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Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis

The endoplasmic reticulum (ER)-Golgi sterol transfer activity of oxysterol-binding protein (OSBP) regulates sphingomyelin (SM) synthesis, as well as post-Golgi cholesterol efflux pathways. The phosphorylation and ER-Golgi localization of OSBP are correlated, suggesting this modification regulates th...

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Autores principales: Goto, Asako, Liu, Xinwei, Robinson, Carolyn-Ann, Ridgway, Neale D.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Cell Biology 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3442410/
https://www.ncbi.nlm.nih.gov/pubmed/22875984
http://dx.doi.org/10.1091/mbc.E12-04-0283
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author Goto, Asako
Liu, Xinwei
Robinson, Carolyn-Ann
Ridgway, Neale D.
author_facet Goto, Asako
Liu, Xinwei
Robinson, Carolyn-Ann
Ridgway, Neale D.
author_sort Goto, Asako
collection PubMed
description The endoplasmic reticulum (ER)-Golgi sterol transfer activity of oxysterol-binding protein (OSBP) regulates sphingomyelin (SM) synthesis, as well as post-Golgi cholesterol efflux pathways. The phosphorylation and ER-Golgi localization of OSBP are correlated, suggesting this modification regulates the directionality and/or specificity of transfer activity. In this paper, we report that phosphorylation on two serine-rich motifs, S381-S391 (site 1) and S192, S195, S200 (site 2), specifically controls OSBP activity at the ER. A phosphomimetic of the SM/cholesterol-sensitive phosphorylation site 1 (OSBP-S5E) had increased in vitro cholesterol and 25-hydroxycholesterol–binding capacity, and cholesterol extraction from liposomes, but reduced transfer activity. Phosphatidylinositol 4-phosphate (PI(4)P) and cholesterol competed for a common binding site on OSBP; however, direct binding of PI(4)P was not affected by site 1 phosphorylation. Individual site 1 and site 2 phosphomutants supported oxysterol activation of SM synthesis in OSBP-deficient CHO cells. However, a double site1/2 mutant (OSBP-S381A/S3D) was deficient in this activity and was constitutively colocalized with vesicle-associated membrane protein–associated protein A (VAP-A) in a collapsed ER network. This study identifies phosphorylation regulation of sterol and VAP-A binding by OSBP in the ER, and PI(4)P as an alternate ligand that could be exchanged for sterol in the Golgi apparatus.
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spelling pubmed-34424102012-11-30 Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis Goto, Asako Liu, Xinwei Robinson, Carolyn-Ann Ridgway, Neale D. Mol Biol Cell Articles The endoplasmic reticulum (ER)-Golgi sterol transfer activity of oxysterol-binding protein (OSBP) regulates sphingomyelin (SM) synthesis, as well as post-Golgi cholesterol efflux pathways. The phosphorylation and ER-Golgi localization of OSBP are correlated, suggesting this modification regulates the directionality and/or specificity of transfer activity. In this paper, we report that phosphorylation on two serine-rich motifs, S381-S391 (site 1) and S192, S195, S200 (site 2), specifically controls OSBP activity at the ER. A phosphomimetic of the SM/cholesterol-sensitive phosphorylation site 1 (OSBP-S5E) had increased in vitro cholesterol and 25-hydroxycholesterol–binding capacity, and cholesterol extraction from liposomes, but reduced transfer activity. Phosphatidylinositol 4-phosphate (PI(4)P) and cholesterol competed for a common binding site on OSBP; however, direct binding of PI(4)P was not affected by site 1 phosphorylation. Individual site 1 and site 2 phosphomutants supported oxysterol activation of SM synthesis in OSBP-deficient CHO cells. However, a double site1/2 mutant (OSBP-S381A/S3D) was deficient in this activity and was constitutively colocalized with vesicle-associated membrane protein–associated protein A (VAP-A) in a collapsed ER network. This study identifies phosphorylation regulation of sterol and VAP-A binding by OSBP in the ER, and PI(4)P as an alternate ligand that could be exchanged for sterol in the Golgi apparatus. The American Society for Cell Biology 2012-09-15 /pmc/articles/PMC3442410/ /pubmed/22875984 http://dx.doi.org/10.1091/mbc.E12-04-0283 Text en © 2012 Goto et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society of Cell BD; are registered trademarks of The American Society of Cell Biology.
spellingShingle Articles
Goto, Asako
Liu, Xinwei
Robinson, Carolyn-Ann
Ridgway, Neale D.
Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis
title Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis
title_full Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis
title_fullStr Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis
title_full_unstemmed Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis
title_short Multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis
title_sort multisite phosphorylation of oxysterol-binding protein regulates sterol binding and activation of sphingomyelin synthesis
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3442410/
https://www.ncbi.nlm.nih.gov/pubmed/22875984
http://dx.doi.org/10.1091/mbc.E12-04-0283
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