cell cycle and cell growth control-2011 text
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Cell Cycle and Cell Growth Control
By Chung-Yih Wang, MD, PhD
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National Taiwan University Medical SchoolUniversity of Wisconsin-MadisonUniversity of Michigan, Ann ArborUniversity of ChicagoNational Yan-Ming UniversityVeterans General Hospital-Taipei, Cancer CenterCheng-Hsin Hospital, Department of Radiotherapy
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Course Organization
Cell cycle controlMitosis Apoptosis Cancer Biology and Novel Treatment
Strategy Against Cancer
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Learn Goals
Understand the stages of the cell cycle Key Factors controlling
the passage of the cell cycle The role of apoptosis in
physiology/ Pathology Controls of apoptosis
Carcinogenesis Oncogenes and tumor
suppressor genes New strategies for
cancer therapy
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Examination
A thorough understanding of the physiological processes and mechanisms is more important than memorizing details and terminology 15 multiple choice questions and two to three
The grades will be adjusted to match normal distribution
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Why bother learning the basic medical science?
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Anti-fungus treatment Organ Transplantation Cancer Treatment Autism
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Rapamycin
Rapamycin was first discovered as a product of the bacterium Streptomyceshygroscopicus [1] in a soil sample from Easter Island originally developed as an
antifungal agent this was abandoned when it
was discovered that it had potent immunosuppressiveand antiproliferativeproperties
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mTOR: mammalian Target of Rapamycin
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Rapamycin
Organ Transplantation used in coronary stents to prevent re-stenosis
following balloon angioplasty Cancer treatment Potential treatment for autism
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Reductionism
Biology is a reductionistic Science To understand the
nature of complex things by reducing them to simpler or more fundamental things
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Holism
the idea that all the properties of a given system cannot be determined or explained by its component parts alone. Instead, the system as a whole determines in an important way how the parts behave.
Aristole: Metaphysics: "The whole is different from the sum of its parts"
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Life History on Earth
Birth of Earth
1/1
Birth of Life
2/17
Eukaryotic Life
9/5
Mammal
12/26
Human
12/3123:59:58
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Cell Cycle
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What You Need to Know?
The stages and their physiological roles of cell cycle ( G1, S, G2, M) Key regulators and their mechanisms of
regulating cell cycle progression (Cyclin, CDK, CKI..)
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Cell Cycle
Cell cycle is the series of events that take place in a dividing cell. These events can be divided in two main parts:
Inter-phase (G1, S, and G2) and M-phase The molecular events that control the cell cycle are
ordered and directional; that is, each process occurs in a sequential fashion and it is impossible to "reverse" the cycle.
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Leland H. Hartwell, R. Timothy Hunt, and Paul M. Nurse won the 2001 Nobel Prize in Physiology or Medicine for their discovery of the central molecules in the regulation of the cell cycle.
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Cell Cycle
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Overview of Cell Cycle
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Techniques and Materials Commonly Used in The
Study of Cell Cycle
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Xenopus laevis (African Clawed Frog)
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Xenopus laevis
Large size Huge amount of cytoplasm Rapid, synchronously
division after fertilization Easy to manipulate
cytoplasm and nucleus S and M with no
interference from G1 or G2 control mechanisms
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Yeast
Saccharomycescerevisae Budding Yeast
Schizosaccharomycespombe Fission yeast
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Yeast
Rapid growth Small genome size (1%
that of mammalian) Easy to manipulate
genetically Can proliferate as
haploid
Easy to isolate and study mutant with phenotype in cell cycle Cell-division-cycle
genes (cdc) Conditional mutation Well conserved cell
cycle control genes from yeast to human
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Cells
3 Categories of Cells Can not divide: Terminally
differentiated cells: Nerve cells, Muscle cells, Red blood cells Normally do not divide:
Liver cells, Mature Lymphocytes Normally keep dividing:
Fertilized egg, Bone marrow, Epithelium
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Control of Cell Cycle Progression
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Cell Cycle
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Cell Fusion Experiment
In the 1970 Whether the cytoplasm
contains the regulatory factors that affect cell cycle activities Cytoplasm of a replicating
cell contains factors that stimulate DNA synthesis
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Cell Fusion Experiment
M+ G1 M+S M+G2
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Cell Fusion Experiment
The transitions from G1 to S and from G2 to M are both under positive control Both are induced by
stimulatory factor(s)
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What is the factor(s) that promotes the transitions from G1 to S and from G2 to M ?
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Isolation of MPF
Maturation Promoting Factor : MPF
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MPF activity is also present in mammalian cells, Hela, and sea urchin eggs Sea urchin eggs fail to
undergo mitosis if put in protein synthesis inhibitor: MPF require new protein synthesis Fluctuation of the protein
during cell cycle: Cyclin
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MPF
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1983 Tim Hunt et, al use 35S methionine to label protein in fertilized and un-fertilized sea urchin egg. What will you expect
the MPF to behave in such experiment?
