Maintenance of genome Flashcards
How does damage to the DNA occur?
Copying errors during DNA replication
* Spontaneous depurination
* Exposure to different agents
e.g. background ionising radiation, UV light, tobacco
What are the five different ways to repair DNA
- Direct reversal of damage
- Base excision
- Nucleotide excision repair
- Homologous recombination repair and non homologous end joining
- DNA mismatch repair
DR: occurs mainly in prokaryotes
BE: corrects DNA damage caused by reactive oxygen species deamination, hydroxylation, spontaneous depurination
NER: - removes adducts that
produce large distortions in DNA
homologous: repairs DNA double strand
breaks
DMR: repairs copy errors made
during replication
How does guanine being methylated become problmatic
- Guanaine binds to cytosine
- but because its been methylated guanine
- O 6 alkyl guanine pairs with thymine.
- on next replication of strand thymine will pair with adenine instead
- this causes a kink in DNA
- causes problems during replication,
- DNA polymerase cant synthesise DNA becasue of kink so cell dies
- if methyl guanine does not cause cell death, it can cause mutageninis, (e.g O-6 alkyl)
How can UV damage DNA
- can cause thymine dimers (two adjacent thymine bases become covalently crossed linked)
- causes a disotrion in DNA
- creates problems when replicating
Way 2
1. Can create 6-4 photoproducts
UV damage can be mutagenic
How does the cell solve UV induced dimmers
in bacteria
can be removed by photolipase
humans dont do this
How do we remove methylated bases
use alkyl tranferase to remove alkyl group
How do we fix strand breaks in sugar phosphate backbone
ligate them back together
In what situations would a cell do base exicision repair
- Spontaneous hydrolytic depurination of DNA.
- Deamination of cytosine (changes into uracil)
- Exposure to reactive small
metabolites (causes DNA adducts)
class switchig is the only time cytsoime is converted into uracil
How does BER work
- remove base using glycoslayes
- this site is now abasic (theres the S-P backbone)
- apuric endonuclease releases that there is a base missing, it removes the site (removal of sugar and base)
- DNA polymerase comes along fills in gap
- ligase and then good to go
How does Nucleotide excision repair work
- works on double stranded not single stranded (not useful in DNA replication)
- Non specfic -> recgonises disortions, not specific adducts
- removes large adducts
- effiecent and error free
how does it work
1. XPC gene recgonises 6, 4 proudct (specfic example)
2. endonueclase cut the distortion (e.g XPB, + D) -> use single stranded binding protein is present to stop DNA being cut up since its vulnerable
2. exonuecluses remove damged bit of DNA (e.g XPF AND G)
3. DNA polymerase fills in Gap
4. then ligase pulls it back together
we use this process usually for UV light
What is XERODERMA PIGMENTOSUM
epidemology, what is it,
- Autosomal recessive disorder
- happens to 1,4 per million
- exteremly sensitive to sun
- develop tumors
- can have neurological abnormalities due to lack of NER
- cells have defect in DNA nucleotide excision repair
- XP cells show a high mutation rate (may be due to unexcised dimers, so incorrect bases are incorporated to oppose damage)
What can mutations in the PTCH 1 gene cause
can cause basal cell carcionmas
patients with XP that also had BCC had problems with PTCH1
DNA replication doesnt stop in XP, so it uses daughter strand gap repair
how does that work
1.DNA polymerase hits thymine dimmer cant get passed it
2.DNA polymerase goes passed it somehow
3.and this leads to daughter strand gaps
4. the thymine dimer is still present + XP variants come in here
What happens in XP variants
- in XP there is a defect in repliation of DNA after UV exposure
- the enxyme DNA polymerase n, is defeinct in this disorder, which makes it unable to replicate DNA past UV photoproducts
- in the absence of the proper DNA polymerase, so XP variants come along
- but these arent great at their job, and can cause muataions to happen
How does DNA double strand break happen
- Caused by errors in replication
- Reactive oxygen species (cellular metabolism) -> hair dying, teeth whitening
exposure to ionising radiation (x rays, cosmic rays, chemo)
BRCA 1 structure
has two critical domains
means its a E3 ubiquntine ligase
- likes putting ubiquitin on other proteins
BRCA 2 structure
-has BCR repeats
-need this to bind to Rad51
-twice as big as BRCA 1
what is the role of BRCA 1 and 2
allow for repair for double strand breaks via
1.Non-homologous end joining
2.Homologous recombination repair
How does homologous recomboination repair work
1.detect break, by MRe11/Rad50/NBs1
2.this cuts DNA and leaves a small amount of single stranded DNA
3.single stranded DNA binds to Rad51
4. Rad51 then looks at undamaged strand, to find exact sequence
5. BRCA1 and 2 load Rad51 onto DNA
6. so you form interlinked DNA
7. the cell does DNA synthesise across the break, a loop forms
8. and tnen the cell can synthesise new strand across this using the undamaged dna strand template
9. problem the two strands connect
9. then you unconnect the two strands
10. ligase and then dip
problem: two strands end up together idk dk what he said
for this process you need to exact temeplate so happens in s phase and stuff
What is the difference between BRCA 1 and 2
BRCA 2 involved in controlling Rad51
BRCA 1 has roles in Rad51 but also functions like checking cell cycle check points
Cells deficient in BRCA1/2 are unusually sensitive to PARP inhibitors
What are PARP inhibitors
PARP is an enzyme that make chains of ADP ribose
the inhibitor prevents this
if you inhibit PARP, it gets trapped in DNA so you get double strand break with protein stuck on end
you need homologous recombination to repair this
if you dont have this the cell can die (double strand breaks are BAD)
when does a cell induce double strand breaks
when making immune cells b and t lymphocytes
during meiosis -> swapping of genes
How does Non homologus end joining work
- Break in DNA is sensed by Ku70/86 heterodimmer
- binds to end of DNA
- recurits kinase called DNA PKcs
- this becomes active kinase, phosphyrlates proteins
- can trigger artemis (trims back DNA to reveal some single stranded DNA)
- single strand DNA undergoes microhomology base pairings
- Flap gets removed
no idea how this works
How does DNA mistmacth repair work
- repairs base mistmatches
- can repair insertion deletion loops due to polymerase slippage -> microsatalite instablity -> important in huntingtons
Can get microstalitie instability, even though you fixed DNA more likely to get mutations
What is the mutator phenotype hypothesis
-if you have base mismacthes and repeated sequences
-if you cannot repair this using DNA mismatch repair,
-then more mutations can happen
-can lead to adenoma to become carcinoma
-you need to get additional mutations
