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Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry

Nature Biotechnology volume 21, pages 813–817 (2003)Cite this article

Abstract

Phosphoinositides (phosphorylated derivatives of phosphatidylinositol, PI) are versatile intracellular signaling lipids whose occurrence in low concentrations complicates direct mass measurements1,2,3. Here we present a sensitive method to detect, identify and quantify phosphatidylinositol phosphate (PIP) and phosphatidylinositol bisphosphate (PIP2) with different fatty acid compositions (phosphoinositide profiles) in total lipid extracts by electrospray ionization mass spectrometry (ESI-MS). Using this method, we detected elevated concentrations of PIP2 in human fibroblasts from patients with Lowe syndrome, a genetic disorder that affects phosphoinositide metabolism4. Saccharomyces cerevisiae cells deficient in enzymes involved in PIP metabolism—Sac1p, a phosphoinositide phosphatase5, and Vps34p and Pik1p, a PI 3-kinase6 and PI 4-kinase7, respectively—showed not only different PIP concentrations but also differential changes in PIP profiles indicating metabolic and/or subcellular pooling. Mass spectrometric analysis of phosphoinositides offers unique advantages over existing approaches and may represent a powerful diagnostic tool for human diseases that involve defective phosphoinositide metabolism.

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Figure 1: Detection, identification and quantification of phosphoinositides in total brain lipid extract by ESI-MS.
Figure 2: Detection of enzymatic products from phosphoinositide kinase and phosphatase reactions in vivo and profiling of phosphoinositides in yeast mutants.

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Acknowledgements

We thank Peter Novick for providing us with yeast strains. This work was supported by grants from the US National Institutes of Health (NS36251, CA46128, DK 54913 and ADA to P.D.C. and DK59635 and DK49230 to G.I.S.).

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Authors and Affiliations

  1. Department of Cell Biology, Howard Hughes Medical Institute, Yale University School of Medicine, Boyer Center for Molecular Medicine, 295 Congress Avenue, New Haven, 06511, Connecticut, USA

    Markus R Wenk, Louise Lucast, Gilbert Di Paolo & Pietro De Camilli

  2. Department of Internal Medicine, Howard Hughes Medical Institute, Yale University School of Medicine, Congress Avenue Building, 300 Cedar Street, New Haven, 06511, Connecticut

    Anthony J Romanelli, Gary W Cline & Gerald I Shulman

  3. The W.M. Keck Mass Spectrometry Resource, Yale University School of Medicine, Boyer Center for Molecular Medicine, 295 Congress Avenue, New Haven, 06511, Connecticut, USA

    Walter McMurray

  4. National Human Genome Research Institute, National Institutes of Health, 49 Convent Dr., Bethesda, 20892-4472, Maryland, USA

    Sharon F Suchy & Robert L Nussbaum

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  1. Markus R Wenk
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Correspondence to Markus R Wenk.

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Wenk, M., Lucast, L., Di Paolo, G. et al. Phosphoinositide profiling in complex lipid mixtures using electrospray ionization mass spectrometry. Nat Biotechnol 21, 813–817 (2003). https://doi.org/10.1038/nbt837

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