Abstract
Phosphoinositides are rapidly turning-over phospholipids that play key roles in intracellular signaling and modulation of membrane effectors. Through technical refinements we have improved sensitivity in the analysis of the phosphoinositide PI, PIP, and PIP2 pools from living cells using mass spectrometry. This has permitted further resolution in phosphoinositide lipidomics from cell cultures and small samples of tissue. The technique includes butanol extraction, derivatization of the lipids, post-column infusion of sodium to stabilize formation of sodiated adducts, and electrospray ionization mass spectrometry in multiple reaction monitoring mode, achieving a detection limit of 20 pg. We describe the spectrum of fatty-acyl chains in the cellular phosphoinositides. Consistent with previous work in other mammalian primary cells, the 38:4 fatty-acyl chains dominate in the phosphoinositides of the pineal gland and of superior cervical ganglia, and many additional fatty acid combinations are found at low abundance. However, Chinese hamster ovary cells and human embryonic kidney cells (tsA201) in culture have different fatty-acyl chain profiles that change with growth state. Their 38:4 lipids lose their dominance as cultures approach confluence. The method has good time resolution and follows well the depletion in < 20 s of both PIP2 and PIP that results from strong activation of Gq-coupled receptors. The receptor-activated phospholipase C exhibits no substrate selectivity among the various fatty-acyl chain combinations.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 513-522 |
| Number of pages | 10 |
| Journal | Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids |
| Volume | 1862 |
| Issue number | 5 |
| DOIs | |
| State | Published - May 1 2017 |
| Externally published | Yes |
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Keywords
- Arachidonic acid
- Lipidomics
- Mass spectrometry
- Phospholipids
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology
Cite this
Fatty-acyl chain profiles of cellular phosphoinositides. / Traynor-Kaplan, Alexis; Kruse, Martin; Dickson, Eamonn J; Dai, Gucan; Vivas, Oscar; Yu, Haijie; Whittington, Dale; Hille, Bertil.
In: Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids, Vol. 1862, No. 5, 01.05.2017, p. 513-522.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Fatty-acyl chain profiles of cellular phosphoinositides
AU - Traynor-Kaplan, Alexis
AU - Kruse, Martin
AU - Dickson, Eamonn J
AU - Dai, Gucan
AU - Vivas, Oscar
AU - Yu, Haijie
AU - Whittington, Dale
AU - Hille, Bertil
PY - 2017/5/1
Y1 - 2017/5/1
N2 - Phosphoinositides are rapidly turning-over phospholipids that play key roles in intracellular signaling and modulation of membrane effectors. Through technical refinements we have improved sensitivity in the analysis of the phosphoinositide PI, PIP, and PIP2 pools from living cells using mass spectrometry. This has permitted further resolution in phosphoinositide lipidomics from cell cultures and small samples of tissue. The technique includes butanol extraction, derivatization of the lipids, post-column infusion of sodium to stabilize formation of sodiated adducts, and electrospray ionization mass spectrometry in multiple reaction monitoring mode, achieving a detection limit of 20 pg. We describe the spectrum of fatty-acyl chains in the cellular phosphoinositides. Consistent with previous work in other mammalian primary cells, the 38:4 fatty-acyl chains dominate in the phosphoinositides of the pineal gland and of superior cervical ganglia, and many additional fatty acid combinations are found at low abundance. However, Chinese hamster ovary cells and human embryonic kidney cells (tsA201) in culture have different fatty-acyl chain profiles that change with growth state. Their 38:4 lipids lose their dominance as cultures approach confluence. The method has good time resolution and follows well the depletion in < 20 s of both PIP2 and PIP that results from strong activation of Gq-coupled receptors. The receptor-activated phospholipase C exhibits no substrate selectivity among the various fatty-acyl chain combinations.
AB - Phosphoinositides are rapidly turning-over phospholipids that play key roles in intracellular signaling and modulation of membrane effectors. Through technical refinements we have improved sensitivity in the analysis of the phosphoinositide PI, PIP, and PIP2 pools from living cells using mass spectrometry. This has permitted further resolution in phosphoinositide lipidomics from cell cultures and small samples of tissue. The technique includes butanol extraction, derivatization of the lipids, post-column infusion of sodium to stabilize formation of sodiated adducts, and electrospray ionization mass spectrometry in multiple reaction monitoring mode, achieving a detection limit of 20 pg. We describe the spectrum of fatty-acyl chains in the cellular phosphoinositides. Consistent with previous work in other mammalian primary cells, the 38:4 fatty-acyl chains dominate in the phosphoinositides of the pineal gland and of superior cervical ganglia, and many additional fatty acid combinations are found at low abundance. However, Chinese hamster ovary cells and human embryonic kidney cells (tsA201) in culture have different fatty-acyl chain profiles that change with growth state. Their 38:4 lipids lose their dominance as cultures approach confluence. The method has good time resolution and follows well the depletion in < 20 s of both PIP2 and PIP that results from strong activation of Gq-coupled receptors. The receptor-activated phospholipase C exhibits no substrate selectivity among the various fatty-acyl chain combinations.
KW - Arachidonic acid
KW - Lipidomics
KW - Mass spectrometry
KW - Phospholipids
UR - http://www.scopus.com/inward/record.url?scp=85013029098&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85013029098&partnerID=8YFLogxK
U2 - 10.1016/j.bbalip.2017.02.002
DO - 10.1016/j.bbalip.2017.02.002
M3 - Article
C2 - 28189644
AN - SCOPUS:85013029098
VL - 1862
SP - 513
EP - 522
JO - Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
JF - Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
SN - 1388-1981
IS - 5
ER -