Overview of Cell Cycle

1
Overview of Cell Cycle

• Interphase G1, S, and G2 phase
• (The end of cell division to The start of
  next cell division)
• Mitosis M phase
• (The period of cell divided into two
  daughter cells)

2
Synthesis at S Phase

• S phase synthetic period
• (DNA is specifically largely replicated)
• (Genetic information is duplicated)
• Interphase
• (Continuous increase of RNA protein)

3
Commitment to Cell Division

• G1 phase START
• (Restriction/ Commitment point beyond S
  phase)
• (Ensure the ability of cell to replicate
  chromosome)
• G2 phase
• (Restriction/ Commitment point beyond
  Mitosis phase)
• (Ensure the ability of cell to divide)
• G1 gt G2

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Orderly Progress of Cell Cycle

• Make a decision
• G1 restriction point
• (Nutrient supply is enough?)
• (Cell mass is sufficient to support
  division?)
• S phase
• (Replication apparatus is ready to
  synthesis DNA? )
• (The newly synthesized DNA is complete?)
• M phase
• (The newly synthesized DNA is complete?)
• (Duplicated chromosome is properly
  segregated?)

Checkpoint A control loop that makes the
initiation of one event in the cell cycle
dependent on the successful completion of earlier
event.
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• Checkpoints at Cell Cycle
• S phase
• Loss of the integrity of the DNA
• (DNA damage break)
• Mitosis phase
• Incompletion of chromosome segregation
• (Unattached kinetochore)

6
Controlled Events of Cell Cycle

• Regulatory Cyclical Events Control Transitions
  between phases
• The newly synthesized of proteins/ the
  degradation of existed proteins
• Activation/ inactivation of molecules
• G1 phase
• Phosphorylation of RB
• S phase
• S phase activator/ protein kinase
• M phase
• M phase protein kinase

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Identify S phase Inducer

• Cell Fusion Experiment
• Characterizes the activators of each phases
• Mix the cells in the presence of chemical of
  viral agents
• The hybrid cell contains two nuclei in a common
  cytoplasm (heterokaryon)

8
Identify M phase Inducer (MPF)

• Xenopus laevis oocytes exp
• Arrested oocytes (G2 phase)
• ? Ovulation
• ? Arrested egg (M phase)
• The cytoplasm of arrested egg can cause the
  arrested oocytes to enter meiosis
• Maturation promoting factor (MPF)
• (Disaggregation of nuclear envelope)
• (Condensation of chromosome)
• (Spindle formation)
• (Phosphorylation a variety of proteins ?
  kinase)

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MPF is the Cdk/Cyclin

• MPF
• Cdc2 a catalytic subunit of serine/theronine
  kinase activity
• Cyclin a regulatory subunit promotes Cdc2
  phosphorylated appropriate substrates
• Cyclin activates Cdc2 by a binding-induced
  conformational change
• Two Forms of MPF
• Cdc2-cyclin A
• Cdc2-cyclin B
• Regulation of MPF
• Cdc2 constantly expressed/ Cyclin is degraded
• Cdc2 is modified by kinase (p)
• Cyclin is regulated by proteolysis
• Cdc2 is modified by phosphatase (-p)

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Yeast Is a Model System for Analyzing Cell Cycle
S. pombe lengthens and then divides with a
septum, while S. cerevisiae buds during a cycle
in which G2 is absent and M occupies the greatest
part.
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Identify Cell Cycle Mutants in S. pombe
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Cdc Genes in the Cell Cycle of S. pombe
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Different Forms of Cdc2-Cyclin in the Stages of
Cell Cycle

• M phase
• Cdc2/Cdc13 (Try-15 p Thr-161 p)
• At late mitosis Cdc13 is degraded
• START
• Cdc2/Cdc13 (Try-15 p Thr-161 p)
• S to M phase
• Cdc2/Cdc13 is activated (Try-15 p Thr-161 p)
  by dephosphorylation

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Identify Cell Cycle Mutants in S. cerevisiae
Haploid yeast cells of either a or a mating type
may reproduce by a mitotic cycle. Cells of
opposite type may mate to form an a/a diploid.
The diploid may sporulate to generate haploid
spores of both a and a types.
Cdc28 is the Cdc2 of S. cerevisiae
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Cell Cycle of S. cerevisiae

• Chromosome cycle
• Chromosome duplication
• G1-S phase
• Cdc8 mutant
• Centrosome cycle
• Spindle pole body segregation
• Organize microtubules to allow chromosome
  segregation
• Cdc31 mutant
• Cytoplasmic phase
• Nuclear migration into the bud
• Cdc24 mutant

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Cell Cycle Control in S. cerevisiae
17
The Cdc25 Phosphatase Wee1 Kinase Control Cdc2
Activity

