Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions

Mathias J. Gerl, Verena Bittl, Susanne Kirchner, Timo Sachsenheimer, Hanna L. Brunner, Christian Lüchtenborg, Cagakan Özbalci, Hannah Wiedemann, Sabine Wegehingel, Walter Nickel, Per Haberkant, Carsten Schultz, Marcus Krüger, Britta Brügger

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Cell membranes contain hundreds to thousands of individual lipid species that are of structural importance but also specifically interact with proteins. Due to their highly controlled synthesis and role in signaling events sphingolipids are an intensely studied class of lipids. In order to investigate their metabolism and to study proteins interacting with sphingolipids, metabolic labeling based on photoactivatable sphingoid bases is the most straightforward approach. In order to monitor protein-lipid-crosslink products, sphingosine derivatives containing a reporter moiety, such as a radiolabel or a clickable group, are used. In normal cells, degradation of sphingoid bases via action of the checkpoint enzyme sphingosine-1-phosphate lyase occurs at position C2-C3 of the sphingoid base and channels the resulting hexadecenal into the glycerolipid biosynthesis pathway. In case the functionalized sphingosine looses the reporter moiety during its degradation, specificity towards sphingolipid labeling is maintained. In case degradation of a sphingosine derivative does not remove either the photoactivatable or reporter group from the resulting hexadecenal, specificity towards sphingolipid labeling can be achieved by blocking sphingosine-1-phosphate lyase activity and thus preventing sphingosine derivatives to be channeled into the sphingolipid-to-glycerolipid metabolic pathway. Here we report an approach using clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated nuclease Cas9 to create a sphingosine-1-phosphate lyase (SGPL1) HeLa knockout cell line to disrupt the sphingolipidto-glycerolipid metabolic pathway. We found that the lipid and protein compositions as well as sphingolipid metabolism of SGPL1 knock-out HeLa cells only show little adaptations, which validates these cells as model systems to study transient protein-sphingolipid interactions.

Original languageEnglish (US)
Article numbere0153009
JournalPLoS One
Volume11
Issue number4
DOIs
StatePublished - Apr 1 2016
Externally publishedYes

Fingerprint

sphingosine
sphingolipids
Sphingolipids
lyases
phosphates
Sphingosine
Lipids
lipids
Proteins
Labeling
proteins
cells
chemical derivatives
Metabolic Networks and Pathways
Derivatives
HeLa Cells
Metabolism
Degradation
degradation
biochemical pathways

ASJC Scopus subject areas

  • Medicine(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Gerl, M. J., Bittl, V., Kirchner, S., Sachsenheimer, T., Brunner, H. L., Lüchtenborg, C., ... Brügger, B. (2016). Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions. PLoS One, 11(4), [e0153009]. https://doi.org/10.1371/journal.pone.0153009

Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions. / Gerl, Mathias J.; Bittl, Verena; Kirchner, Susanne; Sachsenheimer, Timo; Brunner, Hanna L.; Lüchtenborg, Christian; Özbalci, Cagakan; Wiedemann, Hannah; Wegehingel, Sabine; Nickel, Walter; Haberkant, Per; Schultz, Carsten; Krüger, Marcus; Brügger, Britta.

In: PLoS One, Vol. 11, No. 4, e0153009, 01.04.2016.

Research output: Contribution to journalArticle

Gerl, MJ, Bittl, V, Kirchner, S, Sachsenheimer, T, Brunner, HL, Lüchtenborg, C, Özbalci, C, Wiedemann, H, Wegehingel, S, Nickel, W, Haberkant, P, Schultz, C, Krüger, M & Brügger, B 2016, 'Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions', PLoS One, vol. 11, no. 4, e0153009. https://doi.org/10.1371/journal.pone.0153009
Gerl MJ, Bittl V, Kirchner S, Sachsenheimer T, Brunner HL, Lüchtenborg C et al. Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions. PLoS One. 2016 Apr 1;11(4). e0153009. https://doi.org/10.1371/journal.pone.0153009
Gerl, Mathias J. ; Bittl, Verena ; Kirchner, Susanne ; Sachsenheimer, Timo ; Brunner, Hanna L. ; Lüchtenborg, Christian ; Özbalci, Cagakan ; Wiedemann, Hannah ; Wegehingel, Sabine ; Nickel, Walter ; Haberkant, Per ; Schultz, Carsten ; Krüger, Marcus ; Brügger, Britta. / Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions. In: PLoS One. 2016 ; Vol. 11, No. 4.
@article{8d075c2c9b35451bb3f72eeb4d6ad96c,
title = "Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions",
abstract = "Cell membranes contain hundreds to thousands of individual lipid species that are of structural importance but also specifically interact with proteins. Due to their highly controlled synthesis and role in signaling events sphingolipids are an intensely studied class of lipids. In order to investigate their metabolism and to study proteins interacting with sphingolipids, metabolic labeling based on photoactivatable sphingoid bases is the most straightforward approach. In order to monitor protein-lipid-crosslink products, sphingosine derivatives containing a reporter moiety, such as a radiolabel or a clickable group, are used. In normal cells, degradation of sphingoid bases via action of the checkpoint enzyme sphingosine-1-phosphate lyase occurs at position C2-C3 of the sphingoid base and channels the resulting hexadecenal into the glycerolipid biosynthesis pathway. In case the functionalized sphingosine looses the reporter moiety during its degradation, specificity towards sphingolipid labeling is maintained. In case degradation of a sphingosine derivative does not remove either the photoactivatable or reporter group from the resulting hexadecenal, specificity towards sphingolipid labeling can be achieved by blocking sphingosine-1-phosphate lyase activity and thus preventing sphingosine derivatives to be channeled into the sphingolipid-to-glycerolipid metabolic pathway. Here we report an approach using clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated nuclease Cas9 to create a sphingosine-1-phosphate lyase (SGPL1) HeLa knockout cell line to disrupt the sphingolipidto-glycerolipid metabolic pathway. We found that the lipid and protein compositions as well as sphingolipid metabolism of SGPL1 knock-out HeLa cells only show little adaptations, which validates these cells as model systems to study transient protein-sphingolipid interactions.",
author = "Gerl, {Mathias J.} and Verena Bittl and Susanne Kirchner and Timo Sachsenheimer and Brunner, {Hanna L.} and Christian L{\"u}chtenborg and Cagakan {\"O}zbalci and Hannah Wiedemann and Sabine Wegehingel and Walter Nickel and Per Haberkant and Carsten Schultz and Marcus Kr{\"u}ger and Britta Br{\"u}gger",
year = "2016",
month = "4",
day = "1",
doi = "10.1371/journal.pone.0153009",
language = "English (US)",
volume = "11",
journal = "PLoS One",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "4",

