Isolation and partial characterization of a cage of filaments that surrounds the mammalian mitotic spindle.

Abstract

Mitotic cells have been detergent extracted under conditions that support microtubule assembly. When HeLa cells are lysed in the presence of brain tubulin, mitotic-arrested cells nucleate large asters and true metaphase cells yield spindles that remain enclosed within a roughly spherical cage of filamentous material. Detergent-extracted mitotic Chinese hamster ovary (CHO) cells show a similar, insoluble cage but the mitotic apparatus is only occasionally stabilized. In later stages of mitosis, HeLa cages are observed in elongated and furrowed configurations. In the terminal stages of cell division, 2 daughter filamentous networks are connected by the intercellular bridge. When observed by EM the cages include fibers 7-11 nm in diameter. The polypeptide composition of cages isolated from mitotic HeLa cells is complex, but the major polypeptides are a group with MW ranging from 43,000-60,000 daltons and a high-MW polypeptide. CHO cells contain a subset of these proteins which includes a major 58,000-dalton and a high-MW polypeptide. Different antisera (2) directed against the vimentin-containing intermediate filaments bind to polypeptides in the electrophoretic profiles of isolated HeLa and CHO cages and stain the cages, as visualized by indirect immunofluorescence. Apparently the HeLa and CHO cages include intermediate filaments of the vimentin type. The polypeptide composition of HeLa cages suggests that they also contain tonofilaments. The cages apparently form as the cells enter mitosis. These filamentous cages may maintain the structural continuity of the cytoplasm while the cell is in mitosis.