POLARITY AND NUCLEATION OF MICROTUBULES IN POLARIZED EPITHELIAL-CELLS

Microtubules oriented in the apicobasal axis of columnar epithelial ce lls are arranged with a uniform polarity with minus ends toward the ap ical 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 w hich transcellular transport is a requisite step in normal apical memb rane biogenesis, contain microtubules arranged with a similar polarity . In the present study, we explore the question of microtubule polarit y 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 polarit y with minus ends nearest the apical surface. After cold and nocodazol e-induced depolymerization, microtubule regrowth initiated in the apic al 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 g amma-tubulin was present in complexes that sedimented between 10S and 29S; both forms could bind microtubules. The presence of both centroso mal and noncentrosomal gamma-tubulin in apical cytoplasm suggests mult iple mechanisms by which microtubule nucleation might occur in epithel ial cells. (C) 1995 Wiley-Liss, Inc.