Cell cycle

(Molecular Biology of The Cell

chapter 17 :

Alberts, et al., Garland

Science, 2008)

Dr. Ratna Megawati Widharna, SKG, MFT

4 phases of the cell cycle

(< 1 hour)

(10-12 hours)

3
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(3)
(2)

(1)

/ restriction point (late G1)

Cell control system

Based on connected series of biochemical switches
initiates specific cell-cycle event
1. binary on/off : launch events in a complete,
irreversible fashion (e.g. chromosome condensation)
2. Robust & reliable backup mechanism & other
feature system operate effectively under a variety
of conditions & even if some components fail
3. highly adaptable & can be modified to suit
specific cell types/ to respond to specific intracellular
/ extracellular signals

Cell-cycle control system depends on

cyclically Activated Cyclin-Dependent
Protein kinases (Cdks)

Cdk activity at the G2/M checkpoint

phosphorylation of protein that control
chromosome condensation
Nuclear envelope breakdown
Spindle assembly
Other events that occur at the onset of mitosis

Cdk + cyclin protein kinase activity

Cyclin-cdk complexes triggers cell-cycle
events

4 classes of cyclins
(1)

(2)

(3)

All eucaryotic cells require 3

of these classes
(1) G1/S-cyclins : activate Cdk in late G1
help trigger progression through Start
commitment to cell-cycle entry their
levels fall in S phase
(2) S-cyclins : bind Cdks soon after
progression through Start help
stimulate chromosome duplication. Scyclin levels remain elevated until mitosis
contribute to control of some early
mitotic events

All eucaryotic cells require 3

of these classes
(3) M-cyclins : activate Cdk that
stimulate entry into mitosis at the
G2/M checkpoint
In most cells, (4) G1-cyclins : helps
govern the activities of the G1/Scyclins which control progression
through Start in late G1.

The structural basis of Cdk activation

 No cyclin active site in the Cdk prot is partly

obscured by a slab of prot (A)
Cyclin binding slab move away from the active
site partial activation of Cdk enzyme (B)
Full activation of the cyclin-Cdk complex occurs
when a separate kinase (CAK/Cdk-activating kinase)
phosphorylates an amino acid near the entrance of
the Cdk active site -> conformational change
Cdk activity kinase phosphorylate its target
prot effectively induce specific cell-cycle events

 Phosphorylation at a pair of amino acids in

the roof on the kinase active site inhibits
the activity of a cyclin-Cdk complex.
Phosphorylation by Wee1 inhibits Cdk
activity, while dephosphorylation of these
sites by a phosphatase (Cdc25) increases
Cdk activity
Binding of Cdk inhibitor proteins (CKIs) also
regulates cyclin-Cdk complexes

 Cyclin-Cdk-CKI complex reveals that CKI binding

stimulates a large rearrangement in the structure
of the Cdk active site, rendering it inactive
Cell use CKIs primarily to help govern the
activities of G1/S and S-Cdks early in the cell
cycle

The cell cycle control

system depends on cyclical
proteolysis
Progression through the metaphase-to-

anaphase-transition is triggered by protein

destruction, leading to the final stages of cell
division
Key regulator : anaphase-promoting complex/
Cyclosome (APC/C)
APC/C catalyzes the ubiquitylation & destruction
of 2 major prot:
Securin
S- and M-cyclins

APC/C catalyzes the ubiquitylation &

destruction of
1. securin = normally protects the prot linkages
that hold sister chromatid pairs together in
early mitosis
Destruction of securin at the metaphase-to-anaphase
transition activates a protease that separates the
sisters & unleashes anaphase

2. The S&M cyclins

Destroying S&M cyclins INACTIVATES most Cdks in the
cell
many prot phosphorylated cy Cdks from S phase to early
mitosis are dephosphorylated by various phosphatase that are
present in the anaphase cell
dephosphorylation : necessary for the completion of M phase
including the final step in mitosis & the process of cytokinesis

 APC/C remains active in G1 a

stable period of the Cdk inactivity
When G1/S-Cdks are activated in late
G1, the APC/C is turned off
allowing cyclin accumulation to start
the next cell cycle

SCF (another ubiquitin

ligase)
Ubiquitylates certain CKI prot in late
G1 helps control the activation of
S-Cdks & DNA replication

