Department of Medical Biochemistry and Cell Biology, University of Gothenburg, SE-405 30 Gothenburg, Sweden.
Proc Natl Acad Sci U S A. 2012 Apr 10;109(15):5645-50. doi: 10.1073/pnas.1120269109. Epub 2012 Mar 26.
MUC2, the major colonic mucin, forms large polymers by N-terminal trimerization and C-terminal dimerization. Although the assembly process for MUC2 is established, it is not known how MUC2 is packed in the regulated secretory granulae of the goblet cell. When the N-terminal VWD1-D2-D'D3 domains (MUC2-N) were expressed in a goblet-like cell line, the protein was stored together with full-length MUC2. By mimicking the pH and calcium conditions of the secretory pathway we analyzed purified MUC2-N by gel filtration, density gradient centrifugation, and transmission electron microscopy. At pH 7.4 the MUC2-N trimer eluted as a single peak by gel filtration. At pH 6.2 with Ca(2+) it formed large aggregates that did not enter the gel filtration column but were made visible after density gradient centrifugation. Electron microscopy studies revealed that the aggregates were composed of rings also observed in secretory granulae of colon tissue sections. The MUC2-N aggregates were dissolved by removing Ca(2+) and raising pH. After release from goblet cells, the unfolded full-length MUC2 formed stratified layers. These findings suggest a model for mucin packing in the granulae and the mechanism for mucin release, unfolding, and expansion.
黏蛋白 2（MUC2）是结肠的主要黏蛋白，通过 N 端三聚化和 C 端二聚化形成大的聚合物。尽管 MUC2 的组装过程已经确定，但尚不清楚 MUC2 如何在杯状细胞的调节分泌颗粒中包装。当 N 端 VWD1-D2-D'D3 结构域（MUC2-N）在杯状样细胞系中表达时，该蛋白与全长 MUC2 一起储存。通过模拟分泌途径的 pH 和钙条件，我们通过凝胶过滤、密度梯度离心和透射电子显微镜分析了纯化的 MUC2-N。在 pH 7.4 时，MUC2-N 三聚体通过凝胶过滤以单个峰洗脱。在 pH 6.2 并存在 Ca（2+）时，它形成了大的聚集体，这些聚集体不能进入凝胶过滤柱，但在密度梯度离心后变得可见。电子显微镜研究显示，这些聚集体由在结肠组织切片的分泌颗粒中也观察到的环组成。通过去除 Ca（2+）和提高 pH，可以溶解 MUC2-N 聚集体。从杯状细胞释放后，展开的全长 MUC2 形成分层层。这些发现为颗粒中的黏蛋白包装和黏蛋白释放、展开和扩展的机制提供了模型。
