Video
data from "A
Cell Cycle Timer for Asymmetric Spindle Positioning"
Erin K. McCarthy
Campbell, Adam D. Werts, Bob Goldstein
| These
experiments demonstrated that APC-dependent negative regulation of CDK,
long known to
function as a mitotic progression timer, serves a second role as a
timer for spindle displacement during an asymmetric cell division. This
timing mechanism ensures that
spindle assembly is completed before spindle components begin to move
away from the center of the cell. |
Video S1. The Mitotic Spindle Begins to Shift
Approximately 10 s after the
Completion of Chromosome Congression. Movie of the embryo from
Figure
1B–D
expressing histone H2B:GFP and gamma-tubulin:GFP, and a dynamically
annotated version of the corresponding graph. Dotted line in the movie
marks 50% embryo length. As the fluorescence ratio in the center of the
spindle to just outside this region peaks at metaphase, displacement of
the spindle with a compact metaphase plate is evident in the embryo and
the graph. Light blue datapoints in this graph indicate the center of
the two chromatin masses after anaphase.
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Video S2. Disruption of Proteasome Activity Delayed
Both Spindle Displacement and
Anaphase Onset. Three cases of each treatment: wild-type
untreated (top three), rpt-6(RNAi)
(middle three), and c-LβL (bottom three). Small regions of each embryo
are shown, each through most of the mitotic spindle. The red line is
50% embryo length, and each movie starts at nuclear envelope breakdown.
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Video S3. Premature CDK Inactivation Caused Premature
Spindle Displacement and
Anaphase Onset. Three cases of each, untreated (top three) and
flavopiridol-treated
(bottom three). Small regions of each embryo
are shown, each through most of the mitotic spindle. The red line is
50% embryo length, and each movie starts at nuclear envelope breakdown.
Blank frames are
shown at time of flavopiridol-treatment.
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Video S4. Anaphase Timing Is Unaffected When Spindle
Displacement Is Prevented.
WT is wild-type control. The vertical line is
50% embryo length, and each movie starts at nuclear envelope breakdown.
Spindle displacement was
prevented by par-2
RNAi or gpr-1/2 RNAi.
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Video S5. Flavopiridol
Treatment as Nuclear
Envelope Breakdown Began Resulted in Chromosomes Being Spread
over a
Significantly Wider Area Than Normal at the Time of Spindle Displacement. An
embryo laser-permeabilized to flavopiridol as nuclear
envelope breakdown began (right).
Spindle displacement toward the posterior (top) and anaphase both occur
before chromosome congression is complete in the flavopiridol-treated
embryo. The embryo in the right is the bottom embryo among the
flavopiridol-treated embryos in Figure
6A.
The embryo on the left was laser-permeabilized to flavopiridol later
than the embryo on the right was, and chromosome dynamics appear less
affected as expected.
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Video S6. Flavopiridol
Treatment Directly after the Start of Nuclear Envelope Breakdown Resulted Sometimes in
Individual Chromosomes Failing to Associate with a Metaphase Plate by
the Time of Premature Spindle Displacement. As in the previous
video, but this is a treated embryo alone, with an
apparently less severe
defect. One chromosome is not associated with the metaphase plate
during spindle displacement and anaphase. This embryo is also shown as
the center embryo among the flavopiridol-treated embryos in Figure
6A.
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