Niemann-Pick C2 protein regulates sterol transport between plasma membrane and late endosomes in human fibroblasts

Zane Berzina, Lukasz M Solanko, Ahmed S Mehadi, Maria Louise V Jensen, Frederik W Lund, Maciej Modzel, Maria Szomek, Katarzyna A Solanko, Alice Dupont, Gitte Krogh Nielsen, Christian W Heegaard, Christer S Ejsing, Daniel Wüstner

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Abstract

Niemann-Pick disease type C2 is a lipid storage disorder in which mutations in the NPC2 protein cause accumulation of lipoprotein-derived cholesterol in late endosomes and lysosomes (LE/LYSs). Whether cholesterol delivered by other means to NPC2 deficient cells also accumulates in LE/LYSs is currently unknown. We show that the close cholesterol analog dehydroergosterol (DHE), when delivered to the plasma membrane (PM) accumulates in LE/LYSs of human fibroblasts lacking functional NPC2. We measured two different time scales of sterol diffusion; while DHE rich LE/LYSs moved by slow anomalous diffusion in disease cells (D ∼ 4.6∙10-4 μm2/sec; α∼0.76), a small pool of sterol could exchange rapidly with D ∼ 3 μm2/s between LE/LYSs, as shown by fluorescence recovery after photobleaching (FRAP). By quantitative lipid mass spectrometry we found that esterification of13C-labeled cholesterol but not of DHE is reduced 10-fold in disease fibroblasts compared to control cells. Internalized NPC2 rescued the sterol storage phenotype and strongly expanded the dynamic sterol pool seen in FRAP experiments. Together, our study shows that cholesterol esterification and trafficking of sterols between the PM and LE/LYSs depends on a functional NPC2 protein. NPC2 likely acts inside LE/LYSs from where it increases non-vesicular sterol exchange with other organelles.

Original languageEnglish
JournalChemistry and Physics of Lipids
Volume213
Pages (from-to)48-61
ISSN0009-3084
DOIs
Publication statusPublished - 2018

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Sterols
Fibroblasts
Cell membranes
Cell Membrane
Cholesterol
Photobleaching
Proteins
Fluorescence Recovery After Photobleaching
Esterification
Fluorescence
Lipids
Recovery
Lipoproteins
Mass spectrometry
Mutation
dehydroergosterol
Experiments

Cite this

Berzina, Zane ; Solanko, Lukasz M ; Mehadi, Ahmed S ; Jensen, Maria Louise V ; Lund, Frederik W ; Modzel, Maciej ; Szomek, Maria ; Solanko, Katarzyna A ; Dupont, Alice ; Nielsen, Gitte Krogh ; Heegaard, Christian W ; Ejsing, Christer S ; Wüstner, Daniel. / Niemann-Pick C2 protein regulates sterol transport between plasma membrane and late endosomes in human fibroblasts. In: Chemistry and Physics of Lipids. 2018 ; Vol. 213. pp. 48-61.
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title = "Niemann-Pick C2 protein regulates sterol transport between plasma membrane and late endosomes in human fibroblasts",
abstract = "Niemann-Pick disease type C2 is a lipid storage disorder in which mutations in the NPC2 protein cause accumulation of lipoprotein-derived cholesterol in late endosomes and lysosomes (LE/LYSs). Whether cholesterol delivered by other means to NPC2 deficient cells also accumulates in LE/LYSs is currently unknown. We show that the close cholesterol analog dehydroergosterol (DHE), when delivered to the plasma membrane (PM) accumulates in LE/LYSs of human fibroblasts lacking functional NPC2. We measured two different time scales of sterol diffusion; while DHE rich LE/LYSs moved by slow anomalous diffusion in disease cells (D ∼ 4.6∙10-4 μm2/sec; α∼0.76), a small pool of sterol could exchange rapidly with D ∼ 3 μm2/s between LE/LYSs, as shown by fluorescence recovery after photobleaching (FRAP). By quantitative lipid mass spectrometry we found that esterification of13C-labeled cholesterol but not of DHE is reduced 10-fold in disease fibroblasts compared to control cells. Internalized NPC2 rescued the sterol storage phenotype and strongly expanded the dynamic sterol pool seen in FRAP experiments. Together, our study shows that cholesterol esterification and trafficking of sterols between the PM and LE/LYSs depends on a functional NPC2 protein. NPC2 likely acts inside LE/LYSs from where it increases non-vesicular sterol exchange with other organelles.",
author = "Zane Berzina and Solanko, {Lukasz M} and Mehadi, {Ahmed S} and Jensen, {Maria Louise V} and Lund, {Frederik W} and Maciej Modzel and Maria Szomek and Solanko, {Katarzyna A} and Alice Dupont and Nielsen, {Gitte Krogh} and Heegaard, {Christian W} and Ejsing, {Christer S} and Daniel W{\"u}stner",
note = "Copyright {\circledC} 2018 Elsevier B.V. All rights reserved.",
year = "2018",
doi = "10.1016/j.chemphyslip.2018.03.006",
language = "English",
volume = "213",
pages = "48--61",
journal = "Chemistry and Physics of Lipids",
issn = "0009-3084",
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Niemann-Pick C2 protein regulates sterol transport between plasma membrane and late endosomes in human fibroblasts. / Berzina, Zane; Solanko, Lukasz M; Mehadi, Ahmed S; Jensen, Maria Louise V; Lund, Frederik W; Modzel, Maciej; Szomek, Maria; Solanko, Katarzyna A; Dupont, Alice; Nielsen, Gitte Krogh; Heegaard, Christian W; Ejsing, Christer S; Wüstner, Daniel.

