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Vol. 11, Issue 3, 863-872, March 2000
Department of Cell Biology and Physiology, Washington University
School of Medicine, St. Louis, Missouri 63110
In budding yeast, the mitotic spindle is positioned in the neck
between the mother and the bud so that both cells inherit one nucleus.
The movement of the mitotic spindle into the neck can be divided into
two phases: (1) Kip3p-dependent movement of the nucleus to the neck and
alignment of the short spindle, followed by (2) dynein-dependent
movement of the spindle into the neck and oscillation of the elongating
spindle within the neck. Actin has been hypothesized to be involved in
all these movements. To test this hypothesis, we disrupted the actin
cytoskeleton with the use of mutations and latrunculin A (latrunculin).
We assayed nuclear segregation in synchronized cell populations and
observed spindle movements in individual living cells. In synchronized cell populations, no actin cytoskeletal mutant segregated nuclei as
poorly as cells lacking dynein function. Furthermore, nuclei segregated
efficiently in latrunculin-treated cells. Individual living cell
analysis revealed that the preanaphase spindle was mispositioned and
misaligned in latrunculin-treated cells and that astral microtubules
were misoriented, confirming a role for filamentous actin in the early,
Kip3p-dependent phase of spindle positioning. Surprisingly,
mispositioned and misaligned mitotic spindles moved into the neck in
the absence of filamentous actin, albeit less efficiently. Finally,
dynein-dependent sliding of astral microtubules along the cortex and
oscillation of the elongating mitotic spindle in the neck occurred in
the absence of filamentous actin.
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