Difference between APC/C

and
SCF
APC/C activity changes during the cell cycle result

of changes in its association with an activating

subunit-either Cdc20 during anaphase/Cdh1 from late
mitosis through early G1. These subunits helps the
APC/C recognize its target protein
SCF activity depends on subunits F-box prot, which
help the complex recognize its target prot. Unlike
APC/C activity, SCF activity is CONSTANT during the
cell cycle. Ubiquitylation by SCF is controlled by
changes in the phosphorylation state of its target
prot, as F-box subunits recognize only specifically
phosphorylated prot

Major Cell-Cycle Regulatory

Proteins

Start
checkpoints

Chromosome duplication
Early event in mitosis

M-Cdk activation
progress G2/M checkpoint
Events of early mitosis
Leading to alignment of sister chromatids
at the equator of the mitotic spindle
Finally the APC/C + Cdc20 (activator)
triggers the destruction of securin &
cyclins at the metaphase-to-anaphase
transition unleasing sister-chromatid
segregation & completion of mitosis

S phase
2 problems must be solved when
initiating & completing DNA
replication:
1. replication must occur with extreme
accuracy ( the risk of mutations in the
next generation)
2. every nucleotide in the genome must be
copied once and only once, to prevent the
damaging
effects of gene amplification

To ensure that chromosome duplication

occurs only once/cell cycle, the initiation of
DNA replication is divided into 2 distinct
steps

1. In Late mitosis & early G1 when a large complex of initiator

protein = PREREPLICATIVE COMPLEX (Pre-RC) assembles at
origins of replication = LICENSING OF REPLICATION
ORIGINS initiation of DNA synthesis is permitted only at
origins containing a pre-RC
2. ONSET of S phase when components of the pre-RC nucleate
the formation of a larger protein complex = PREINITIATION
COMPLEX unwinds the DNA helix & loads DNA
polymerases & other replication enzymes onto the DNA
Strands initiating DNA synthesis pre RC is dismantled
& cannot be reassembled at the origin until the following
G1 origins can be activated only once/cell cycle

Assembly of the pre-RC

Inhibited by Cdk activity
Stimulated by APC/C
In late mitosis & early G1
Cdk activity
APC/C

At the onset of S-phase activation of

S-Cdk triggers the formation of a
preinitiation complex initiates DNA
synthesis

Pre-RC is partly dismantled

S-Cdk
M-Cdk
APC/C
Until late mitosis, new pre-RCs
cannot be assembled at fired origins
until the cell cycle is complete

ORC binds to replication

origins throughout the cell
cycle
In late mitosis and early G1, the protein

Cdc6 & Cdt1 bind to the ORC at origins &

help load a group of 6 related prot = Mcm
prot complex pre-RC & the origin is now
licensed for replication
The 6 Mcm prot of the pre-RC form a ring
around DNA serve as the major DNA
helicase that unwinds the origin DNA when
DNA synthesis begins & as the replication
forks move out from the origin

 Central purpose of pre-RC: to load the helicase

that will play a central part in the subsequent
DNA replication process
Once the pre-RC has assembled in G1, the
replication origin is ready to fire
The activation of S-Cdk in late G1 triggers the
assembly of several additional prot complexes
at the origin, leading to the formation of a
giant preinitiation complex that unwinds the
helix & begins DNA synthesis

 At the same time it initiates DNA replication,

S-Cdk triggers the disassembly of some preRC components at the origin
Cdks phosphorylates ORC & Cdc6
inhibition
Inactivation of the APC/C in late G1 also
helps turn off pre-RC assembly
In late mitosis and early G1, the APC/C
triggers the destruction of a prot GEMININ,
that binds and inhibits the pre-RC
component Cdt1
When APC/C is turned off in late G1,
GEMININ accumulates and inhibits Cdt1

 At the end of mitosis, APC/C

activation inactivation of Cdks and
destruction of geminin pre-RC
components are dephosphorylated
and Cdt1 is activated allowing preRC assembly to prepare the cell for
the S phase

Cohesins help hold sister

chromatids together
2 of the subunits of cohesin are
members of SMC proteins (Structural
Maintenance of Chromosomes)
Cohesin forms giant ring-like
structures

