Cancer:

1
Feedback Control Between S-phase and Mitosis
Feedback control ensures dependency of cell cycle
events
2
DNA Damage Checkpoints
Sensors (i.e. Rad9)
Transducers
Targets
Targets
From Susan Forsburg, Salk Institute
http//pingu.salk.edu/forsburg/cclecture.html
3
Chromosome Movements During Mitosis
From Rieder and Salmon (1998) Trends Cell Biol.
8310
4
What if There Were No Checkpoint?
Lodish et al. Figure 13-33
5
The Mitotic Spindle Assembly Checkpoint Ensures
the Order of Mitotic Events
What has to Happen for Proper Chromosome
Segregation? - Sister Chromatid Cohesion
established in S-phase - Chromosome condensation
in G2 - Mitotic Spindle Assembly in Mitosis -
Chromosome alignment in Mitosis -
Metaphase/Anaphase transition (M/A) -securin
degradation leads to cohesion destruction
Mitotic checkpoint has to be upstream of M/A -
Once you start destroying cohesion theres no
going back
6
The Mitotic Spindle Assembly Checkpoint
aka Kinetochore Checkpoint Mitotic
Checkpoint Metaphase Checkpoint Spindle
Checkpoint
Mitotic Checkpoint Inhibits Cohesion
Destruction -via inhibition of APCCdc20
Mitotic Checkpoint Also Inhibits Mitotic
Exit -via inhibition of APCCdh1
7
The Metaphase to Anaphase (M/A) Transition
From Zhou et al. (2002) J. Cell Sci. 1153547
8
How do We Know Theres a Checkpoint?
Prometaphase
Metaphase
Adapted From Shimoda and Solomon (2002) Cell 1099
9
How do We Know Theres a Checkpoint?
Prometaphase
Metaphase
Regardless of the length of Prometaphase, Metaphse
lasts 20 minutes
Adapted From Shimoda and Solomon (2002) Cell 1099
10
Cohesion is Established in S-phase
From Susan Forsburg, Salk Institute
http//pingu.salk.edu/forsburg/cclecture.html
11
Cohesion is Established in S-phase
Pds5
Scc3
Scc1
Smc1
Smc3
Cohesin
Condensin
From Uhlman (2001) Cur. Op. Cell Biol. 13754
12
Cohesion is Established in S-phase
Cohesin
Condensin
Chromosome Cohesion
Chromosome Compaction
Scissile Subunit (Scc1) Cleaved at M/A
From Uhlman (2001) Cur. Op. Cell Biol. 13754
13
Condensin is Required for Chromosome Compaction
Cohesin is Required for Chromosome Compaction
14
Condensin is Required for Chromosome Compaction
Cohesin is Required for Chromosome Compaction
Cohesin links Condensin from adjacent sister
chromatids
15
From Susan Forsburg, Salk Institute
http//pingu.salk.edu/forsburg/cclecture.html
16
The Metaphase to Anaphase (M/A) Transition
From Zhou et al. (2002) J. Cell Sci. 1153547
17
Evidence for the Existence of a Checkpoint
Needle
20 min.
Micromanipulation of Chromosomes in Grasshopper
Spermatocytes
Adapted From Nicklas (1997) Science 275632
18
Micromanipulation of Chromosomes in Grasshopper
Spermatocytes
From Nicklas (1997) Science 275632
19
Kinetochores Generate the Checkpoint Signal
Adapted From Shimoda and Solomon (2002) Cell 1099
20
Kinetochores Generate the Checkpoint Signal
20 min
Laser Ablate Kinetochore from last Mono-oriented
Chromosome
Adapted From Shimoda and Solomon (2002) Cell 1099
21
From Rieder et al. (1995) J. Cell Biol. 130941
22
Laser
Anaphase
From Rieder et al. (1995) J. Cell Biol. 130941
23
Unattached Kinetochores Cause a Checkpoint Delay
From Musacchio and Hardwick (2002) Nature Reviews
2731
24
Both Attachment and Tension are Monitored
3F3 Epitope Phosphorylation of unknown proteins
-dims upon attachment -disappears upon
tension
From Rieder and Salmon (1998) Trends Cell Biol.
8310
25
Both Attachment and Tension are Monitored
Mono-oriented Grasshopper Spermatocyte Chromosomes
Relaxed
Tension Applied
3F3 Epitope Present in the absence of tension
3F3 Epitope Disappears when tension applied
Adapted From Nicklas (1997) Science 275632
26
One More Yeast Screen MADs and BUBs
Search for Genes Responsible for Feedback Control
of Mitosis
Hoyt Budding Uninhibited by Benomyl Bub1,
Bub2, Bub3
Li Mitotic Arrest Deficient Mad1, Mad2,
Mad3
27
Spindle Assembly Checkpoint Components
28
APC is the Target of the Mitotic Checkpoint
The Metaphase to Anaphase (M/A) Transition
From Zhou et al. (2002) J. Cell Sci. 1153547
29
(No Transcript)
30
Potential Checkpoint Targets
31
APC is the Target of the Mitotic Checkpoint
From Zhou et al. (2002) J. Cell Sci. 1153547
32
APC is the Target of the Mitotic Checkpoint
Cdc2/Cyclin B
Mad2 Mad3/BubR1
Bub2
Cdc2/Cyclin B
Adapted from Peters (2002) Mol. Cell 9931
33
Spindle Assembly Checkpoint Pathways
Securin
Separase
Adapted From Susan Forsburg, Salk Institute
http//pingu.salk.edu/forsburg/cclecture.html
34
Mitotic Exit is a Spindle Checkpoint Target
Bub2 active NO Cdc14 Release
Mitotic Exit Network (MEN)
Bub2 inactive Cdc14 Released
From Susan Forsburg, Salk Institute
http//pingu.salk.edu/forsburg/cclecture.html
35
Bub2 Prevents Cdc14 Release/Cdh1 Activation
Bub2
Bub2
36
APC is the Target of the Mitotic Checkpoint
Cdc2/Cyclin B
Mad2 Mad3/BubR1
Bub2
Cdc2/Cyclin B
Adapted from Peters (2002) Mol. Cell 9931
37
Spindle Assembly Checkpoint Pathways
Securin
Separase
From Susan Forsburg, Salk Institute
http//pingu.salk.edu/forsburg/cclecture.html
38
Checkpoint is Mediated by Rearrangement of Mad2
Complexes
Mad2 Inhibits APCCdc20
From Millband et al. (2002) Trends Cell Biol.
12205
39
Mad2 Cycles Through Kinetochore and Inhibits Cdc20
Shah and Cleveland (2000) Cell 103997
40
(No Transcript)