Odd chromosome movement and inaccurate chromosome distribution in mitosis and meiosis after treatment with protein kinase inhibitors

Abstract

Errors in chromosome orientation in mitosis and meiosis are inevitable, but normally they are quickly corrected. We find that such errors usually are not corrected in cells treated with protein kinase inhibitors. Highly inaccurate chromosome distribution is the result. When grasshopper spermatocytes were treated with the kinase inhibitor 6-dimethylaminopurine (DMAP), 84% of maloriented chromosomes failed to reorient; in anaphase, both partner chromosomes were distributed to the same daughter cell. These chromosomes were observed for a total of over 60 h, and not a single reorientation was seen. In contrast, in untreated cells, maloriented chromosomes invariably reoriented, and quickly: in 10 min, on average. A second protein kinase inhibitor, genistein, had exactly the same effect as DMAP. DMAP affected PtK1 cells in mitosis as it did spermatocytes in meiosis: improper chromosome orientations persisted, leading to frequent errors in distribution. We micromanipulated chromosomes in spermatocytes treated with DMAP to learn why maloriented chromosomes often fail to reorient. Reorientation requires the loss of improper microtubule attachments and the acquisition of new, properly directed kinetochore microtubules. Micromanipulation experiments disclose that neither the loss of old nor the acquisition of new microtubules is sufficiently affected by DMAP to account for the indefinite persistence of malorientations. Drug treatment causes a novel form of chromosome movement in which one kinetochore moves toward another kinetochore. Two kinetochores in the same chromosome or in different chromosomes can participate, producing varied, dance-like movements executed by one or two chromosomes. These kinetochore-kinetochore interactions evidently are at the expense of kinetochore-spindle interactions. We propose that malorientations persist in treated cells because the kinetochores have numerous, short microtubules with a free end that can be captured by a second kinetochore. Kinetochores capture each other's kinetochore microtubules, leaving too few sites available for the efficient capture of spindle microtubules. Since the efficient capture of spindle microtubules is essential for the correction of errors, failure of capture allows malorientations to persist. Whether the effects of DMAP actually are due to protein kinase inhibition remains to be seen. In any case, DMAP reveals interactions of one kinetochore with another, which, though ordinarily suppressed, have implications for normal mitosis.