Microtubules are required for chromosome motion and directed cell locomotion. Recent experiments had demonstrated that microtubules are dynamic polymers and that dynamic behavior is modulated throughout the cell cycle. A major aim of these experiments is to quantitate microtubule dynamic behavior throughout the cell cycle and to examine the function and regulation of microtubule dynamics. To address these questions, fluorescent or biotinylated derivatives of tubulin will be microinjected into living cells and the rate and pattern of microtubule turnover examined using fluorescence or electron microscopy. Microtubule dynamic behavior will be examined in interphase and correlated with cell motility. The dramatic reorganization of the interphase microtubule array at the transition from interphase to mitosis will be examined in living cells. Spindle fiber formation during prometaphase will be characterized; particular attention will be paid to the interaction of the kinetochore with spindle microtubules. Does capture and release of microtubules within the kinetochore fiber occur during prometaphase? The half-time of kinetochore-microtubule interactions will be measured to determine if the affinity of microtubules for the kinetochore changes during mitosis. Finally, experiments will be initiated to identify regulatory molecules involved in modulation of microtubule dynamics in living cells. The results of this, research will provide significant new insight into the mechanisms of cell motility and mitosis.
