Berrie Christopher P, Iurisci Cristiano, Piccolo Enza, Bagnati Renzo, Corda Daniela
Department of Cell Biology and Oncology, Consorzio Mario Negri Sud, Santa Maria Imbaro, Italy.
Methods Enzymol. 2007;434:187-232. doi: 10.1016/S0076-6879(07)34011-1.
Many and various experimental techniques have been developed to fully analyze the intracellular signaling pathways of membrane phosphoinositides and their water-soluble derivatives. This chapter concentrates mainly on the range of lipid-derived, water-soluble signaling molecules that can be produced in cells from these membrane phosphoinositides, for which we and others have proposed biological roles. These include lysophosphatidylinositol, produced via phospholipase A(1/2) activities on phosphatidylinositol; cyclic inositol phosphates, produced via phosphatidylinositol/lysophosphatidylinositol-specific phospholipase C activities; and glycerophosphoinositols, produced via lysophospholipase A(2/1) activities on their corresponding lysophosphoinositides. While the methodologies described in this chapter are aimed more specifically at the separation, identification, and quantification of monophosphorylated glyceropho sphoinositols and other similarly charged inositol-containing products of the membrane phosphoinositides in cell extracts, they can be equally well applied to the full range of lysophosphoinositides, glycerophosphoinositols, inositol phosphates, and further inositol-containing water-soluble products of the phosphoinositides (e.g., cyclic inositol phosphates, methylphosphoinositol phosphates).
人们已经开发出许多各种各样的实验技术，用于全面分析膜磷酸肌醇及其水溶性衍生物的细胞内信号传导途径。本章主要关注可从这些膜磷酸肌醇在细胞中产生的一系列脂质衍生的水溶性信号分子，我们和其他人已为其提出了生物学作用。这些包括通过磷脂酶A(1/2)对磷脂酰肌醇的作用产生的溶血磷脂酰肌醇；通过磷脂酰肌醇/溶血磷脂酰肌醇特异性磷脂酶C活性产生的环肌醇磷酸酯；以及通过溶血磷脂酶A(2/1)对其相应的溶血磷脂酰肌醇的作用产生的甘油磷酸肌醇。虽然本章所述方法更具体地针对细胞提取物中膜磷酸肌醇的单磷酸化甘油磷酸肌醇和其他带类似电荷的含肌醇产物的分离、鉴定和定量，但它们同样可很好地应用于溶血磷脂酰肌醇、甘油磷酸肌醇、肌醇磷酸酯以及磷酸肌醇的其他含肌醇水溶性产物（例如环肌醇磷酸酯、甲基磷酸肌醇磷酸酯）的全范围分析。
