Quantification of Signaling Lipids by Nano-Electrospray Ionization Tandem Mass Spectrometry (Nano-ESI MS/MS)

Lipids, such as phosphoinositides (PIPs) and diacylglycerol (DAG), are important signaling intermediates involved in cellular processes such as T cell receptor (TCR)-mediated signal transduction. Here we report identification and quantification of PIP, PIP2 and DAG from crude lipid extracts. Capitalizing on the different extraction properties of PIPs and DAGs allowed us to efficiently recover both lipid classes from one sample. Rapid analysis of endogenous signaling molecules was performed by nano-electrospray ionization tandem mass spectrometry (nano-ESI MS/MS), employing lipid class-specific neutral loss and multiple precursor ion scanning for their identification and quantification. Profiling of DAG, PIP and PIP2 molecular species in primary human T cells before and after TCR stimulation resulted in a two-fold increase in DAG levels with a shift towards 1-stearoyl-2-arachidonoyl-DAG in stimulated cells. PIP2 levels were slightly reduced, while PIP levels remained unchanged.

[1]  M. Hermansson,et al.  Electrospray Ionization Mass Spectrometry and Exogenous Heavy Isotope-labeled Lipid Species Provide Detailed Information on Aminophospholipid Acyl Chain Remodeling* , 2008, Journal of Biological Chemistry.

[2]  S. Feske.  Calcium signalling in lymphocyte activation and disease , 2007, Nature Reviews Immunology.

[3]  Xianlin Han,et al.  Shotgun lipidomics of cardiolipin molecular species in lipid extracts of biological samples Published, JLR Papers in Press, January 31, 2006. , 2006, Journal of Lipid Research.

[4]  Michael Loran Dustin,et al.  The lymphocyte function-associated antigen-1 receptor costimulates plasma membrane Ras via phospholipase D2 , 2007, Nature Cell Biology.

[5]  K. Takase,et al.  [T cell activation]. , 1995, Ryumachi. [Rheumatism].

[6]  R. Murphy,et al.  Electrospray ionization tandem mass spectrometry of glycerophosphoethanolamine plasmalogen phospholipids , 2004, Journal of the American Society for Mass Spectrometry.

[7]  A. Suzuki,et al.  Mammalian phosphoinositide kinases and phosphatases. , 2009, Progress in lipid research.

[8]  W. J. Dyer,et al.  A rapid method of total lipid extraction and purification. , 1959, Canadian journal of biochemistry and physiology.

[9]  J. Eichberg,et al.  Diphosphoinositide and triphosphoinositide in animal tissues. Extraction, estimation and changes post mortem. , 1965, The Biochemical journal.

[10]  Xianlin Han,et al.  Electrospray ionization mass spectroscopic analysis of human erythrocyte plasma membrane phospholipids. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[11]  J. Vickers.  Extraction of polyphosphoinositides from platelets: comparison of a two-step procedure with a common single-step extraction procedure. , 1995, Analytical biochemistry.

[12]  R. Taguchi,et al.  Reversed-phase LC/MS method for polyphosphoinositide analyses: changes in molecular species levels during epidermal growth factor activation in A431 cells. , 2008, Analytical chemistry.

[13]  G. Liebisch,et al.  Lipid profiling of lipoproteins by electrospray ionization tandem mass spectrometry. , 2011, Biochimica et biophysica acta.

[14]  K. Sauer,et al.  Lipid signaling in T-cell development and function. , 2010, Cold Spring Harbor perspectives in biology.

[15]  F. Wieland,et al.  The Membrane Domains Occupied by Glycosylphosphatidylinositol-anchored Prion Protein and Thy-1 Differ in Lipid Composition* , 2004, Journal of Biological Chemistry.

[16]  Fredrik Karpe,et al.  Quantification of PtdInsP3 molecular species in cells and tissues by mass spectrometry , 2011, Nature Methods.

[17]  M. Mann,et al.  Electrospray ionization for mass spectrometry of large biomolecules. , 1989, Science.