In: Chemistry and Physics of Lipids, Vol. 213, 2018, p. 48-61.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Niemann-Pick C2 protein regulates sterol transport between plasma membrane and late endosomes in human fibroblasts

AU - Berzina, Zane

AU - Solanko, Lukasz M

AU - Mehadi, Ahmed S

AU - Jensen, Maria Louise V

AU - Lund, Frederik W

AU - Modzel, Maciej

AU - Szomek, Maria

AU - Solanko, Katarzyna A

AU - Dupont, Alice

AU - Nielsen, Gitte Krogh

AU - Heegaard, Christian W

AU - Ejsing, Christer S

AU - Wüstner, Daniel

N1 - Copyright © 2018 Elsevier B.V. All rights reserved.

PY - 2018

Y1 - 2018

N2 - Niemann-Pick disease type C2 is a lipid storage disorder in which mutations in the NPC2 protein cause accumulation of lipoprotein-derived cholesterol in late endosomes and lysosomes (LE/LYSs). Whether cholesterol delivered by other means to NPC2 deficient cells also accumulates in LE/LYSs is currently unknown. We show that the close cholesterol analog dehydroergosterol (DHE), when delivered to the plasma membrane (PM) accumulates in LE/LYSs of human fibroblasts lacking functional NPC2. We measured two different time scales of sterol diffusion; while DHE rich LE/LYSs moved by slow anomalous diffusion in disease cells (D ∼ 4.6∙10-4 μm2/sec; α∼0.76), a small pool of sterol could exchange rapidly with D ∼ 3 μm2/s between LE/LYSs, as shown by fluorescence recovery after photobleaching (FRAP). By quantitative lipid mass spectrometry we found that esterification of13C-labeled cholesterol but not of DHE is reduced 10-fold in disease fibroblasts compared to control cells. Internalized NPC2 rescued the sterol storage phenotype and strongly expanded the dynamic sterol pool seen in FRAP experiments. Together, our study shows that cholesterol esterification and trafficking of sterols between the PM and LE/LYSs depends on a functional NPC2 protein. NPC2 likely acts inside LE/LYSs from where it increases non-vesicular sterol exchange with other organelles.

AB - Niemann-Pick disease type C2 is a lipid storage disorder in which mutations in the NPC2 protein cause accumulation of lipoprotein-derived cholesterol in late endosomes and lysosomes (LE/LYSs). Whether cholesterol delivered by other means to NPC2 deficient cells also accumulates in LE/LYSs is currently unknown. We show that the close cholesterol analog dehydroergosterol (DHE), when delivered to the plasma membrane (PM) accumulates in LE/LYSs of human fibroblasts lacking functional NPC2. We measured two different time scales of sterol diffusion; while DHE rich LE/LYSs moved by slow anomalous diffusion in disease cells (D ∼ 4.6∙10-4 μm2/sec; α∼0.76), a small pool of sterol could exchange rapidly with D ∼ 3 μm2/s between LE/LYSs, as shown by fluorescence recovery after photobleaching (FRAP). By quantitative lipid mass spectrometry we found that esterification of13C-labeled cholesterol but not of DHE is reduced 10-fold in disease fibroblasts compared to control cells. Internalized NPC2 rescued the sterol storage phenotype and strongly expanded the dynamic sterol pool seen in FRAP experiments. Together, our study shows that cholesterol esterification and trafficking of sterols between the PM and LE/LYSs depends on a functional NPC2 protein. NPC2 likely acts inside LE/LYSs from where it increases non-vesicular sterol exchange with other organelles.

U2 - 10.1016/j.chemphyslip.2018.03.006

DO - 10.1016/j.chemphyslip.2018.03.006

M3 - Journal article

C2 - 29580834

VL - 213

SP - 48

EP - 61

JO - Chemistry and Physics of Lipids

JF - Chemistry and Physics of Lipids

SN - 0009-3084

ER -