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Ch' 18' Cancer and DNA Repair

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Title: Ch' 18' Cancer and DNA Repair


1
Ch. 18. Cancer and DNA Repair
  • What is cancer?
  • 1) Ability to proliferate uncontrollably and
    ignore usual signals to differentiate or undergo
    apoptosis
  • Apoptosis programmed cell death
  • Carcinogenesis cancer development
  • 2) Resulted from genetic predisposition,
    environmental factors or infections

2
Cancer is a genetic disease
  • ? Damage to DNA ? activate proto-oncogenes or
    inactivate tumor suppressor genes
  • oncogenes viral genes which are ?responsible
    for transforming cells (cancer) and ?activated
    versions of normal cellular genes
    (proto-oncogenes)
  • - proto-oncogene
  • - tumor suppressor

3
Genetic alterations in cancer
  • Gene amplifications
  • Deletions
  • Chromosomal translocations
  • Point mutations
  • Epigenetic changes
  • Hypo and hyper methylation in promoter regions
  • Modifications of histone

4
Figure 18.02 Action of proto-oncogenes and tumor
suppressor genes.
5
Multiple defects contribute to cancer
  • Cell division and differentiation
  • Apoptosis
  • Telomere erosion
  • Contact inhibition
  • Angiogenesis
  • Metastasis
  • Cancer therapy
  • ? non-specific treatment suppressing cell
    proliferation
  • ? specific herceptin (HER2), Gleevec (Bcr-Abl)

6
Figure 18.03a Contact inhibition.
7
  • 2. DNA damage and Repair
  • DNA damage? repair OR mutation
  • DNA can be damaged in different ways
  • UV induces covalent linkages between
  • adjacent thymine bases ?interferes normal
  • replication transcription
  • 2) Ionizing radiation direct action or
    indirectly by formation of free radicals
  • 3) Chemicals (carcinogens, mutagens) react
    with DNA, benzopyrene (in tobacco smoke),
    aflatoxin (produced by mold)
  • 4) Reactive oxygen species produced during
    cellular metabolism chemical modification of DNA
    (gt100 modifications, 8-oxoguanine)
  • 5) Nonenzymatic reactions under physiological
    conditions hydrolysis of the N-glycosidic bond
    or deamination of cytosine
  • 6) Errors introduced during normal DNA
    replication

8
  • Repair enzymes restore damaged DNA
  • DNA photolyase in bacteria, restore thymidine
    dimer
  • A Methyltransferase remove methyl groups of
    O6-methylguanine
  • Mismatch repair system
  • ? MutS bind to the mispair
  • ? Endonuclease cleave the strand
  • ? Helicase unwinds the helix (The DNA
    defective region is fixed)

9
Figure 18.04 the mismatch repair protein MutS
bound to DNA.
10
Base excision repair corrects the most frequent
DNA lesions
1) Uracil-DNA glycosylase Substrate
catalysis 2) AP endonuclease nick the DNA
backbone at the abasic ribose 3) DNA polymerase
replace a base gap or as many as 10 bases (flap
endonuclease instead of AP endonuclease). 4) DNA
ligase seal the nick
11
Nucleotide excision repair targets the second
most common form of DNA damage
  • target DNA damage from UV or oxidation
  • 30 bases removed
  • Enzymes identified by studies using two
    hereditary diseases
  • a. Cockayne syndrome mutations in any of 5
    enzymes to detect RNA polymerase to be stalled
  • b. Xeroderma pigmentosum mutations in any of 7
    enzymes participate in nucleotide excision repair
  • error-prone DNA polymerase (?)

12
Some damage can be repaired through recombination
  • A single-strand break
  • Double-strand break
  • End-joining can be mutagenic
  • ataxia telangiectasia a disease caused by
    recombination defective
  • Expression of DNA repair proteins
  • 1) proteins to carry out recombinations
    constitutive expression? DNA strand breaks
    unavoidable?
  • 2) Proteins related to other DNA repair
    mechanisms inducible expression, constitutive
    expression could be harmful to cells

13
Figure 18.11 Recombination to bypass a
single-strand gap.
14
Figure 18.12 Homolgous recombination to replicate
past a double-strand break.
15
Figure 18.13 End joining of broken DNA.
16
3. Cell Cycle Control
  • Genetic unstability of cancer cells
  • Cell cycle S (synthesis)-G2-M(mitosis)-G1-S
  • Check points
  • ? cyclin/ cdk (cyclin-dependent kinases)
    ubiquitin proteasome (cyclin degradation)
  • ? presence of growth-promoting signals, DNA
    damage, telomere length, proper spindle formation
  • 4) Not well?? Halt the cell cycle, promote DNA
    repair or kill the cell by apoptosis