Mitosis part 1
An abrupt in M-Cdk activity at the
G2/M checkpoint triggers the events
of early mitosis (prophase,
prometaphase, and metaphase). MCdk and several other mitotic protein
kinase phosphorylate a variety of
proteins, leading to the assembly of
the mitotic spindle & its attachment
to the sister chromatid pairs

Mitosis part 2
The 2nd major part of mitosis begins at the
metaphase-to-anaphase transition, when the APC/C
triggers the destruction of securin, liberating a
protease that cleaves cohesin initiates
separation of the sister chromatids
The APC/C also triggers the destruction of cyclins
Cdk inactivation and the dephosphorylation of Cdk
targets required for all events of late M phase ,
including completion of anaphase, disassembly of
the mitotic spindle, & division of cell by cytokinesis

Dephosphorylation activates
M-Cdk at the onset of
mitosis
M-Cdk activation begins with the
accumulation of M-cyclin
In embryonic cell cycles, the synthesis of MCyclin is constant throughout the cell cycle,
and M-cyclin accumulation results from the
high stability of the protein in interphase
In most cell types, M-cyclin synthesis
increases during G2 and M, owing primarily
to an increase in M-cyclin gene transcription

 The increase in M-cyclin protein leads to a

corresponding accumulation of M-Cdk (the
complex of Cdk1 and M-cyclin) as the cell
approaches mitosis
Although the Cdk in these complexes is
phosphorylated at an activation site by
the Cdk-activating kinase (CAK), the
protein kinase Wee1 holds it in inactive
state by inhibitory phosphorylation at 2
neighbouring sites

 Thus, by the time the cell reaches

the end of G2, it contains an
abundant stockpile of M-Cdk that is
primed and ready to act but is
suppressed by phosphates that block
the active site of kinase

What triggers the activation

of the M-Cdk stockpile?
Activation of protein phosphatase Cdc25,
which removes the inhibitory phosphatases
that restrain M-Cdk. At the same time, the
inhibitory activity of the kinase Wee1 is
suppressed, further ensuring that M-Cdk
activity increases. The mechanism that
unleash Cdc25 activity (and supress Wee1)
in early mitosis are not well understood. One
possibility is that the S-Cdks that are active
in G2 and early prophase stimulate Cdc25

 Interestingly, Cdc25 can also be activated, at least in

part, by its target, M-Cdk. M-Cdk may also inhibit the
inhibitory kinase Wee1. The ability of M-Cdk to activate
its own activator (Cdc25) and inhibits its own inhibitor
(Wee1) suggests that M-Cdk activation in mitosis
involves positive feedback loops
According to this model, the partial activation of Cdc25
(perhaps by S-Cdk) leads to partial activation of a
subpopulation of M-Cdk complexes, which then
phosphorylate more Cdc25 and Wee1 molecules. This
leads to more M-Cdk activation, and so on. Such
mechanism would quickly promote the complete
activation of all the M-Cdk complexes in the cell

Condensin helps configure

duplicated chromosomes for
separation

Related to that of cohesin complex

(holds sister chromatid together)
Contains 2 SMC subunits like those of
cohesin + 3 non-SMC subunits
Condensin may form a ring-like
structure that somehow uses the
energy provided by ATP hydrolysis to
promote the compaction & resolution
of sister chromatids

Condensin
Able to change the coiling of DNA
molecules in a test tube important for
chromosome condensation during
mitosis
Phosphorylation of condensin subunits
by M-Cdk stimulates this coiling activity,
providing 1 mechanism by which M-Cdk
may promote chromosome restructuring
in early mitosis

The mitotic spindle is a

microtubule-based machine
Chromosome segregation depends on
mitotic spindle
The minus end of which are focused at the
2 spindle poles, and the plus end of which
radiate outward from the poles
The plus end of microtubules =
INTERPOLAR MICROTUBULES interact with
the plus ends of microtubules from the
other pole, resulting in an antiparallel array
in the spindle midzone

 The plus ends of other microtubules-the

kinetochore microtubules are attached
to sister chromatid pairs at large protein
structures called KINETOCHORES, which
are located at the centromere of each
sister chromatid
Finally, many spindles also contain astral
microtubules that radiate outward from
the poles and contact the cell cortex,
helping to position the spindle in the cell

