Fatty-acyl chain profiles of cellular phosphoinositides

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 spectr...

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Published in:Biochimica et biophysica acta. Molecular and cell biology of lipids Vol. 1862; no. 5; pp. 513 - 522
Main Authors: Traynor-Kaplan, Alexis, Kruse, Martin, Dickson, Eamonn J., Dai, Gucan, Vivas, Oscar, Yu, Haijie, Whittington, Dale, Hille, Bertil
Format: Journal Article
Language:English
Published: Netherlands Elsevier B.V 01.05.2017
Subjects:
animal ovaries
Animals
Arachidonic acid
butanol
cell culture
Chinese hamsters
CHO Cells
Cricetinae
Cricetulus
derivatization
detection limit
electrospray ionization mass spectrometry
fatty acids
Fatty Acids - chemistry
Fatty Acids - isolation & purification
ganglia
Humans
kidney cells
Lipidomics
Mass spectrometry
monitoring
Phosphatidylinositols - chemistry
Phosphatidylinositols - isolation & purification
phospholipase C
Phospholipids
Phospholipids - chemistry
Phospholipids - isolation & purification
pineal body
receptors
Signal Transduction
sodium
Spectrometry, Mass, Electrospray Ionization - methods
PLC
MRM
Lipidomics
Phospholipids
DAG
Arachidonic acid
IP3
PIP
SCG
PI
D-PI
TQ
Mass spectrometry
PIP2
ISSN:1388-1981, 1879-2618
Online Access:Get full text
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Summary: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 20pg. 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 <20s 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. •Refined quantitative mass spectrometry of cell phosphoinositides•Resolution of many fatty-acyl chain combinations in phosphoinositides•Dominance of 38:4 lipids in mammalian primary cells but not in cultured cells•Change of fatty-acyl profile with culture conditions•Fast kinetics (<10s) of phosphoinositide depletion after receptor activation
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ISSN:1388-1981
1879-2618
DOI:10.1016/j.bbalip.2017.02.002