}

TY - JOUR

T1 - Sphingosine-1-phosphate lyase deficient cells as a tool to study protein lipid interactions

AU - Gerl, Mathias J.

AU - Bittl, Verena

AU - Kirchner, Susanne

AU - Sachsenheimer, Timo

AU - Brunner, Hanna L.

AU - Lüchtenborg, Christian

AU - Özbalci, Cagakan

AU - Wiedemann, Hannah

AU - Wegehingel, Sabine

AU - Nickel, Walter

AU - Haberkant, Per

AU - Schultz, Carsten

AU - Krüger, Marcus

AU - Brügger, Britta

PY - 2016/4/1

Y1 - 2016/4/1

N2 - Cell membranes contain hundreds to thousands of individual lipid species that are of structural importance but also specifically interact with proteins. Due to their highly controlled synthesis and role in signaling events sphingolipids are an intensely studied class of lipids. In order to investigate their metabolism and to study proteins interacting with sphingolipids, metabolic labeling based on photoactivatable sphingoid bases is the most straightforward approach. In order to monitor protein-lipid-crosslink products, sphingosine derivatives containing a reporter moiety, such as a radiolabel or a clickable group, are used. In normal cells, degradation of sphingoid bases via action of the checkpoint enzyme sphingosine-1-phosphate lyase occurs at position C2-C3 of the sphingoid base and channels the resulting hexadecenal into the glycerolipid biosynthesis pathway. In case the functionalized sphingosine looses the reporter moiety during its degradation, specificity towards sphingolipid labeling is maintained. In case degradation of a sphingosine derivative does not remove either the photoactivatable or reporter group from the resulting hexadecenal, specificity towards sphingolipid labeling can be achieved by blocking sphingosine-1-phosphate lyase activity and thus preventing sphingosine derivatives to be channeled into the sphingolipid-to-glycerolipid metabolic pathway. Here we report an approach using clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated nuclease Cas9 to create a sphingosine-1-phosphate lyase (SGPL1) HeLa knockout cell line to disrupt the sphingolipidto-glycerolipid metabolic pathway. We found that the lipid and protein compositions as well as sphingolipid metabolism of SGPL1 knock-out HeLa cells only show little adaptations, which validates these cells as model systems to study transient protein-sphingolipid interactions.

AB - Cell membranes contain hundreds to thousands of individual lipid species that are of structural importance but also specifically interact with proteins. Due to their highly controlled synthesis and role in signaling events sphingolipids are an intensely studied class of lipids. In order to investigate their metabolism and to study proteins interacting with sphingolipids, metabolic labeling based on photoactivatable sphingoid bases is the most straightforward approach. In order to monitor protein-lipid-crosslink products, sphingosine derivatives containing a reporter moiety, such as a radiolabel or a clickable group, are used. In normal cells, degradation of sphingoid bases via action of the checkpoint enzyme sphingosine-1-phosphate lyase occurs at position C2-C3 of the sphingoid base and channels the resulting hexadecenal into the glycerolipid biosynthesis pathway. In case the functionalized sphingosine looses the reporter moiety during its degradation, specificity towards sphingolipid labeling is maintained. In case degradation of a sphingosine derivative does not remove either the photoactivatable or reporter group from the resulting hexadecenal, specificity towards sphingolipid labeling can be achieved by blocking sphingosine-1-phosphate lyase activity and thus preventing sphingosine derivatives to be channeled into the sphingolipid-to-glycerolipid metabolic pathway. Here we report an approach using clustered, regularly interspaced, short palindromic repeats (CRISPR)-associated nuclease Cas9 to create a sphingosine-1-phosphate lyase (SGPL1) HeLa knockout cell line to disrupt the sphingolipidto-glycerolipid metabolic pathway. We found that the lipid and protein compositions as well as sphingolipid metabolism of SGPL1 knock-out HeLa cells only show little adaptations, which validates these cells as model systems to study transient protein-sphingolipid interactions.

UR - http://www.scopus.com/inward/record.url?scp=84964653334&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84964653334&partnerID=8YFLogxK

U2 - 10.1371/journal.pone.0153009

DO - 10.1371/journal.pone.0153009

M3 - Article

C2 - 27100999

AN - SCOPUS:84964653334

VL - 11

JO - PLoS One

JF - PLoS One

SN - 1932-6203

IS - 4

M1 - e0153009

ER -