Meads T, Schroer T A
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218, USA.
Cell Motil Cytoskeleton. 1995;32(4):273-88. doi: 10.1002/cm.970320404.
Microtubules oriented in the apicobasal axis of columnar epithelial cells are arranged with a uniform polarity with minus ends toward the apical surface, suggesting that these cytoskeletal filaments might serve as a substrate for polarized movement of membrane vesicles within the cell. It is not known whether hepatocytes, a cuboidal epithelium in which transcellular transport is a requisite step in normal apical membrane biogenesis, contain microtubules arranged with a similar polarity. In the present study, we explore the question of microtubule polarity and possible mechanisms for nucleation in the epithelial cell lines WIF-B (hepatocyte), Caco-2 (intestine), and Madin-Darby canine kidney (MDCK). Caco-2 microtubules in the apicobasal axis had uniform polarity with minus ends nearest the apical surface. After cold and nocodazole-induced depolymerization, microtubule regrowth initiated in the apical region in all three cell types. The apex of WIF-B and Caco-2 cells contained two pools of gamma-tubulin: one associated with centrosomes and the other delocalized under the apical membrane. Non-centrosomal gamma-tubulin was present in complexes that sedimented between 10S and 29S; both forms could bind microtubules. The presence of both centrosomal and noncentrosomal gamma-tubulin in apical cytoplasm suggest multiple mechanisms by which microtubule nucleation might occur in epithelial cells.
沿柱状上皮细胞顶基轴排列的微管具有统一的极性，负端朝向顶端表面，这表明这些细胞骨架细丝可能作为细胞内膜泡极化运动的底物。目前尚不清楚肝细胞（一种立方上皮细胞，其中跨细胞运输是正常顶端膜生物发生的必要步骤）是否含有极性相似的微管。在本研究中，我们探讨了上皮细胞系WIF-B（肝细胞）、Caco-2（肠）和Madin-Darby犬肾（MDCK）中微管极性问题和成核的可能机制。沿顶基轴的Caco-2微管具有统一的极性，负端最靠近顶端表面。在冷诱导和诺考达唑诱导解聚后，所有三种细胞类型的微管在顶端区域开始重新生长。WIF-B和Caco-2细胞的顶端含有两池γ-微管蛋白：一池与中心体相关，另一池在顶端膜下呈非定位状态。非中心体γ-微管蛋白存在于沉降在10S至29S之间的复合物中；两种形式都能结合微管。顶端细胞质中同时存在中心体和非中心体γ-微管蛋白，提示上皮细胞中微管成核可能存在多种机制。