 In most somatic animal cells, each spindle

pole is focused at centrosome
Each centrosome consists of pericentriolar
matrix that surrounds a pair of centrioles
The pericentriolar matrix nucleates a
radial array of microtubules, with their
fast-growing plus ends projecting outward
and their minus ends assoc with the
centrosome

The matrix contains

A variety of proteins, including
Microtubule-dependent motor proteins
coiled-coil proteins that link the motors
to the centrosome
Structural proteins
Components of the cell-cycle control
system
-tubulin ring complex responsible for
nucleating microtubules

Microtubule-dependent motor
proteins govern spindle assembly
and function

Kinesin-related prot : move toward the plus

end of microtubules
Dynein : move toward the minus end of
microtubules
Kinesin-5 protein: contains 2 motor
domains that interact with plus ends of
antiparallel microtubules in the spindle
midzone slide the 2 antiparallel
microtubules past each other toward the
spindle poles, forcing the poles apart

 Kinesin-14 protein: minus end directed motors

with a single motor domain & other domains that
can interact with a different microtubule. They
can cross-link antiparallel microtubules at the
spindle midzone and tend to pull the poles
together
Kinesin 4 & kinesin 10 : chromokinesin plus end
directed motors associated with chromosome
arms and push the attached chromosome away
from the pole (/ the pole away from chromosome)

 Dynein : minus end directed-motors

together with associated protein: organize
microtubules at various cellular locations
They link the plus ends of astral
microtubules to components of the actin
cytoskeleton at the cell cortex, e.g. by
moving toward the minus end of the
microtubules, the dynein motors pull the
spindle poles toward the cell cortex and
away from each other

2 mechanisms collaborate in the

assembly of a bipolar mitotic
spindle

1. Depends on the ability of mitotic

chromosomes to nucleate and
stabilize microtubules and on the
ability of the various motor proteins
to organize microtubules into a
bipolar array, with minus ends
focused at 2 spindle poles and plus
ends interacting with each other in
the spindle midzone

2 mechanisms collaborate in the

assembly of a bipolar mitotic
spindle

2. depends on the ability of

centrosomes to help form the spindle
poles
Each of the pair of centrosomes
nucleates a radial array of
microtubules
The 2 centromeres facilitate bipolar
spindle assembly by providing a pair of
fabricated spindle poles

Centrosomes duplication
occurs early in the cell cycle
Centrosome duplication begins at about the same
time as the cell enters S phase
The G1/S-Cdk (complex of cyclin E and Cdk2 in
animal cells) that triggers cell cycle entry also
initiates centrosome duplication
The 2 centrioles in the centrosome separate, and
each nucleates the formation of a single new
centriole 2 centriole pairs within an enlarged
pericentrilar matrix. This centrosome pair remains
together on 1 side of the nucleus until the cell enters
mitosis

Centrosome ~ chromosome
duplication
Use a semi-conservative mechanism
of duplication 2 halves separate
and serve as templates for
construction of a new half
Must replicate once and only once
per cell cycle to ensure that the
cell enters mitosis with only 2 copies

M-Cdk initiates spindle

assembly in prophase
M-Cdk activity initiates spindle assembly
2 centrosomes move apart along the
nuclear envelope & the plus ends of the
microtubules between them interdigitate to
form the interpolar microtubules of the
developing spindle
The amount of tubulin ring complexes in
each centrosomes greatly, the ability of
centrosomes to nucleate new microtubules
= CENTROSOMES MATURATION

M-Cdk initiates spindle

assembly in prophase (2)
Minus end directed dynein motor protein at
the plus ends of astral microtubules provide
the major force these motors are anchored
at the cell cortex/on the nuclear envelope and
their movement toward the microtubule
minus end pulls the centrosome apart
Interaction between the centrosomal
microtubules & the cell cortex allow actinmyosin bundles in the cortex to pull the
centrosomes further apart

 Finally, kinesin 5 motors cross-link the

overlapping, antiparralel ends of interpolar
microtubules and push the poles apart
Dynein & kinesin-5 motors : centrosome
separation and spindle length
Kinesin 14 protein : minus end directed
motors and interact with a microtubule from
1 pole while traveling toward the minus end
of an antiparallel microtubule from the other
pole they tend to pull the poles together

