title: Oxysterols inhibit phosphatidylcholine synthesis via ERK docking and phosphorylation of CTP:phosphocholine cytidylyltransferase.
abstract: Surfactant deficiency contributes to acute lung injury and may result from the elaboration of bioactive lipids such as oxysterols. We observed that the oxysterol 22-hydroxycholesterol (22-HC) in combination with its obligate partner, 9-cis-retinoic acid (9-cis-RA), decreased surfactant phosphatidylcholine (PtdCho) synthesis by increasing phosphorylation of the regulatory enzyme CTP:phosphocholine cytidylyltransferase-alpha (CCTalpha). Phosphorylation of CCTalpha decreased its activity. 22-HC/9-cis-RA inhibition of PtdCho synthesis was blocked by PD98059 or dominant-negative ERK (p42 kinase). Overexpression of constitutively active MEK1, the kinase upstream of p42 kinase, increased CCTalpha phosphorylation. Expression of truncated CCTalpha mutants lacking proline-directed sites within the C-terminal phosphorylation domain partially blocked oxysterol-mediated inhibition of PtdCho synthesis. Mutagenesis of Ser315 within CCTalpha was both required and sufficient to confer significant resistance to 22-HC/9-cis-RA inhibition of PtdCho synthesis. A novel putative ERK-docking domain N-terminal to this phosphoacceptor site was mapped within the CCTalpha membrane-binding domain (residues 287-300). The results are the first demonstration of a physiologically relevant phosphorylation site and docking domain within CCTalpha that serve as targets for ERKs, resulting in inhibition of surfactant synthesis.
authors.affiliation: Department of Internal Medicine, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City 52242, USA.
authors.forename: Marianna
authors.initials: M
authors.lastname: Agassandian
authors.forename: Jiming
authors.initials: J
authors.lastname: Zhou
authors.forename: Linda A
authors.initials: LA
authors.lastname: Tephly
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Ryan
authors.forename: A Brent
authors.initials: AB
authors.lastname: Carter
authors.forename: Rama K
authors.initials: RK
authors.lastname: Mallampalli
mesh_terms: D000077556:Alitretinoin; D000818:Animals; D001665:Binding Sites; D019916:Choline-Phosphate Cytidylyltransferase / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D005419:Flavonoids / Q000494:pharmacology; D005799:Genes, Dominant; D015151:Immunoblotting; D047468:Immunoprecipitation; D008168:Lung / Q000166:cytology; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D016296:Mutagenesis; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010713:Phosphatidylcholines / Q000378:metabolism*; D010766:Phosphorylation; D011392:Proline / Q000737:chemistry; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D011987:Receptors, Steroid / Q000502:physiology*; D013261:Sterols / Q000378:metabolism*; D013501:Surface-Active Agents / Q000494:pharmacology; D013997:Time Factors; D014158:Transcription, Genetic; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology; D000098825:Oxysterol Binding Proteins
chemical_list: D018076:DNA, Complementary; D005419:Flavonoids; D010713:Phosphatidylcholines; D011987:Receptors, Steroid; D013261:Sterols; D013501:Surface-Active Agents; D000098825:Oxysterol Binding Proteins; D000077556:Alitretinoin; D014212:Tretinoin; D011392:Proline; D048049:Extracellular Signal-Regulated MAP Kinases; D019950:Mitogen-Activated Protein Kinase 1; D019916:Choline-Phosphate Cytidylyltransferase; C500611:Pcyt1a protein, mouse; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one