Soya Seçkin, Şahar Umut, Karaçalı Sabire
Molecular Biology Section, Department of Biology, Faculty of Science, Ege University, Izmir, Turkey.
Invert Neurosci. 2016 Sep;16(3):8. doi: 10.1007/s10158-016-0191-6. Epub 2016 Jun 24.
Glycoconjugates have various functions in differentiation, development, aging and in all aspects of normal functioning of organisms. The reason for increased research on this topic is that glycoconjugates locate mostly on the cell surface and play crucial biological roles in the nervous system including brain development, synaptic plasticity, learning, and memory. Considering their roles in the nervous system, information about their existence in the insect nervous system is rather sparse. Therefore, in order to detect monosaccharide content of N- and O-glycans, we carried out capLC-ESI-MS/MS analysis to determine the concentration changes of glucose, mannose, galactose, N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), fucose, xylose, arabinose, and ribose monosaccharides in the nervous system of Bombyx mori during development and aging processes. In addition to LC-MS, lectin blotting was done to detect quantitative changes in N- and O-glycans. Developmental stages were selected as 3rd (the youngest sample), 5th (young) larval instar, motionless prepupa (the oldest sample), and pupa (adult development). Derivatization of monosaccharides was performed with a solution of PMP agent and analyzed with capLC-ESI-MS/MS. For lectin blotting, determination of glycan types was carried out with Galanthus nivalis agglutinin and Peanut agglutinin lectins. In all stages, the most abundant monosaccharide was glucose. Although all monosaccharides were present most abundantly in the youngest stage (3rd instar), they are generally reduced gradually during the aging process. It was observed that amounts of monosaccharides increased again in the pupa stage. According to lectin blotting, N- and O-linked glycoproteins expressions were different and there were some specific glycoprotein expression differences between stages. These findings suggest that the glycosylation state of proteins in the nervous system changes during development and aging in insects in a similar fashion to that reported for vertebrates.
糖缀合物在分化、发育、衰老以及生物体正常功能的各个方面都具有多种功能。对这一主题研究增加的原因是，糖缀合物大多位于细胞表面，在神经系统中发挥着关键的生物学作用，包括大脑发育、突触可塑性、学习和记忆。考虑到它们在神经系统中的作用，关于它们在昆虫神经系统中存在情况的信息相当稀少。因此，为了检测N - 聚糖和O - 聚糖的单糖含量，我们进行了capLC - ESI - MS/MS分析，以确定家蚕神经系统在发育和衰老过程中葡萄糖、甘露糖、半乳糖、N - 乙酰葡糖胺（GlcNAc）、N - 乙酰半乳糖胺（GalNAc）、岩藻糖、木糖、阿拉伯糖和核糖等单糖的浓度变化。除了液相色谱 - 质谱分析外，还进行了凝集素印迹法以检测N - 聚糖和O - 聚糖的定量变化。发育阶段选择为第3龄（最年轻的样本）、第5龄（年轻）幼虫期、静止蛹期（最老的样本）和蛹期（成虫发育阶段）。单糖的衍生化用PMP试剂溶液进行，并通过capLC - ESI - MS/MS分析。对于凝集素印迹法，用雪花莲凝集素和花生凝集素确定聚糖类型。在所有阶段，最丰富的单糖是葡萄糖。尽管所有单糖在最年轻阶段（第3龄）含量最高，但在衰老过程中它们通常会逐渐减少。观察到单糖含量在蛹期再次增加。根据凝集素印迹法，N - 连接和O - 连接糖蛋白的表达不同，并且各阶段之间存在一些特定的糖蛋白表达差异。这些发现表明，昆虫神经系统中蛋白质的糖基化状态在发育和衰老过程中发生变化，其方式与脊椎动物中报道的相似。