 M-Cdk and Aurora A phosphorylate

kinesin-5 motors and stimulate them
to drive centrosome separation
Aurora-A and Plk also phosphorylate
components of the centrosome
promote its maturation

The completion of spindle assembly in

animal cells requires nuclear envelope
breakdown
Begin when M-Cdk phosphorylates several subunits
the giant nuclear pore complexes in the nuclear
envelope
Initiates the disassembly of nuclear pore complexes
and their dissociation from the envelope
M-Cdk also phosphorylates components of the
nuclear lamina, the structural framework that lies
beneath the envelope leads to disassembly of
the nuclear lamina & the breakdown of the
envelope membranes into small vesicles

Microtubules instability
increases greatly in mitosis
(interphase)
Dinamyc instability:
Growth shrinkage : Catastrophe
Shrinkage growth : Rescue
New microtubules are continually being
created to balance the loss of those that
disappear completely by depolymerization
Long microtubules a >>> # of shorter
& more dynamic microtubules surrounding
each centromere

Prometaphase

T1/2 microtubules

Metaphase
dramatically
In microtubule instability
Dense and dynamic
array of spindle
Ability of centrosomes to
microtubules that are
ideally suited for
nucleate microtubule
capturing sister
chromatids

M-Cdk initiates these changes by phosphorylating 2

classes of protein that control microtubule dynamics
these contains microtubule-dependent motor
protein and microtubule-associated protein (MAPs)

2 classes of protein govern

microtubule dynamics in
mitosis

CATASTROPHE Factors : destabilize

microtubule arrays by frequency of
catastrophes. One of these protein :
kinesin-related protein that does not
function as a motor
MAP : stabilizing microtubules :
frequency of rescues shrinkage to
growth, growth rate, shrinkage
rate of microtubules

Bi-orientation is achieved by
trial and error
Sister kinetochores are constructed in a
back-to back orientation that reduces the
likelihood that both kinetochores can face
the same spindle pole
Incorrect attachment are highly unstable
while correct attachments are locked in place
Tension when a sister chromatid pair is
properly bi-oriented on the spindle, the 2
kinetochores are pulled in opposite directions
by strong poleward forces

 Sister-chromatid cohesion resists

these poleward forces high level of
tension within kinetochores
High tension at the kinetochore shuts
off the inhibitory signal,
strengthening microtubule
attachment formation a thick
kinetochore fiber composed of
multiple microtubules

Aurora B
Generate the inhibitory signal that
reduces the strength of microtubule
attachment in the absence of tension
It phosphorylates several components
of the microtubule attachment site,
sites affinity for a microtubule plus end
Inactivated when bi-orientation occurs
kinetochore phosphorylation and
affinity of the attachment site

The APC/C triggers sisterchromatid separation and the

completion of mitosis

After M-Cdk has triggered the complex

rearrangements that occur in early mitosis, the cell
cycle reaches its climax with the separation of the
sister chromatids at the metaphase-to-anaphase
transition
Although M-Cdk activity sets the stage for this
event, the anaphase-promoting complex (APC/C)
throws the switch that initiates sister-chromatid
separation by ubiquitylating several mitotic
regulatory proteins and thereby trigerring their
destruction

The APC/C triggers sisterchromatid separation and the

completion of mitosis

During metaphase, cohesins holding the

sister chromatids together resist the
poleward forces that pull the sister
chromatid apart. Anaphase begins with a
sudden disruption of sister-chromatid
cohesion, which allows the sisters to
separate and mote to opposite poles of the
spindle
The APC/C initiates the process by targeting
the inhibitory protein securin for destruction.

