Tubulin Tyrosine Ligase-like Genes ttll3 and ttll6 Maintain Zebrafish Cilia Structure and Motility
Tubulin Tyrosine Ligase-like Genes ttll3 and ttll6 Maintain Zebrafish Cilia Structure and Motility
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DOI：
10.1074/jbc.m110.209817
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发表时间：
2011-04-01
影响因子：
4.8
通讯作者：
Drummond, Iain A.
中科院分区：
文献类型：
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作者：
Pathak, Narendra;Austin, Christina A.;Drummond, Iain A.
微管蛋白翻译后修饰产生微管异质性并调节微管功能，并由微管蛋白酪氨酸连接酶样（TTLL）蛋白催化。在斑马鱼胚胎的整装免疫染色中，使用单甘氨酰化、多甘氨酰化和谷氨酰化微管蛋白的特异性抗体，我们观察到微管蛋白修饰的不同的组织特异性模式。纤毛中的微管蛋白修饰模式与纤毛细胞中ttll 3和ttll 6的表达相关。所有斑马鱼微管蛋白酪氨酸连接酶样基因的表达筛选显示，ttll 1在脑神经元中，ttll 4在肌肉中，ttll 7在耳基板中有额外的组织特异性表达。ttll 3的敲除消除纤毛微管蛋白的甘氨酰化，但对纤毛结构和运动性具有令人惊讶的温和影响。类似地，ttll 6的敲低强烈降低纤毛微管蛋白谷氨酰化，但仅部分影响纤毛结构和运动性。ttll 3和ttll 6的功能丧失导致纤毛运动几乎完全丧失，并诱导多种轴丝超微结构缺陷，类似于先前在斑马鱼fleer突变体中观察到的缺陷，这些突变体显示缺乏微管蛋白谷氨酰化。一致地，我们发现fleer突变体也缺乏微管蛋白糖基化。这些结果表明，微管蛋白糖基化和谷氨酰化在维持纤毛结构和运动性方面具有重叠的功能，并且fleer/dyf-1 TPR蛋白是两种类型的微管蛋白翻译后修饰所需的。
Tubulin post-translational modifications generate microtubule heterogeneity and modulate microtubule function, and are catalyzed by tubulin tyrosine ligase-like (TTLL) proteins. Using antibodies specific to monoglycylated, polyglycylated, and glutamylated tubulin in whole mount immunostaining of zebrafish embryos, we observed distinct, tissue-specific patterns of tubulin modifications. Tubulin modification patterns in cilia correlated with the expression of ttll3 and ttll6 in ciliated cells. Expression screening of all zebrafish tubulin tyrosine ligase-like genes revealed additional tissue-specific expression of ttll1 in brain neurons, ttll4 in muscle, and ttll7 in otic placodes. Knockdown of ttll3 eliminated cilia tubulin glycylation but had surprisingly mild effects on cilia structure and motility. Similarly, knockdown of ttll6 strongly reduced cilia tubulin glutamylation but only partially affected cilia structure and motility. Combined loss of function of ttll3 and ttll6 caused near complete loss of cilia motility and induced a variety of axonemal ultrastructural defects similar to defects previously observed in zebrafish fleer mutants, which were shown to lack tubulin glutamylation. Consistently, we find that fleer mutants also lack tubulin glycylation. These results indicate that tubulin glycylation and glutamylation have overlapping functions in maintaining cilia structure and motility and that the fleer/dyf-1 TPR protein is required for both types of tubulin post-translational modification.