Molecular Biology of the Cell by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morgan, Martin Raff, Keith Roberts, Peter Walter by by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morg

994 Chapter 17: The Cell Cycle
metaphase-to-anaphase transition. When the last sister-chromatid pair is properly
bi-oriented, this block is removed, allowing sister-chromatid separation to
occur.
The negative checkpoint signal depends on several proteins, including
Mad2, which are recruited to unattached kinetochores (Figure 17–39). Detailed
structural analyses of Mad2 suggest that the unattached kinetochore acts like
an enzyme that catalyzes a change in the conformation of Mad2, so that Mad2,
together with other proteins, can bind and inhibit Cdc20–APC/C.
In mammalian somatic cells, the spindle assembly checkpoint determines
the normal timing of anaphase. The destruction of securin in these cells begins
moments after the last sister-chromatid pair becomes bi-oriented on the spindle,
and anaphase begins about 20 minutes later. Experimental inhibition of the
checkpoint mechanism causes premature sister-chromatid separation and anaphase.
Surprisingly, the normal timing of anaphase does not depend on the spindle
assembly checkpoint in some cells, such as yeasts and the cells of early frog
and fly embryos. Other mechanisms, as yet unknown, must determine the timing
of anaphase in these cells.
Chromosomes Segregate in Anaphase A and B
The sudden loss of sister-chromatid cohesion at the onset of anaphase leads to sister-chromatid
separation, which allows the forces of the mitotic spindle to pull the
sisters to opposite poles of the cell—called chromosome segregation. The chromosomes
move by two independent and overlapping processes. The first, anaphase
A, is the initial poleward movement of the chromosomes, which is accompanied
by shortening of the kinetochore microtubules. The second, anaphase B, is the
separation of the spindle poles themselves, which begins after the sister chromatids
have separated and the daughter chromosomes have moved some distance
apart (Figure 17–40).
Chromosome movement in anaphase A depends on a combination of the two
major poleward forces described earlier. The first is the force generated by microtubule
depolymerization at the kinetochore, which results in the loss of tubulin
subunits at the plus end as the kinetochore moves toward the pole. The second
is provided by microtubule flux, which is the poleward movement of the microtubules
toward the spindle pole, where minus-end depolymerization occurs. The
relative importance of these two forces during anaphase varies in different cell
types: in embryonic cells, chromosome movement depends mainly on microtubule
flux, for example, whereas movement in yeast and vertebrate somatic cells
results primarily from forces generated at the kinetochore.
Spindle-pole separation during anaphase B depends on motor-driven mechanisms
similar to those that separate the two centrosomes in early mitosis. Plusend
directed kinesin-5 motor proteins, which cross-link the overlapping plus
ends of the interpolar microtubules, push the poles apart. In addition, dynein
motors that anchor astral microtubule plus ends to the cell cortex pull the poles
apart (see Figure 17–25).
Although sister-chromatid separation initiates the chromosome movements
of anaphase A, other mechanisms also ensure correct chromosome movements
in anaphase A and spindle elongation in anaphase B. Most importantly,
the completion of a normal anaphase depends on the dephosphorylation of Cdk
substrates, which in most cells results from the APC/C-dependent destruction of
cyclins. If M-cyclin destruction is prevented—by the production of a mutant form
that is not recognized by the APC/C, for example—sister-chromatid separation
generally occurs, but the chromosome movements and microtubule behavior of
anaphase are abnormal.
The relative contributions of anaphase A and anaphase B to chromosome segregation
vary greatly, depending on the cell type. In mammalian cells, anaphase
B begins shortly after anaphase A and stops when the spindle is about twice its
metaphase length; in contrast, the spindles of yeasts and certain protozoa primarily
use anaphase B to separate the chromosomes at anaphase, and their spindles
elongate to up to 15 times their metaphase length.
Figure 17–39 Mad2 protein on
unattached kinetochores. This
fluorescence micrograph shows a
mammalian cell in prometaphase, with
the mitotic spindle in green and the sister
chromatids in blue. One sister-chromatid
pair is attached to only one pole of the
spindle. Staining with anti-Mad2 antibodies
indicates that Mad2 is bound to the
kinetochore of the unattached sister
chromatid (red dot, indicated by red
arrow). A small amount of Mad2 is
associated with the kinetochore of the
sister chromatid MBoC6 that m17.45/17.39
is attached to the
spindle pole (pale dot, indicated by white
arrow). (From J.C. Waters et al., J. Cell Biol.
141:1181–1191, 1998. With permission
from the authors.)