 Before anaphase, securin binds to and

inhibits the activity of a protease, called
separase
The destruction of securin at the end of
metaphase releases separase which is then
free to cleave one of the subunits of
cohesin
The cohesin fall away and the sister
chromatids abruptly and synchronously
separate

 In addition to securin, the APC/C also

targets the S and M-cyclins for
destruction, leading to the loss of
most Cdk activity in anaphase
Cdk inactivation allows phosphatases
to dephosphorylates the many Cdk
target substrates in the cell, as
rewuired for the completion of mitosis
and cytokinesis

The APC/C triggers sisterchromatid separation and the

completion of mitosis

If the APC/C triggers anaphase, what

activates the APC/C?
APC/C activation requires the protein
Cdc20, which binds and activates the
APC/C in mitosis
At least 2 processes regulate Cdc20
and its association with the APC/C

 1. Cdc20 synthesis increases as the cell

approaches mitosis, owing to an increase
in the transcription of its gene
2. Phosphorylation of the APC/C helps
Cdc20 kinases that phosphorylates and
thus activate the APC/C is M-Cdk. M-Cdk
is not only triggers the early mitotic
events leading up to metaphase, but it
also sets the stage for progression into
anaphase

 The ability of M-Cdk to promote

Cdc20/APC/C activity creates a
negative feedback loop: M-Cdk sets
in motion a regulatory process that
leads to cyclin destruction and thus
its own activation

 During metaphase, cohesins holding the

sister chromatids together resist the
poleward forces that pull the sister
chromatids apart.
Anaphase begins with a sudden disruption
of sister-chromatid cohesion, which allows
the sisters to separate and move to opposite
poles of the spindle
The APC/C initiates the process by targeting
the inhibitory protein securin for destruction

 Before anaphase, securin binds to

and inhibits the activity of a protease
called separase
The destruction of securin at the end
of metaphase releases separase,
which is then free to cleave one of
the subunits of cohesin. The cohesin
falls away and the sister chromatids
abruptly and synchronously separate

Unattached chromosomes block sisterchromatid separation : The spindle

assembly checkpoint
Spindle assembly checkpoint
mechanism that is activated by drug
treatment and blocks progression
through the metaphase-to-anaphase
transition
The checkpoint mechanism ensures
that cells do not enter anaphase until
all chromosomes are correctly bioriented on the mitotic spindle

 Any kinetochore that is not properly attached

to the spindle sends out a negative signal that
blockes Cdc20-APC/C activation and thus
blocks the metaphase-to-anaphase transition.
Only when the last kinetochore is properly
attached is this block removed, allowing sisterchromatid separation to occur
Inappropriately attached kinetochores
somehow generate a diffusible signal that
inhibits Cdc-20-APC/C activity throughout the
cell

 Several proteins, including Mad2 are recruited

to unattached kinetochores and are required for
spindle assembly checkpoint to function
Unattached kinetochore acts like an enzyme
that catalyzes the change in the conformation
of Mad2 so that Mad2 can bind and inhibit
Cdc20-APC/C
The destruction of securin in mamalian somatic
cells begins moments after the last sister
chromatid pair becomes bi-oriented on the
spindle and anaphase begins 20 minutes later

Chromosomes segregate in
anaphase A and B
Sudden loss of sister-chromatid cohesion
at the onset of anaphase sisterchroatied separation allows forces of the
mitotic spindle to pull the sisters to
opposite poles of the cell = CHROMOSOME
SEGREGATION
Chromosomes move by 2 independent and
overlapping processes:
Anaphase A
Anaphase B

Anaphase A
Initial poleward movement of the chromosomes
Accompanied by shortening of kinetochore
microtubules
Chromosome movement depends on the
combination of 2 major poleward force
1. Force generated by microtubule
depolymerization at the kinetochore results
in loss of tubulin subunits at the plus end as the
kinetochore moves toward the pole
2. Provided by microtubule flux, which is the
poleward movement of the microtubules toward
the spindle pole, where minus end
depolymerization occurs

Anaphase B
Separation of the spindle poles themselves, which begins
after the sister chromatids have separated and the
daughter chromosomes have moved some distance
apart
Spindle pole separation depends on motor-driven
mechanism
Plus end directed kinesin 5 motor proteins, which crosslinke 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

 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 mutatnt form that is not
recognized by APC/C, e.g. sister-chromatid
separation generally occurs but the
chormosome movements and microtubule
behaviour of anaphase are abNormal

Segregated chromosomes are

packaged in daughter nuclei at
telophase
2 sets of chromosome are packaged
into a pair of daughter nuclei
1st major event : disassembly of the
mitotic spindle, followed by reformation of the nuclear envelope
Initially, nuclear membrane
fragments associate with the surface
of individual chromosomes

 These membrane fragments fuse to partly

enclose clusters of chromosomes and then
coalesce to re-form the complete nuclear
envelope
Nuclear pore complexes are incorporated
into the envelope
The nuclear lamina re-forms
And the envelope once again become
continuous with the endoplasmic reticulum

 Once the nuclear envelope has reformed, the pore complexes pump in
nuclear proteins, the nucleus expands
and the condensed mitotic chromosomes
are reorganized into their interphase
state, allowing gene transcription to
resume. A new nucleus has been created,
and mitosis is complete. All that remains
is for the cell to complete its division into
2.

