A Hybrid Approach Enabling Large-Scale Glycomic Analysis of Post-Golgi Vesicles Reveals a Transport Route for Polysaccharides
A Hybrid Approach Enabling Large-Scale Glycomic Analysis of Post-Golgi Vesicles Reveals a Transport Route for Polysaccharides
复制标题
DOI：
10.1105/tpc.18.00854
复制
发表时间：
2019-03-01
期刊：
影响因子：
11.6
通讯作者：
Drakakaki，Georgia
中科院分区：
文献类型：
--
作者：
Wilkop，Thomas;Pattathil，Sivakumar;Drakakaki，Georgia
植物内膜系统通过其动态途径促进多糖、相关酶和糖蛋白的运输。尽管参与细胞壁生物合成的酶已被鉴定，但对以膜为基础的糖组分的运输知之甚少。这部分归因于在生化测定特定囊泡中的多糖含量方面的技术挑战。在这里，我们引入一种混合方法来解决这一限制。通过结合囊泡分离和大规模碳水化合物抗体排列技术，我们绘制了一个初步的大规模图谱，描述了拟南芥(Arabitopsis Thaliana)中PLANTS61(SYP61)跨高尔基网络隔室的Synaxin(SYP61)的糖类分布。识别特定非纤维素碳水化合物表位的抗体库使我们能够在分离的小泡中识别一系列不同的多糖，包括果胶、木葡聚糖(XyGs)和阿拉伯半乳糖蛋白。拟南芥SYP61突变体细胞壁中XyG和果胶特异性表位的变化证实了我们的发现。我们的数据提供了SYP61囊泡参与结构多糖和糖蛋白的运输和沉积的证据。我们方法的适应可以使研究表征植物和动物系统中不同囊泡种群的糖类及其在由亚细胞标志物、发育阶段或环境刺激所定义的糖运输中的各自作用。
The plant endomembrane system facilitates the transport of polysaccharides, associated enzymes, and glycoproteins through its dynamic pathways. Although enzymes involved in cell wall biosynthesis have been identified, little is known about the endomembrane-based transport of glycan components. This is partially attributed to technical challenges in biochemically determining polysaccharide cargo in specific vesicles. Here, we introduce a hybrid approach addressing this limitation. By combining vesicle isolation with a large-scale carbohydrate antibody arraying technique, we charted an initial large-scale map describing the glycome profile of the SYNTAXIN OF PLANTS61 (SYP61)trans-Golgi network compartment in Arabidopsis (Arabidopsis thaliana). A library of antibodies recognizing specific noncellulosic carbohydrate epitopes allowed us to identify a range of diverse glycans, including pectins, xyloglucans (XyGs), and arabinogalactan proteins in isolated vesicles. Changes in XyG- and pectin-specific epitopes in the cell wall of an ArabidopsisSYP61mutant corroborate our findings. Our data provide evidence that SYP61 vesicles are involved in the transport and deposition of structural polysaccharides and glycoproteins. Adaptation of our methodology can enable studies characterizing the glycome profiles of various vesicle populations in plant and animal systems and their respective roles in glycan transport defined by subcellular markers, developmental stages, or environmental stimuli.