Lipid Signaling Protocols

Lipid Signaling Protocols assembles in a single volume the various tools and methodologies needed by the interested investigator to unravel lipid dependent signaling and cell function. Divided into two convenient sections, the volume begins by summarizing the physical properties of hydrophobic metabolites as well as the physical methodologies used for their analysis, which leads to the second section and its selection of biological methods, focused around the most relevant lipids, their corresponding metabolizing enzymes, and the recognition proteins. Following the highly successful Methods in Molecular Biology series format, the chapters provide readily reproducible laboratory protocols, lists of necessary materials and reagents, and tips on troubleshooting and avoiding known pitfalls.

Lipid Systems and Techniques
Eicosanoids: Generation and Detection in Mammalian Cells	p. 5
Lipidomic Analysis of Phospholipids and Related Structures by Liquid Chromatography-Mass Spectrometry	p. 25
Measurement of Polyphosphoinositides in Cultured Mammalian Cells	p. 43
Inositol Lipid-Dependent Functions in Saccharomyces cerevisiae: Analysis of Phosphatidylinositol Phosphates	p. 59
Methods for the Determination of the Mass of Nuclear PtdIns4P, PtdIns5P, and PtdIns(4,5)P[subscript 2]	p. 75
Lipid Quantification and Structure Determination of Nuclear Envelope Precursor Membranes in the Sea Urchin	p. 89
Solution and Solid-State NMR of Lipids	p. 111
Spatial Localization of PtdInsP[subscript 2] in Phase-Separated Giant Unilamellar Vesicles with a Fluorescent PLC-delta 1 PH Domain	p. 135
Studying Cell-to-Cell Interactions: An Easy Method of Tethering Ligands on Artificial Membranes	p. 145
Ceramide-Induced Transbilayer (Flip-Flop) Lipid Movement in Membranes	p. 155
Effects of Sterols on the Development and Aging of Caenorhabditis elegans	p. 167
Proteomic Analysis of the Sterol-Mediated Signaling Pathway in Caenorhabditis elegans	p. 181
Lipid Metabolism and Recognition
Studying the Subcellular Localization and DNA-Binding Activity of FoxO Transcription Factors, Downstream Effectors of PI3K/Akt	p. 201
Measurement of PTEN Activity in vivo by Imaging Phosphorylated Akt	p. 213
Analysis of Phosphatidylinositol 3,4,5 Trisphosphate 5-Phosphatase Activity by in vitro and in vivo Assays	p. 223
Measuring Phospholipase D Activity in Insulin-Secreting Pancreatic [beta]-Cells and Insulin-Responsive Muscle Cells and Adipocytes	p. 241
Protein Kinase C as an Effector of Lipid-Derived Second Messengers	p. 253
Detection of Myotubularin Phosphatases Activity on Phosphoinositides in vitro and ex vivo	p. 265
Quantification of Multiple Phosphatidylinositol 4-Kinase Isozyme Activities in Cell Extracts	p. 279
Phospholipid-Interacting Proteins by Solution-State NMR Spectroscopy	p. 291
Revealing Signaling in Single Cells by Single- and Two-Photon Fluorescence Lifetime Imaging Microscopy	p. 307
Phosphoinositide (PI) 3-Kinase Assays	p. 345
Measurement of Phosphatidylinositol and Phosphatidylcholine Binding and Transfer Activity of the Lipid Transport Protein PITP	p. 363
In vitro Reconstitution of Activation of PLC[epsion] by Ras and Rho GTPases	p. 379
Assaying Endogenous Phosphatidylinositol-4-Phosphate 5-Kinase (PIP5K) Activities	p. 391
Preparation of Membrane Rafts	p. 403
Index	p. 415
Table of Contents provided by Ingram. All Rights Reserved.

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Lipid Signaling Protocols

Summary

Table of Contents

Supplemental Materials