Transcript Document

Ch 7 Mutation
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Heritable change in DNA
Random
Source of genetic variation
may be advantageous, deleterious,
neutral
Mutation (+ sexual reproduction)
+ natural selection = evolution
FYI: Adaptation vs mutation
• Adaptation – not proven
– environment induces specific adaptive change
– Ex. E. coli + ampicillin – some cells become
resistant in response
• Mutation
– mutation is random
– E. coli DNA mutates randomly – statistically,
one will result in amp resistance, this cell grows
resulting in species adaptation
Types of mutations
• germ-line mutation
– in sperm or egg (meiosis), will be passed to offspring
• somatic mutation
– body cells, not passed on to offspring
• spontaneous mutation
– natural aspect of DNA replication
• induced
– UV light, chemicals, X-ray etc.
Germ line or somatic?
cat
Spontaneous?
Induced?
Drosophila
Euphorbia
basal cell tumor
Mutation and natural selection
Point mutation
• single base substitution,
deletion, or addition
Point mutations
Base pair substitutions
transition = G to A
transversion = ?
T to C
Missense mutation
 Wrong amino acid encoded
Sickle cell Hb
Subsititution,
addition or
deletion?
TS or TV?
Which codon
position?
• Glu hydrophilic
• Val hydrophobic (reduces solubility
of the molecule)
• Non-conservative mutation
Red blood cells
Conservative mutation
Example: Val to Leu both hydrophobic
Nonsense mutation
– Premature stop codon
– Codon AAA  UAA
lys
stop
5' - ATG ACT CAC CGA GCG CGA AGC TGA - 3‘
3' - TAC TGA GTG GCT CGC GCT TCG ACT
5' mRNA5' - AUG ACU CAC CGA GCG CGA AGC UGA - 3‘
Protein: Met Thr His Arg Ala Arg Ser Stop
5' - ATG ACT CAC TGA GCG CGA AGC TGA - 3‘
3' - TAC TGA GTG ACT CGC GCT TCG ACT - 5'
mRNA: 5' - AUG ACU CAC UGA GCG CGU AGC UGA - 3'
Protein: Met Thr His Stop
Neutral mutation
- mutation does not change amino acid folding
- codon AAA  AGA
lys
arg
(both basic aa)
Silent mutation
– No amino acid change, usually 3rd position TS
codon GCA  GCG
arg
arg
Frameshift mutation
– Shifts reading frame -->
scrambled message
– May lead to a premature stop
codon
The sun was hot but the man did
not get his hat.
Transition or transversion and which
codon position?
TATACCTAT
TATACCCTA
Other mutations
• Larger insertions, trinucleotide repeats,
deletions, duplications, translocation,
inversions
Trinucleotide repeat pg. 475
Fragile X Syndrome
• Xq27.3 FMR-1 gene
• X-linked dominant
• 2nd leading cause of mental retardation
(1/4000 males, 1/6000 females)
(Reprinted from Medical Genetics, 2nd ed., Jorde LB, et al, ©2000, with permission from
Elsevier Science.)
• CGG repeat in promoter region
– Normal = 6-54 repeats
– Normal carriers = 55 – 200 repeats
(premutation expansion of repeat)
– Fragile X syndrome = 200 – 1300 repeats
triplet repeat amplification
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FMR-1  FMRP (mRNA binding protein)
FMRP regulates translation of mRNA
Affects synaptic plasticity – signaling between neurons for learning and
memory
>200 repeats and FMR1 gene silenced  retardation
Caused by slipping of DNA polymerase during replication
Trinucleotide repeats
Mutation rate
Human estimate is ~1 x 10-6 spontaneous
mutations/gene /generation
Mutation rates per gene differ.
May be hot spots
Causes of mutations
1. Spontaneous mistakes by DNA
polymerase = replication errors
Most repaired by DNA proofreading and repair systems
2. Induced mutations
• Chemicals
Ethidium bromide
Mustard gas, industrial waste, PCBs, DDT
• Base analogues
– Example: Bromouracil substitutes for thymine
• Radiation – sun (UV), radon, X-ray
– Breaks, dimers…
– High doses kill cells
The Ames test
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Indirect assay for carcinogens
Plate 108 auxotrophic bacteria (his-)
Add filter disc with chemical
Look for revertants to his+
= mutations
• Add rat liver extract to look at metabolism
Spontaneous reversion rate
to His+ colonies
1000X higher rate
with mutagen
Repair of mutations
1. Direct reversal
Example: Mismatch repair
– 3’  5’ proofreading by DNA polymerase
Also, UV repair (bacteria) and alkylation repair (prokaryotes)
2. Excision repair
Ex. base excision repair
- single strand incision at both sides of error;
- Excise mistake
- DNA synthesis to replace the gap
- Ligation
- Also UV repair prokaryotes, methyl mismatch repair, SOS
response
Defective excision repair: Xeroderma
pigmentosum (1/250,000) 9q34.1
• Autosomal recessive
• Extreme sun sensitivity
– High risk of skin cancer (1000X) average by
age 8
– Metastatic melanoma
• Corneal damage
Knockout mice for excision repair
(XPA gene)
• XPA gene knocked out
• UV sensitive and develop tumors
• Study cancer, drugs
Small deletion example: Cystic
fibrosis (1989)
• CFTR gene (chromosome 7)
• Encodes a transmembrane
chloride channel active in
cells that line the lungs
Cystic fibrosis recessive mutation
p508
Will the mRNA message be in frame?
Effects of p508 CFTR mutation
• The 3 base deletion results in a
channel protein that does not
function correctly
• Thick mucus in the lungs,
infections, lung stress
Phenotypic effects of mutation:
Loss of function
• Drosophila white eye gene
• Wildtype = red eyes
Gain of function mutation
• Antennapedia (Drosophila)– legs
on head (ectopic) in place of
antennae
– Legs are normally expressed in
second thoracic segment
Knockout mice for excision repair
(XPA gene)
• Genetically engineered to not express XPA gene
• Mice are UV sensitive and develop tumors
• Use mice for DNA repair/cancer study
Defective DNA mismatch repair:
Hereditary nonpolyposis colon cancer
• Inherited predisposition to colon cancer
• ~4 people in family have had colorectal
cancer, multiple generations, before age 50
Small deletion example: Cystic
fibrosis (1989)