Cell biology of spatial regulation of formation of the cell wall

1.在红豆杉边缘凹陷的形成过程中，皮质微管参与了细胞壁的局限性沉积。2.建立了观察墙体三维结构的方法。用这种方法在豌豆上胚轴中发现，纤维素微纤维在进入坑区后立即中断。这表明次生加厚的质膜(PM)结构域和凹坑区域分别激活和失活纤维素合成酶。3.半纤维素仅分布在次生加厚层中。这表明纤维素和半纤维素的沉积至少在一定程度上受相同机制的控制。4.甲胺磷对微管的破坏与次生增厚的沉积模式一样，表现为纤维素和半纤维素的无序沉积。这意味着微管定义了增厚和凹陷之间的边界。5.基于上述结果，我提出了如下假说：携带纤维素合成酶和半纤维素的高尔基体小泡只与次生增厚的PM融合。纤维素合成酶携带到PM中，在增稠的PM中合成纤维素微纤维，但进入窖池PM时失去活性。皮质微管界定了增厚的PM区域和凹陷之间的边界。6.为了验证这一假说，我们尝试利用Closterium细胞分离和鉴定与壁合成相关的PM蛋白。大量合成纤维素的细胞质膜蛋白与少量合成纤维素的细胞质膜蛋白在双向PAGE图谱上有一定的差异。

1.Involvement of cortical microtubules in localized deposition of the cell wall was demonstrated in formation of bordered pit in Taxus. 2.A method to observe three dimensional structure of the wall was established. It was found in pea epicotyls by this method that cellulose microfibrils are interrupted immediately after entering into the pit areas. This indicates that the plasma membrane(PM) domains of secondary thickenings and pit areas activate and inactivate cellulose synthase, respectively. 3.Hemicellulose was distributed only in secondary thickenings. This indicates that depositions of cellulose and hemicellulose are controlled at least in part by the same mechanism. 4.Disruption of microtubules by amiprophos-methyl disordered deposition patterns of cellulose and hemicellulose in the same way as did the deposition pattern of the secondary thickening. This means that microtubules define the boundary between the thickening and the pit. 5.Based on the above results, I proposed the following hypothesis : Golgi vesicles carrying cellulose synthase and hemicelluloses fuse only with the PM of the secondary thickenings. Cellulose synthases carried to the PM synthesize cellulose microfibrils in the PM of the thickening but lose their activity when entering into the PM of the pit. Cortical microtubules define the boundary between the PM domains in the thickening and the pit. 6.Toward proving this hypothesis, separation and identification of PM proteins related to wall synthesis was tried using Closterium cells. Some differences in spot patterns of two dimensional PAGE were detected between PM proteins of cells synthesizing much and little cellulose.