Heparan sulfate polymerization in Drosophila
Heparan sulfate polymerization in Drosophila
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DOI：
10.1074/jbc.m509138200
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发表时间：
2006-01-27
影响因子：
4.8
通讯作者：
Kitagawa, H
中科院分区：
文献类型：
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作者：
Izumikawa, T;Egusa, N;Kitagawa, H
硫酸肝素（HS）链的形成是由EXT（遗传性多发性外泌素基因）家族成员编码的糖基转移酶催化的。对影响果蝇形态因子信号通路的突变进行遗传筛选，发现了3个基因，分别编码人类EXT1、EXT2和EXTL3的同源基因，分别为tout-velu （ttv）、tout-velu的姊妹基因（sotv）和tout-velu的兄弟基因（botv）。到目前为止，只有BOTV/DEXTL3具有糖基转移酶的体外活性，它具有n -乙酰氨基葡萄糖转移酶- i （GlcNAcT-I）和n -乙酰氨基葡萄糖转移酶- ii （GlcNAcT-II）活性，负责HS的链起始和延伸，而没有葡萄糖醛酸转移酶- ii （GlcAT-II）活性。在这里，我们证明TTV/ DEXT1和SOTV/DEXT2具有生物合成HS主干重复双糖单元所需的GlcNAcT-II和GlcAT-II活性，并且与TTV或SOTV单独表达相比，TTV与SOTV共表达显著增强了糖基转移酶活性。此外，以BOTV和由TTV和SOTV组成的酶配合物（TTV- SOTV）为受体底物，在连接区类似物上聚合了HS。与人类相比，TTV-SOTV没有GlcNAcT-I活性，这表明BOTV/DEXT3是一个EXT-Like基因，具有HS起始所需的GlcNAcT-I活性，对于果蝇HS链的生物合成是必不可少的。因此，在果蝇中TTV、SOTV和BOTV这三个EXT成员都是果蝇全长HS生物合成所必需的。
The formation of heparan sulfate (HS) chains is catalyzed by glycosyltransferases encoded by EXT (hereditary multiple exostosin gene) family members. Genetic screening for mutations affecting morphogen signaling pathways in Drosophila has identified three genes, tout-velu (ttv), sister of tout-velu (sotv), and brother of tout-velu (botv), which encode homologues of human EXT1, EXT2, and EXTL3, respectively. So far, in vitro glycosyltransferase activities have been demonstrated only for BOTV/DEXTL3, which harbors both N-acetylglucosaminyltransferase-I ( GlcNAcT-I) and N-acetylglucosaminyltransferase-II (GlcNAcT-II) activities responsible for the chain initiation and elongation of HS, and no glucuronyltransferase-II (GlcAT-II) activity. Here we demonstrated that TTV/ DEXT1 and SOTV/DEXT2 had GlcNAcT-II and GlcAT-II activities required for the biosynthesis of repeating disaccharide units of the HS backbone, and the coexpression of TTV with SOTV markedly augmented both glycosyltransferase activities when compared with the expression of TTV or SOTV alone. Moreover, the polymerization of HS was demonstrated on a linkage region analogue as an acceptor substrate by BOTV and an enzyme complex composed of TTV and SOTV (TTV- SOTV). In contrast to human, TTV-SOTV exhibited no GlcNAcT-I activity, indicating that BOTV/DEXT3, which is an EXT-Like gene and possesses GlcNAcT-I activity required for the initiation of HS, is indispensable for the biosynthesis of HS chains in Drosophila. Thus, all three EXT members in Drosophila, TTV, SOTV, and BOTV, are required for the biosynthesis of full-length HS in Drosophila.