17
3. Cell Cycle Control
5) Signal transduction kinase, phosphatase and
other components, about 20 of the protein
coding genes works a s the network with
redundancy eg Rb (tumor suppressor
genes)
18
Some members of the DNA-damage checkpoint pathway
have been identified
  • Check point proteins may resemble PCNA (the
    eukaryotic sliding-clamp protein that enhances
    the processivity of DNA polymerase)
  • ?ATM a defective gene in ataxia telangiectasia,
    a protein kinase
  • A large protein with a DNA-binding domain
  • To monitor DNA damaged or incompletely replicated
  • Able to bind to double-strand breaks and
    activates repair by recombination
  • Kinase substrate BRCA1 (breast cancer causing
    gene), p53
  • p-BRCA1 bringing together repair proteins,
    binds to double stranded breaks

19
  • P53 plays a central role in cancer
  • Tumor suppressor one of the most mutated genes
    in cancer (gt50 )
  • P53 level controlled by its degradation rate
  • DNA damage ? activate ATM ? phosphorylate p53 ?
    loss of p53 binding protein mediating p53
    degradation (p53 to be stabilized)
  • Active p53 (transcription factor)
  • synthesize a protein that inhibits cyclin
    dependent kinase ? block cell cycle progression
    (time to repair DNA)
  • Induce synthesis of DNA repair enzyme and
    ribonucleotide reductase
  • ? Induce synthesis of apoptosis proteins

20
Figure 18.19a Structure of the core domain of P53.
21
Box 18-A Gene expression profiling ( microarray)
Functional genomics gene expression and gene
regulation
  • Gene Expression
  • Single gene, spatial/temporal expression
  • Genome-wide expression profiling
  • Gene Regulation
  • Overexpression/Misexpression
  • Wrong place/wrong time

22
The Challenge
  • Traditional gene expression studies involve one
    or a few genes
  • How can we observe the expression of most or all
    of the genes in an organism?

23
Microarrays Based on nucleic acid hybridization
  • Dot-blots Known sequences at specific positions
  • Test thousands of genes at once

Global gene expression profiling
24
Target Labeling
Two color hybridization Compare gene expression
in 2 samples by allowing differentially-labeled
cDNA targets to competetively hybridize to
microarray probes.
25
Spotted Microarray 2 Color Hybridization
26
Box 18-B Cell death necrosis, apoptosis,
autophagy
27
Cell deathapoptosis
  • Programmed cell death
  • Cell intrinsic mechanism for suicide
  • Regulated by variety of cell signaling
  • Nuclear condensation and fragmentation (200bp),
    cleavage of chromosomal DNA into intranucleosomal
    fragments, packaging of the deceased cell into
    apoptotic bodies w/o plasma membrane breakdown
  • Absence of inflammation around dying cells
  • Resulted from activation of caspase cascade
  • Require ATP

28
Cell deathnecrosis
  • A passive form of cell death
  • Initiated by cellular accidents (toxic insults,
    physical damage)
  • Vacuolation of the cytoplasm, breakdown of the
    plasma membrane, induction of inflammation around
    dying cells
  • Programmed necrosis tissue culture phenomena?

29
Cell deathautophagy
  • Eat oneself during nutrient stress, cellular
    constituents are degraded for energy production
  • Formation of double membrane vesicles in cytosol
    ? encapsulating organells ? fusion with lysosome
    ? contents are degraded and recycled
  • Death cells digest themselves to death
  • Autophagy survial or suicide pathway or both?

30
  • Box 18-B Apoptosis
  • a form of programmed cell death in multicellular
    organisms.
  • Orderly cellular self destruction
  • crucial for survial of multicellular organisms as
    cell division
  • involves a series of biochemical events leading
    to a characteristic cell morphology and death
  • changes to the cell membrane, cell shrinkage,
    nuclear fragmentation, chromatin condensation,
    and chromosomal DNA fragmentation
  • cellular debris whose results do not damage the
    organism differentiate
  • cf necrosis
  • 5) Specific genes carrying out cascade reactions
    caspases, cytochrome c

31
Importance of Apoptosis
  • Important in normal physiology / development
  • Development Immune systems maturation,
    Morphogenesis, Neural development
  • Adult Immune privilege, DNA Damage and wound
    repair.
  • Excess apoptosis
  • Neurodegenerative diseases
  • Deficient apoptosis
  • Cancer
  • Autoimmunity
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