The mouse C2C12 myoblast cell surface N-linked glycoproteome: identification, glycosite occupancy, and membrane orientation.
The mouse C2C12 myoblast cell surface N-linked glycoproteome: identification, glycosite occupancy, and membrane orientation.
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DOI：
10.1074/mcp.m900195-mcp200
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发表时间：
2009-11
期刊：
影响因子：
--
通讯作者：
Wollscheid B
中科院分区：
文献类型：
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作者：
Gundry RL;Raginski K;Tarasova Y;Tchernyshyov I;Bausch-Fluck D;Elliott ST;Boheler KR;Van Eyk JE;Wollscheid B
内源性再生和修复机制负责替换死亡和受损的细胞，以维持或增强组织和器官的功能，骨骼肌是内源性修复机制的最好例子之一。虽然调控卫星细胞和成肌细胞向肌纤维分化的分子机制尚不完全清楚，但感知和响应环境的细胞表面蛋白起着重要作用。细胞表面捕获技术被用于揭示成肌细胞的细胞表面n -连接糖蛋白亚蛋白质组，并鉴定成肌细胞分化的潜在标记。从小鼠C2C12成肌细胞中鉴定出128个真正暴露于细胞表面的n -连接糖蛋白，包括117个跨膜蛋白、4个糖基磷脂酰肌醇锚定蛋白、5个细胞外基质蛋白和2个膜相关蛋白。该数据集显示了36个带分化注释的蛋白簇，并确认了235个n-链糖基化位点的占用。鉴定蛋白质胞外结构域的n -糖基化位点可以确定鉴定的蛋白质在质膜内的取向。发现一个糖蛋白跨膜取向与Swiss-Prot注释不一致，而其他14个蛋白质的模糊注释得到解决。在验证实验中发现，当成肌细胞向肌管分化时，几种已鉴定的n-连接糖蛋白（包括水通道蛋白-1和β-肌聚糖）的总体丰度会发生变化。因此，该策略和数据揭示了成肌细胞表面亚蛋白质组的复杂性，并揭示了成肌细胞向肌管分化过程中细胞中间体的临床重要特征的新靶点。
Endogenous regeneration and repair mechanisms are responsible for replacing dead and damaged cells to maintain or enhance tissue and organ function, and one of the best examples of endogenous repair mechanisms involves skeletal muscle. Although the molecular mechanisms that regulate the differentiation of satellite cells and myoblasts toward myofibers are not fully understood, cell surface proteins that sense and respond to their environment play an important role. The cell surface capturing technology was used here to uncover the cell surface N-linked glycoprotein subproteome of myoblasts and to identify potential markers of myoblast differentiation. 128 bona fide cell surface-exposed N-linked glycoproteins, including 117 transmembrane, four glycosylphosphatidylinositol-anchored, five extracellular matrix, and two membrane-associated proteins were identified from mouse C2C12 myoblasts. The data set revealed 36 cluster of differentiation-annotated proteins and confirmed the occupancy for 235 N-linked glycosylation sites. The identification of the N-glycosylation sites on the extracellular domain of the proteins allowed for the determination of the orientation of the identified proteins within the plasma membrane. One glycoprotein transmembrane orientation was found to be inconsistent with Swiss-Prot annotations, whereas ambiguous annotations for 14 other proteins were resolved. Several of the identified N-linked glycoproteins, including aquaporin-1 and β-sarcoglycan, were found in validation experiments to change in overall abundance as the myoblasts differentiate toward myotubes. Therefore, the strategy and data presented shed new light on the complexity of the myoblast cell surface subproteome and reveal new targets for the clinically important characterization of cell intermediates during myoblast differentiation into myotubes.