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MPF
Protein synthesis inhibitor blocked MPF activity 35S Methionine labelling
followed by electrophoresis identify possible candidates for MPF The potential MPF will
appear and disappear at specific time corresponding to cell cycle
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MPF
Partially purified MPF stimulate the incorporation of 32P into proteins Suggesting that MPF is
a protein kinase Purified MPF contains
two subunits, 32kDa and 45 kDa
cDNA of putative cyclin were cloned mRNA of cyclin, when
injected to Xenopusoocyte, can promote cell cycle progression
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Yeast cdc
Ts
Cell-division-cycle genes (cdc)
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MPF/Yeast cdc
Yeast cdc2 gene encode a 34kDa protein kinase p34cdc2 Antibody against cdc2
react with 32kDa protein of MPF
Vertebrate cdc2 homologue dose not fluctuate during the cell cycle, which suggest that this 32kDa kinase depend on another protein
45 kDa : Cyclin
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MPF
An activities identified in frog oocytes that promote mitosis MPF activity fluctuate
during the cell cycle Two components, one
needs novel protein synthesis: Cyclin, the other one contains kinase activity: CDK
Human, Frog, Sea urchin, and yeast are all close relatives
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MPF
Consist of two subunits One regulatory subunit:Cyclin One subunit with kinase activity: Cdk
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Fission yeast The same Cdk (cdc2) is
responsible for G1 and M
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Mammalian Cell Cycle
There are four classes of cyclins Each cyclin binds to and
regulate the activity of a Cdk G1/S Cyclin: commit the
cell to DNA replication S Cyclin: Initiation of
DNA replication M Cyclin: mitosis G1 Cyclin: Start
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How is CDK Regulated During Cell Cycle?
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Cyclin Regulates Cdk
Binding of cyclin1.causes a conformation
change and activation of Cdk
2. Directs it to specific target
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Wee1 encodes a tyrosine kinase Cdc25 encodes a
phosphotase
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Regulation of CDK in yeast
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Phosphorylation
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Regulation of CDK
Cyclin Binding Cdk Phosphorylation Inhibitory Stimulatory Cdk Inhibitors Controlled Proteolysis Subcellualr Localization
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Cdk Inhibitory Proteins (CKIs)
P21 , P27 Crucial In the
pathogenesis of Cancer
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Proteolysis Control (G1, S)
Cyclin and CKI destruction by a Ubiquitin-conjugating system
G1/S: SCF Ubiquitylation by SCF is
controlled by phosphrylation, only specifically phosphorylated proteins are recognized
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Proteolysis Control (M)
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Subcellular Localization (Cyclin B1, M-cyclin)
G2 M
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Cdk Regulation
Cyclin Binding Cdk Phosphorylation Inhibitory Stimulatory
Controlled Proteolysis Cdk Inhibitors Subcellualr Localization
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S Phase Initiation
How does a cell know when to start DNA synthesis
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CDK activity in G1 is suppressed by: accumulation of CKIs, inhibition of cyclingene transcriptionWhat control the transcription of cyclin gene?
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Rb in G1-S
Transcription of G1/S and S-Cyclin requires E2F E2F is inhibited by
binding with pRb
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Retinoblastoma (Rb) is a rapidly developing cancer which develops in the cells of retina . A very treatable cancer There are two forms of the disease, heritable
form and a non-heritable form.
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S Phase Initiation Slow accumulation of G1-Cdk
during G1 G1-Cdk phosphorylates pRb E2F enhance its own and S-
Cyclin transcription S-Cdk increases pRb
phosphorylation S-Cdk phosphorylates and
inactivate P27-CKI E2F promote transcription of
genes required for DNA synthesis
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S Phase Initiation
Result: Rapid and Complete activation of S-Cdk and
S-initiation A typical example of biological control
system with multiple feedback loop
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Checkpoint
Cell cycle progress will stop if: DNA is damaged DNA is not properly
replicated during S Chromosomes are not well
aligned during M Checkpoints ensure the
integrity of genome during cell cycle
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DNA Damage Chcekpoints
G1: Phosphorylation and
activation of of P53 , Transcription
activation of P21
G2 Phosphorylation and
destruction of CDC25 G2CDK remains
inactive without CDC25
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G1 DNA Damage Checkpoint
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ATM (Ataxia-telangiectasia Mutation)
Story of Ataxia-telangiectasia a rare, neurodegenerative, inherited disease affects the cerebellum, immune system and
predispose to cancer increased sensitivity to ionizing radiation, but
normal response to other forms of radiation such as ultraviolet light
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Entry into Mitosis
M-cyclin transcription increase during G2 and M Gradual accumulation of
M-Cdk, which are phosphrylated by CAK, but remain inactive due to inhibitory phosphorylationby wee1
Activation of Cdc25 phosphotase leads to activation of M-Cdk, which then positively enhance its own activation
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Entry into Mitosis