[18]  A. Shevchenko,et al.  Quantitative profiling of phospholipids by multiple precursor ion scanning on a hybrid quadrupole time-of-flight mass spectrometer. , 2002, Analytical chemistry.

[19]  M. Philips,et al.  Compartmentalized Ras/MAPK signaling. , 2006, Annual review of immunology.

[20]  Xianlin Han,et al.  Toward fingerprinting cellular lipidomes directly from biological samples by two-dimensional electrospray ionization mass spectrometry. , 2004, Analytical biochemistry.

[21]  Xianlin Han,et al.  Global analyses of cellular lipidomes directly from crude extracts of biological samples by ESI mass spectrometry: a bridge to lipidomics. , 2003, Journal of lipid research.

[22]  Kim Ekroos,et al.  High-throughput shotgun lipidomics by quadrupole time-of-flight mass spectrometry. , 2009, Journal of chromatography. B, Analytical technologies in the biomedical and life sciences.

[23]  H. Brown,et al.  Glycerophospholipid identification and quantitation by electrospray ionization mass spectrometry. , 2007, Methods in enzymology.

[24]  Kai Simons,et al.  Automated identification and quantification of glycerophospholipid molecular species by multiple precursor ion scanning. , 2006, Analytical chemistry.

[25]  G. Prestwich,et al.  Measurement and immunofluorescence of cellular phosphoinositides. , 2004, Methods in molecular biology.

[26]  W. Lehmann,et al.  Quantitative analysis of biological membrane lipids at the low picomole level by nano-electrospray ionization tandem mass spectrometry. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[27]  F. Cooke.  Measurement of polyphosphoinositides in cultured mammalian cells. , 2009, Methods in molecular biology.

[28]  D. Corda,et al.  CD28 co‐stimulates TCR/CD3‐induced phosphoinositide turnover in human T lymphocytes , 2001, European journal of immunology.

[29]  A. Bielawska,et al.  Determination of ceramides and diglycerides by the diglyceride kinase assay. , 2001, Analytical biochemistry.

[30]  I. Batty,et al.  The regulation of membrane to cytosol partitioning of signalling proteins by phosphoinositides and their soluble headgroups. , 2005, Biochemical Society transactions.

[31]  Christer S. Ejsing,et al.  Lipid profiling by multiple precursor and neutral loss scanning driven by the data-dependent acquisition. , 2006, Analytical chemistry.

[32]  H. Callender,et al.  Quantification of diacylglycerol species from cellular extracts by electrospray ionization mass spectrometry using a linear regression algorithm. , 2007, Analytical chemistry.

[33]  M. Wakelam,et al.  Methods for analyzing phosphoinositides using mass spectrometry. , 2011, Biochimica et biophysica acta.

[34]  A. Godi,et al.  The role of the phosphoinositides at the Golgi complex. , 2005, Biochimica et biophysica acta.

[35]  S. Fleischer,et al.  Two dimensional thin layer chromatographic separation of polar lipids and determination of phospholipids by phosphorus analysis of spots , 1970, Lipids.

[36]  F. Hsu,et al.  Structural determination of glycosphingolipids as lithiated adducts by electrospray ionization mass spectrometry using low-energy collisional-activated dissociation on a triple stage quadrupole instrument , 2001, Journal of the American Society for Mass Spectrometry.

[37]  Michael J O Wakelam,et al.  Analysis of intact phosphoinositides in biological samples Published, JLR Papers in Press, April 21, 2006. , 2006, Journal of Lipid Research.

[38]  M. Karnovsky,et al.  Extraction of polyphosphoinositides with neutral and acidified solvents. A comparison of guinea-pig brain and liver, and measurements of rat liver inositol compounds which are resistant to extraction. , 1970, Biochimica et biophysica acta.

[39]  I. Bird.  Analysis of cellular phosphoinositides and phosphoinositols by extraction and simple analytical procedures. , 1994, Methods in molecular biology.