 Earlier : phosphorylation of various proteins

by M-Cdk promotes spindle assembly,
chromosome condensation and nuclear
envelope break-down in early mitosis
Dephosphorylation of these same proteins is
required for spindle disassembly and the reformation of daughter nuclei in telophase
These dephosphorylation and the completion
of mitosis could be triggered by inactivation of
Cdks, the activation of phosphatases, or both

 Although Cdk inactivation-resulting

primarily from cyclin destruction-si
mainly responsible in most cells,
some cells also rely on activation of
phosphatases

meiosis
Loss of arm cohesion in meiosis I
depends on APC/C activation leads to
securin destruction , separase activation
and cohesin cleavage along the arms
In contrast to mitosis, cohesin complexes
near the centromeres remain uncleaved
in meiosis I because cohesin in that
region is protected from separase

 Sister-chromatid pairs therefore remained

linked at their centromeres throughtout
meiosis I, allowing their correct biorientation on the spindle in meiosis II
The mechanisms that block cohesin
cleavage at the centromere in meiosis I are
removed in meiosis II
At the onset of anaphase II, APC/C activation
therefore triggers centromeric cohesin
cleavage and sister-chromatid separation

Cytokinesis
The final step in the cell cycle = division of cytoplasm
1st visible change : sudden appearance of a
pucker/cleavage furrow, on the cell surface
The furrow rapidly deepens and spreads the arround
the cell until it completely divides the cell in 2
The process underlying this process: contractile ring-a
dynamic assembly composed of actin filaments,
myosin II filaments, and many structural and
regulatory proteins
During anaphase, the ring assembles just beneath
the plasma membrane

 The ring gradually contracts, and, at the same time,

fusion of intracellular vesicles with the plasma
membrane inserts new membrane adjacent to the ring
This addition of membrane compensates for the
increase in surface area that accompanies cytoplasmic
division
When ring contraction is completed, membrane
insertion and fusion seal the gap between the
daughter cells
Thus, cytokinesis can be considered to occur in 4
stages : initiation, contraction, membrane insertion
and completion

Local activation of RhoA triggers

assembly and contraction of the
contractile ring

RhoA, a small GTPase of the Ras superfamily

controls the assembly and function of the
contractile ring at the site of cleavage
RhoA is activated at cell cortex at the future
division site, where it promotes actin filament
formation, myosin II assembly, and ring
contraction. It promotes actin filament
formation by activating formins, and it
promotes myosin II assembly and contractions
by activating multiple protein kinase, including
the Rho-activated kinase Rock

 These kinases phosphorylate the

regulatory myosin light chain (RMLC),
which is one of the subunits of
myosin II
Phosphorylation of the RMLC
stimulates bipolay myosin II filament
formation and motor activity, thereby
promoting the assembly and
contraction of the actin-myosin ring

 Like other GTPases, RhoA is inactive when

bound to GDP and active when bound to
GTP
The local activation of RhoA at the
cleavage furrow is though to depend on a
Rho guanine nucleotide exchange factor
(RhoGEF), which is found at the cell cortex
at the future division site and stimulates
the release of GDP and binding of GTP to
RhoA

The microtubules of the mitotic

spindle determine the plane of
animal cell division

Cytokinesis must occur only after the 2

sets of chromosomes are fully
segregated from @ other, and the site of
division must be placed between the 2
sets of daughter chromosomes
ensuring that @ daughter cell receives a
complete set
Correct timing & positioning of
cytokinesis depend on mitotic spindle

Cytokinesis occurs at the

correct time
During anaphase, the spindle generates
signals that initiate furrow formation at a
position midway between the spindle poles
ensuring that division occurs between the 2
sets of separated chromosomes
contributes to correct timing of cytokinesis in
late mitosis
Dephosphorylation of Cdk substrates, which
depends on cyclin destruction in metaphase
and anaphase initiates cytokinesis

