Angata Kiyohiko, Lee Wenjau, Mitoma Junya, Marth Jamey D, Fukuda Minoru
Glycobiology Program, Cancer Center, Burnham Institute for Medical Research, La Jolla, California, USA.
Methods Enzymol. 2006;417:25-37. doi: 10.1016/S0076-6879(06)17003-2.
Recent studies demonstrate that carbohydrates synthesized by specific glycosyltransferases play important roles in the development of the central nervous system. Among these carbohydrates, polysialic acid is a unique glycan that modulates functions of the neural cell adhesion molecule (NCAM) by attenuating NCAM-mediated interaction between neural cells. During brain development, polysialic acid is synthesized in a specific spatiotemporal pattern by two polysialyltransferases, ST8SiaII and ST8SiaIV. To study in vivo the roles of polysialic acid synthesized by each respective enzyme, we generated ST8SiaII and ST8SiaIV knockout mice. Single knockout ST8SiaII or ST8SiaIV mice show polysialic acid expression patterns differing from wild type, and those patterns indicate different roles of each gene during neural development. In this chapter, we discuss methods used to analyze polysialyltransferase knockout mice using immunohistochemistry of brain and primary cultures of neurons.
最近的研究表明，由特定糖基转移酶合成的碳水化合物在中枢神经系统发育中发挥重要作用。在这些碳水化合物中，多唾液酸是一种独特的聚糖，它通过减弱神经细胞粘附分子（NCAM）介导的神经细胞间相互作用来调节NCAM的功能。在大脑发育过程中，多唾液酸由两种多唾液酸转移酶ST8SiaII和ST8SiaIV以特定的时空模式合成。为了在体内研究由每种酶合成的多唾液酸的作用，我们构建了ST8SiaII和ST8SiaIV基因敲除小鼠。单基因敲除的ST8SiaII或ST8SiaIV小鼠显示出与野生型不同的多唾液酸表达模式，这些模式表明每个基因在神经发育过程中的不同作用。在本章中，我们将讨论使用脑免疫组织化学和神经元原代培养来分析多唾液酸转移酶基因敲除小鼠的方法。
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