[40]  Xianlin Han,et al.  Shotgun lipidomics: electrospray ionization mass spectrometric analysis and quantitation of cellular lipidomes directly from crude extracts of biological samples. , 2005, Mass spectrometry reviews.

[41]  A. Tuininga,et al.  Identification of molecular species of glycerophospholipids and sphingomyelin using electrospray mass spectrometry. , 1994, Journal of lipid research.

[42]  M. Gross,et al.  Quantification of diacylglycerol molecular species in biological samples by electrospray ionization mass spectrometry after one-step derivatization. , 2007, Analytical chemistry.

[43]  E. Gottfried,et al.  Lipids of human leukocytes: relation to celltype. , 1967, Journal of lipid research.

[44]  D. Decamp,et al.  A targeted mass spectrometric analysis of phosphatidylinositol phosphate species Published, JLR Papers in Press, May 16, 2005. DOI 10.1194/jlr.D500010-JLR200 , 2005, Journal of Lipid Research.

[45]  Christer S. Ejsing,et al.  Global analysis of the yeast lipidome by quantitative shotgun mass spectrometry , 2009, Proceedings of the National Academy of Sciences.

[46]  J. Takemoto,et al.  Analysis of phosphoinositides in protein trafficking. , 2000, Methods.

[47]  Kai Simons,et al.  Membrane lipidome of an epithelial cell line , 2011, Proceedings of the National Academy of Sciences.

[48]  R. Murphy,et al.  Detection of the abundance of diacylglycerol and triacylglycerol molecular species in cells using neutral loss mass spectrometry. , 2007, Analytical biochemistry.

[49]  Takao Shimizu,et al.  A shotgun tandem mass spectrometric analysis of phospholipids with normal-phase and/or reverse-phase liquid chromatography/electrospray ionization mass spectrometry. , 2005, Rapid communications in mass spectrometry : RCM.

[50]  S. Schurmans,et al.  Phosphoinositide phosphatases in a network of signalling reactions , 2007, Pflügers Archiv - European Journal of Physiology.

[51]  Y. Shoenfeld,et al.  [The T cell receptor]. , 1985, Harefuah.

[52]  Morgan Huse,et al.  The T-cell-receptor signaling network , 2009, Journal of Cell Science.

[53]  L. Cantley,et al.  Pathways for phosphoinositide synthesis. , 1999, Chemistry and physics of lipids.

[54]  F. Hsu,et al.  Characterization of phosphatidylinositol, phosphatidylinositol-4-phosphate, and phosphatidylinositol-4,5-bisphosphate by electrospray ionization tandem mass spectrometry: A mechanistic study , 2000, Journal of the American Society for Mass Spectrometry.

[55]  P. Falardeau,et al.  Quantification of diacylglycerols by capillary gas chromatography-negative ion chemical ionization-mass spectrometry. , 1993, Analytical biochemistry.

[56]  Pietro De Camilli,et al.  Phosphoinositides in cell regulation and membrane dynamics , 2006, Nature.

[57]  W. Hubbard,et al.  Quantitation of 1-stearoyl-2-arachidonoyl-sn-3-glycerol in human basophils via gas chromatography-negative ion chemical ionization mass spectrometry. , 1996, Analytical biochemistry.

[58]  J. Folch,et al.  A simple method for the isolation and purification of total lipides from animal tissues. , 1957, The Journal of biological chemistry.

[59]  S. Grinstein,et al.  SopB promotes phosphatidylinositol 3-phosphate formation on Salmonella vacuoles by recruiting Rab5 and Vps34 , 2008, The Journal of cell biology.

[60]  S. Blanksby,et al.  Advances in mass spectrometry for lipidomics. , 2010, Annual review of analytical chemistry.

[61]  Henric Olsson,et al.  Nonradioactive methods for the assay of phosphoinositide 3-kinases and phosphoinositide phosphatases and selective detection of signaling lipids in cell and tissue extracts. , 2003, Analytical biochemistry.

[62]  R. Nussbaum,et al.  Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry , 2003, Nature Biotechnology.