Ch' 18' Cancer and DNA Repair

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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

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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

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Genetic alterations in cancer

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

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Figure 18.02 Action of proto-oncogenes and tumor
suppressor genes.
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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)

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Figure 18.03a Contact inhibition.
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• 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

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• 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)

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Figure 18.04 the mismatch repair protein MutS
bound to DNA.
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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
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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 (?)

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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

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Figure 18.11 Recombination to bypass a
single-strand gap.
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Figure 18.12 Homolgous recombination to replicate
past a double-strand break.
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Figure 18.13 End joining of broken DNA.
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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

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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)
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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

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• 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

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Figure 18.19a Structure of the core domain of P53.
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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

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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?

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Microarrays Based on nucleic acid hybridization

• Dot-blots Known sequences at specific positions
• Test thousands of genes at once

Global gene expression profiling
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Target Labeling
Two color hybridization Compare gene expression
in 2 samples by allowing differentially-labeled
cDNA targets to competetively hybridize to
microarray probes.
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Spotted Microarray 2 Color Hybridization
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Box 18-B Cell death necrosis, apoptosis,
autophagy
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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

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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?

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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?

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• 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

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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