Department of Biochemistry and Biophysics, Texas A&M University, College Station, Texas 77843, USA.
J Neurosci. 2010 May 5;30(18):6466-76. doi: 10.1523/JNEUROSCI.5253-09.2010.
In vertebrates, sialylated glycans participate in a wide range of biological processes and affect the development and function of the nervous system. While the complexity of glycosylation and the functional redundancy among sialyltransferases provide obstacles for revealing biological roles of sialylation in mammals, Drosophila possesses a sole vertebrate-type sialyltransferase, Drosophila sialyltransferase (DSiaT), with significant homology to its mammalian counterparts, suggesting that Drosophila could be a suitable model to investigate the function of sialylation. To explore this possibility and investigate the role of sialylation in Drosophila, we inactivated DSiaT in vivo by gene targeting and analyzed phenotypes of DSiaT mutants using a combination of behavioral, immunolabeling, electrophysiological, and pharmacological approaches. Our experiments demonstrated that DSiaT expression is restricted to a subset of CNS neurons throughout development. We found that DSiaT mutations result in significantly decreased life span, locomotor abnormalities, temperature-sensitive paralysis, and defects of neuromuscular junctions. Our results indicate that DSiaT regulates neuronal excitability and affects the function of a voltage-gated sodium channel. Finally, we showed that sialyltransferase activity is required for DSiaT function in vivo, which suggests that DSiaT mutant phenotypes result from a defect in sialylation of N-glycans. This work provided the first evidence that sialylation has an important biological function in protostomes, while also revealing a novel, nervous system-specific function of alpha2,6-sialylation. Thus, our data shed light on one of the most ancient functions of sialic acids in metazoan organisms and suggest a possibility that this function is evolutionarily conserved between flies and mammals.
在脊椎动物中,唾液酸化糖参与了广泛的生物学过程,并影响神经系统的发育和功能。虽然糖基化的复杂性和唾液酸转移酶之间的功能冗余为揭示唾液酸化在哺乳动物中的生物学作用提供了障碍,但果蝇只拥有一种独特的脊椎动物型唾液酸转移酶,即果蝇唾液酸转移酶(DSiaT),与哺乳动物的同源性很高,这表明果蝇可能是研究唾液酸化作用的合适模型。为了探索这种可能性并研究唾液酸化在果蝇中的作用,我们通过基因靶向使 DSiaT 在体内失活,并结合行为学、免疫标记、电生理学和药理学方法分析 DSiaT 突变体的表型。我们的实验表明,DSiaT 表达在整个发育过程中局限于中枢神经系统神经元的一个子集。我们发现,DSiaT 突变导致寿命显著缩短、运动异常、温度敏感麻痹和神经肌肉接头缺陷。我们的结果表明,DSiaT 调节神经元兴奋性,并影响电压门控钠离子通道的功能。最后,我们表明唾液酸转移酶活性是 DSiaT 在体内功能所必需的,这表明 DSiaT 突变表型是由于 N-糖基化的唾液酸化缺陷所致。这项工作首次提供了证据表明,唾液酸化在原口动物中具有重要的生物学功能,同时也揭示了α2,6-唾液酸化的一种新的、神经系统特异性功能。因此,我们的数据揭示了唾液酸在后生动物中的一个最古老的功能之一,并表明这个功能在果蝇和哺乳动物之间可能是进化保守的。