How does the mitotic

spindle specify the site of
division?
1. Astral stimulation model : postulates
that the astral microtubules carry furrowinducing signals to the cell cortex, where
they are somehow focused into a ring
halfway between the spindle poles
experiment in large embryonic cells
cleavage furrow forms midway between 2
aster, even when the 2 centrosomes
nucleating the asters are not connected to
each other by a mitotic spindle (Fig 17-54)

How does the mitotic

spindle specify the site of
division?
2. Central spindle stimulation model :

spindle midzone/central spindle

generates a furrow-inducing signal that
specifies the site of furrow formation at
the cell cortex
The overlapping interpolar microtubules
of the central spindle associate with
numerous signaling proteins, including
proteins that may stimulate RhoA

How does the mitotic

spindle specify the site of
division?

3. astral microtubules promote the

local relaxation of actin-myosin
bundles at the cell cortex
Cortical relazation is minimal at the
spindle equator promoting cortical
contraction at that site

Membrane-enclosed organelles must

be distributed to daughter cell during
cytokinesis
How do the various membrane-enclosed
organelles segregate when a cell divides?
Organelles e.g mitochondria and
chloroplasts are usu present in large
enough number sto be safely inherited
The ER in interphase cell is continuous
with the nuclear membrane and is
organized by the microtubule
cytoskeleton

 Upon entry into M phase, the reorganization of the

microtubules and breakdown of the nuclear
envelope releases the ER.
ER remains largely intact and is cut in 2 during
cytokinesis
The Golgi apparatus is reorganized and fragmented
during mitosis. Golgi fragments associate with the
spindle poles & r thereby distributed to opposite
ends of the spindle, ensuring that @ daughter cell
inherits the materials needed to reconstruct the
Golgi in telophase

Some cells reposition their

spindle to divide
asymmetrically

Most animal cells divide symmetrically :

the contractile ring forms around the
equator of the parent cell, producing 2
daughter cells of equal size and with
the same components
This symmetry results from the
placement of the mitotic spindle, which
in most cases tends to center itself in
the cytoplasm

Mitosis can occur without

cytokinesis

The G1 phase is a stable

state of Cdk inactivity
Key regulatory event in late M phase : inactivation
of Cdks, which is driven primarily by APC//C
dependent cyclin destruction
Inactivation of Cdks in late M phase has many
functions :
It triggers the events of late mitosis
Promotes cytokinesis enables the synthesis of
prereplicative complexes at DNA replication origins
Provides a mechanism for resetting the cell-cycle
control system to a state of Cdk inactivity as the
cell prepares to enter a new cell cycle

 In most cells, this state of Cdk inactivity generates

a G1 gap phase, during which the cell grows and
monitors its environment before committing a new
division
The destruction of M-cyclin in late mitosis soon
leads to inactivation of all APC/C activity in an
embryonic cell
This APC/C inactivation immediately after mitosis
is especially useful in rapid embryonic cell cycles,
as it allows the cell to quickly begin accumulating
new M-cyclin for the next cycle (Fig 17-60A)

 Rapid cyclin accumulation

immediately after mitosis is not
useful, for cells with cell cycles
containing a G1 phase
Mechanism 1 : use another APC/C
activating protein, called Cdh1, a
close relative of Cdc20

Cell Cycle
Zack Cook
Start the Tour

Prophase
Metaphase
Interphase

Anaphase
Cytokinesis
Telophase

End Show

Interphase
Cells Mature
Chromosomes copy themselves

Back

Prophase
Nucleur Membrane Disappears
Spindle Fibers Form at the cells
poles

Back

Metaphase
Chromosomes line up in the middle
of cells
2 spindle fibers attach to each of the
23 chromosome pairs

Back

Anaphase
Chromosome pairs split
Spindle Fibers pull chromosomes to
each pole

Back

Telophase
2 nuclei form at poles
Mitosis complete
Membrane pinches in, dividing the
cell

Back

Cytokinesis
2 new daughter cells are formed

Back

Mitotic chromosomes
promote bipolar spindle
assembly

Kinetochores attach sister

chromatids to the spindle