery has not been elucidated. Therefore, the function of CDP-Gro in mammals is largely unknown. Here, we reveal that cultured human cells and mouse tissues contain CDP-Gro using liquid chromatography tandem-mass spectrometry (LC-MS/MS). By performing the enzyme activity assay of candidate recombinant proteins, we found that ethanolamine-phosphate cytidylyltransferase (PCYT2), the key enzyme in de novo phosphatidylethanolamine biosynthesis, has CDP-Gro synthetic activity from glycerol-3-phosphate (Gro3P) and CTP. In addition, knockdown of PCYT2 dramatically reduced cellular CDP-Gro. These results indicate that PCYT2 is a CDP-Gro synthase in mammals. Furthermore, we found that the expression of functionally glycosylated α-DG is increased by reducing PCYT2 expression. Our results suggest an important role for CDP-Gro in the regulation of α-DG function in mammals.

• Authors: Rieko Imae, Hiroshi Manya, Hiroki Tsumoto, Yuri Miura, Tamao Endo
• Journal: Journal of biochemistry (J Biochem) Vol. 170 Issue 2 Pg. 183-194 (Oct 11 2021) ISSN: 1756-2651 [Electronic] England
• PMID: 34255834 (Publication Type: Journal Article)
• Copyright: © The Author(s) 2021. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

Chemical References

• FKTN protein, human
• Membrane Proteins
• Nucleoside Diphosphate Sugars
• Phosphatidylethanolamines
• Polysaccharides
• Dystroglycans
• phosphatidylethanolamine
• Cytidine Diphosphate
• cytidine diphosphate glycerol
• FKRP protein, human
• Pentosyltransferases
• RNA Nucleotidyltransferases
• Ethanolamine-phosphate cytidylyltransferase
• Phosphoric Monoester Hydrolases
• glycerol-1-phosphatase
• Glycerol

Topics

• Animals
• Chromatography, Liquid (methods)
• Cytidine Diphosphate (metabolism)
• Dystroglycans (metabolism)
• Glycerol (metabolism)
• Glycosylation
• HEK293 Cells
• Humans
• Male
• Mammals
• Membrane Proteins (metabolism)
• Mice
• Mice, Inbred C57BL
• Nucleoside Diphosphate Sugars (metabolism)
• Pentosyltransferases (metabolism)
• Phosphatidylethanolamines (metabolism)
• Phosphoric Monoester Hydrolases (metabolism)
• Polysaccharides (metabolism)
• RNA Nucleotidyltransferases (metabolism)
• Tandem Mass Spectrometry (methods)