11.1 DNA damage and repair

Question 1

What is slippage?

Answer 1

• A mistake DNA poly makes by “slipping” when replicating repetitive regions
• This can lead to expansion of the repetitive sequence and trinucleotide repeat disorders

Question 2

What can chemical exposures (nitrates, smoke, other compounds) do to DNA?

Answer 2

destroy or chemically modify nitrogenous bases

Question 3

What does UV radiation do to DNA?

Answer 3

• introduces covalent bonds between adjacent thyamine bases (dimerization), leading to an interruption of the H bonds, creating a large bulge into the double helix structure
• interferes with DNA replication and gene expression

Question 4

What can reactive oxygen species do to DNA?

Answer 4

• Damage dna by creating nicks in the phosphate backbone of one strand, or a complete cleavage of dsDNA

Question 5

What can infectious diseases do to DNA?

Answer 5

triggers double stranded breaks in the cell lining of stomach

Question 6

What can viruses do to dna?

Answer 6

insert their genomes into host cells, sometimes disrupting normal functions of that region of the genome

Question 7

Direct repair:

Answer 7

• recognizes and directly removes a chemical modification, restoring the dna to its original form
• certain types (such as methylation or alkylation) are common
  each type of direct repair enzyme recognizes a specific modification.
• the phosphate backbone of a DNA chain is not interrupted

Question 8

Base excision repair:

Answer 8

removes and replaces a single nucleotide

Question 9

Mismatch repair and nucleotide excision repair

Answer 9

both involve removing a large segment of one of the DNA strands, and filling it in with newly synthesized dna.

Question 10

What can repair double stranded breaks in dna?

Answer 10

• nonhomologous end joining
• homologous recombination

Question 11

nonhomologous end joining

Answer 11

glues broken ends of DNA back together

Question 12

homologous recombination

Answer 12

uses the homologous chromosome to copy and repair the broken dna pieces

Question 13

What dna repair pathways can be a mutagenic process?

Answer 13

• nonhomologous end joining and homologous recombination, but this is less dangerous than leaving a double stranded break in the genome

Question 14

What is mismatch repair?

Answer 14

• fixes mismatches left behind by DNA polymerase

Question 15

What are the steps of mismatch repair?

Answer 15

1. Recognizing the mismatch
2. Resecting a portion of DNA by nucleases, leaving a gap
3. Resynthesizing the gap w polymerases
4. Sealing the gap with ligases

Question 16

How does the mismatch repair pathway know which is the correct nucleotide when it finds a mismatch?

Answer 16

• over time, the dna in our cells becomes methylated. Older strands have methylation patterns
• MMR proteins use methylation state to determine what is the old vs new strand of dna, then the unmethylated strand is removed and the methylated strand is used as the template strand for new dna synthesis
• MMR proteins are found tethered to the replisome during DNA replication, probably through the sliding clamp loader to ensure it can faithfully repair mismatches

Question 17

What does Base excision repair do?

Answer 17

• fixes mismatches or chemical modification to a base by removing and replacing 1 nucleotide

Question 18

How does base excision repair work?

Answer 18

• fixes or chemically modifies a base by removing and replacing a single nucleotide
• involves cutting and repairing the phosphate backbone

Question 19

What are glycosylases used for?

Answer 19

• Used in base excision repair (BER)
• They recognize specific base-pairing mistakes and remove a base, leaving a hole.
• There are more than different glycosylases in cells

Question 20

What do AP endonucleases do?

Answer 20

• Used in Base exision repair, remove sugar phosphate backbone

Question 21

Steps of base excision repair

Answer 21

• Glycosylases recognize specific base-pairing mistakes and remove a base, leaving a hole
• AP endonucleases remove sugar phosphate backbone
• DNA pol adds a new nucleotide
• Ligase seals the nick

Question 22

What is a fault of BER?

Answer 22

• BER is not always good at determining which nucleotide is the “correct” nucleotide, so sometimes the fix leads to a permanent mutation

Question 23

What does Nucleotide Excision Repair (NER) usually fix?

Answer 23

buldges in DNA from UV radiation or slippage

Question 24

What does NER do?

Answer 24

• recognizes buges and misshapen DNA structures
• Uses endonucleases to cleave the damaged DNA and surrounding nucleotides
• Then, polymerase synthesizes new dna and ligase seals the backbone

Question 25

What types of overhangs can ds breaks cause?

Answer 25

5' overhangs, 3' overhangs, 5' and 3' overhangs

Question 26

What happens during NHEJ?

Answer 26

* exonucleases and polymerases create blunt (even) ends by either nibbling away at the overhang, or by filling in the overhang * These blunt ended pieces of dna are then joined by ligases to repair the break. * The dsDNA break is repaired, but there are always INDELs (short insertions or deletions)... so it is a mutagenic process

Question 27

What happens in homologous recombination repair?

Answer 27

* ds breaks are fixed by resecting a larger portion of the DNA w exonucleases, creating large overhangs * Through a process known as strand invasion and a complex of rad proteins, the homologous chromosomes are found and then used as a template for new DNA synthesis * The repaired strand contains sequences that are identical to the homologous chromosome, and the template homologous chromosome will contain some of the dna from the damaged chromosome

Question 28

What happen as a result of homolgous recombination repair (HR)?

Answer 28

* the repaired strand contains sequence identical to the homologous chromosome * the template homologous chromosome will contain some of the dna from the damaged chromosome * this can lead to a loss of heterozygocity, where all the alleles of a particular sequence are the same or gene conversion (where sequences are moved between the homologous chromosomes)

Question 29

How many new mutations per year in somatic tissues and germline cells?

Answer 29

* somatic tissues: 2-60 new mutations/year * germline cells: 1-2 new mutations/year

Question 30

What type of mutation can our dna repair pathways not fix?

Answer 30

* large insertions * ex. HPV

Question 31

What is genetic anticipation?

Answer 31

* nucleotide repeats have a tendency to increase with each generation * this can lead to early onset and increased severity of symptoms, a phenomena known as genetic anticipation