• Cdc25
• Phosphatase
• Remove inhibitory phosphate from Tyr-15 of Cdc2
• Wee1
• Kinase
• Phosphorylate Tyr-15 of Cdc25

S. pombe
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Cdc18 Controls S phase Rum1 Inactivates
MPF
S. cerevisiae
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DNA Damage Triggers a Checkpoint
Prevents a defect in genetic information Blocks
the cell cycle progression Damaged DNA is
repaired DNA damage ? ATM kinase ?
phosphorylates Chk2 ? phosphorylates/ inactivate
Cdc25
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DNA Damage Triggers the G2 Checkpoint
DNA damage ? ATM kinase ? phosphorylates Chk2 ?
phosphorylates/ inactivate Cdc25
21
Checkpoints function at each stage of the cell
cycle in S. cerevisiae
DNA damage ? Sensor ? Transducer ? Effector
22
Animal Cell Cycle Is Controlled by Many
Cdk-Cyclin Complexes.

• Many dimeric kinase complex (Cdk-cyclin) function
  in animal cells
• Cdk2 Cdk4
• Form a complex with G1 cyclins

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Control of Mitosis in Animal Cells Requires
Phosphorylation and Dephosphorylation of MPF

• Cdc2
• Thr-161 is phosphorylated by Cdc2-activating
  kinase (CAK) Wee1
• Cdc25
• Phosphatase
• Remove inhibitory phosphate from Tyr-15 of Cdc2

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Several Cdk-Cyclin Complexes Are Active at G1
and S Phases

• Cdk2, Cdk4, Cdk5
• Thr-161 is phosphorylated by Cdc2-activating
  kinase (CAK) Wee1
• D cyclins
• Active at G0/G1 phase
• Controlled by synthesis/ degradation
• E cyclins
• Active at G1/S phase
• Controlled by synthesis/ degradation
• A cyclins
• Active at S phase and G2/M phases (only)
• Controlled by synthesis/ degradation

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Which Protein Is Activated by Cdk-Cyclin Complex?

• RB
• Is a target of cdk-G1 cyclin (D) complexes
• Is a tumor suppressor
• RB-E2F ? transcription inhibited
• RB-p E2F ? transcription activated
• Cdk-cyclin complex phosphorylates RB

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Cdk Inhibitors Represent Alternative Ways to
Control Cell Cycle

• Cdk-cyclin inhibitors
• CKI
• CKI has its specificity to different cdk-cyclin
• Binds to cdk-cyclin
• Interferes with the phosphorylation of RB

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Target of CKI to Protein Degradation Controls
G1/S Progression

• Sic1
• A CKI in S. cerevisiae
• Binds to CDC28-CLB in G1 phase
• Sic1 is degraded when cells enter S phase
• Sic1 is degraded by proteolysis
• SCF (Cdc53-Skp1-Cdc4) E3 ligase confers the
  proteolysis of Sic1
• p27, p21, and cyclinE are also substrates of SCF
  E3 ligase

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Protein Degradation Also Regulates G2/M
Progression

• Cyclin A
• Is degraded in metaphase
• Cyclin B
• Is degraded in anaphase
• Pds1p
• Is degraded in anaphase
• Ensures sister chromatids to seperate

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Anaphase Promoting Complex/ Cyclosome (APC/C)
Regulates Proteolysis at G2/M Phase

• E3 ubiquitin ligase
• Activated by phosphorylation
• Degrade
• B-type cyclin, Pds1p, and the molecules
  unnecessary for mitosis
• Nucleotide synthesis enzyme
• DNA replication complex
• subunit

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The Proteolytic Activity of APC/C Ensure the
Proper Segregation of Sister Chromatids

• Pds1p/Securin
• Is degraded by APC/C
• Esp1/Separin
• Is activated by degradation of securin

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The APC/C Is Maintained at Inactive State until
All Kinetochores Are Attached

• Mad proteins
• Inhibit the activity of APC/C by binding to CDC20
• Bub proteins
• Inhibit the activity of APC/C by binding to CDH1

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Exit from Mitosis Is Controlled by Cdc14

• Cdc14
• A phosphatase dephosphorylates Cdh1 ? activate
  APC/C
• A phosphatase dephosphorylates Sic1 ? inactivate
  mitotic cyclins
• In nucleolus at interphase

33
Cell Structure Is Reorganized at Mitosis
34
Spindle Assembly Microtubule Nucleation
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Overview of Cell Cycle

• Protein synthesis/ degradation
• Transcription/ ubiquitin E3 ligase
• Activate/ Inactivate
• Post-translational modifications
• Coordinated events through cell cycle
• Conserved machinery during evolution