POLARITY ORIENTATION AND ASSEMBLY PROCESS OF MICROTUBULE BUNDLES IN NOCODAZOLE-TREATED, MAP2C-TRANSFECTED COS CELLS

Microtubule bundles reminiscent of those found in neuronal processes a re formed in fibroblasts and Sf9 cells that are transfected with the m icrotubule-associated proteins tau, MAP2, or MAP2c. To analyze the ass embly process of these bundles and its relation to the microtubule pol arity, we depolymerized the bundles formed in MAP2c-transfected COS ce lls using nocodazole, and observed the process of assembly of microtub ule bundles after removal of the drug in cells microinjected with rhod amine-labeled tubulin. Within minutes of its removal, numerous short m icrotubule fragments were observed throughout the cytoplasm. These sho rt fragments were randomly oriented and were already bundled. Somewhat longer, but still short bundles, were then found in the peripheral cy toplasm. These bundles became the primordium of the larger bundles, an d gradually grew in length and width. The polarity orientation of micr otubules in the reformed bundle as determined by ''hook'' procedure us ing electron microscope was uniform with the plus end distal to the ce ll nucleus. The results suggest that some mechanism(s) exists to orien t the polarity of microtubules, which are not in direct continuity wit h the centrosome, during the formation of large bundles. The observed process presents a useful model system for studying the organization o f microtubules that are not directly associated with the centrosomes, such as those observed in axons.