Origins of Mutations and DNA Replication/Repair

Question 1

3 common DNA lesions

Answer 1

1. Depurination – spontaneous hydrolyzation of bond between base and deoxyribose resulting in loss of purine
2. Deamination – loss of amine group due most commonly to spontaneous conversion of cytosine to uracil
3. Pyrimidine dimers – dimerization of adjacent thymine bases

Question 2

Difference between somatic and germline mutations

Answer 2

Somatic – affects only cell where mutation occurred and progeny, can be problematic if resulting in unregulated proliferation/survival of cell, if mutation occurs early in embryogenesis (can cause mosaicism)
Germline – pass on mutation to offspring of affected individual

Question 3

Gene mutation

Answer 3

Variation in nucleotide sequence (freq. = 10^-10 per bp per cell division/10^-5 or -6 per locus) – often occurs due to nucleotide exchange, insertion, or deletion

Question 4

Chromosome mutation

Answer 4

Rearrangements, deletions, or duplications of chromosomal regions (freq. = 6 x 10^-4 per cell division) – often follows DNA double strand breaks or faulty recombination (MORE LIKELY to cause disease than gene mutation)

Question 5

Genome mutation

Answer 5

Surplus or loss of chromosomes (freq. = 10^-2 per cell division) – often occurs due to mis-segregation of chromosomes during meiosis or mitosis (MORE LIKELY to cause disease than gene mutation)

Question 6

Point mutation (4 types)

Answer 6

Mutations resulting in exchange of one nucleotide for another, arising from replication errors, mutagenic chemicals, radiation, or repair of damaged DNA

1. Silent
2. Missense
3. Nonsense
4. RNA processing mutant

Question 7

Silent mutation

Answer 7

Does not change amino acid sequence of encoded protein

Question 8

Missense mutation

Answer 8

Changes amino acid sequence of encoded protein

Question 9

Nonsense mutation

Answer 9

Generates stop codon (UGA, UAA, UAG)

Question 10

Incorrect recombination (mechanisms leading to small insertions or deletions)

Answer 10

If homologous recombination between DNA molecules is done incorrectly due to misalignment, unequal crossover will occur, inserting or deleting genetic material (especially common at loci with repetitive DNA)

Question 11

Strand slippage during replication (mechanisms leading to small insertions or deletions)

Answer 11

Newly synthesized DNA strand may loop out in a single-stranded form, causing the strand to be replicated a second time, which creates DNA with extra repeated segments

Question 12

Intercalating agents (mechanisms leading to small insertions or deletions)

Answer 12

Chemical mutagens intercalating into DNA, distorting the structure of the molecule and resulting in insertion or deletion of one or more nucleotides during DNA replication

Question 13

Frameshift mutation

Answer 13

Insertion or deletion mutation that is not a multiple of three, altering the codon structure downstream of the mutation to create different protein products (responsible for 25% of human genetic diseases)

Question 14

Describe the process of meiosis.

Answer 14

• -Prophase I of meiotic division sees chromosomes condense and become visible
• -Homologous chromosomes (maternal and paternal copies) align in synapsis process, held tightly together by synaptonemal complex
• -Homologous chromosomes exchange homologous fragments by crossing over
• -Synaptonemal complex disintegrates and chromosomes are held together by chiasmata (sites of crossovers)
• -In metaphase, chrosomes are positioned at equator
• -In anaphase, homologous chromosomes are separated in disjunction
• -Cell divides to give rise to two haploid cells, each with 23 chromosomes
• -Sister chromatids are divided equally between two daughter cells in second meiotic division

Question 15

Crossing over

Answer 15

Exchange of homologous fragments on homologous chromosomes that leads to genetic recombination (VERY IMPORTANT!)

Question 16

Disjunction

Answer 16

Proper separation of homologous chromosomes in anaphase

Question 17

Nondisjunction

Answer 17

Incomplete separation of chromosomes in anaphase, leading to germ cells with surplus or missing chromosomes (increases with maternal age)

Question 18

Which autosomes does trisomy occur in?

Answer 18

13 (Patau syndrome), 18 (Edward syndrome), 21 (Down syndrome) – this is a GENOME MUTATION

Question 19

Mosaicism

Answer 19

Cells with different genotypes occurring in the same person that results from somatic mutations in embryogenesis (occurs in 2-4% of trisomy 21)

Question 20

Proofreading activity of DNA polymerases

Answer 20

DNA polymerase δ has 3’ –> 5’ exonuclease activity which edits 3’ end of growing DNA strand; if polymerase detects wrong base pairing, 3’ –> 5’ exonuclease activity hydrolyzes new phosphodiester bond and polymerase tries again

Question 21

Strand-directed mismatch repair

Answer 21

DNA repair enzymes recognize nicks and gaps in newly synthesized strand and distortions in the DNA double helix that result from incorrect base pairing; these enzymes remove the incorrect base pair and surrounding area which is then filled in by DNA polymerase

Question 22

HHNPCC (Lynch syndrome)

Answer 22

Heritable cancer syndrome that causes 80% chance of developing colorectal cancer (and others); caused by defects in the mismatch repair system

Question 23

Base excision repair

Answer 23

DNA glycosylases recognize specific types of altered base and catalyze their removal hydrolytically (ex. removal by uracil DNA glycosylase of cytosine that has been deaminated to uracil)

Question 24

Steps of base excision repair

Answer 24

1. Nucleotide is spontaneously altered to the wrong one
2. Uracil DNA glycosylase deletes the wrong nucleotide
3. AP endonuclease cuts the backbone
4. DNA phosphodiesterase removes deoxyribose phosphate group
5. Gap is filled by DNA polymerase and backbone repaired by DNA ligase

Question 25

Nucleotide excision repair

Answer 25

Repairs damage caused by agents that result in large changes in structure of DNA (ex. pyrimidine dimers)

Question 26

How is a pyrimidine dimer repaired?

Answer 26

1. Enzyme complex scans DNA molecule and recognizes distortions in double helix
2. Helicase separates strands and phosphodiester bond is cut on either side of affected area
3. Oligonucleotide containing lesion (section of strand including dimer) is peeled off
4. Gap in DNA helix produced by nucleotide excision is filled/fixed with DNA polymerase and ligase

Question 27

Xeroderma pigmentosa

Answer 27

Disorder characterized by extreme sensitivity of the skin to sunlight, pigmentation changes, and skin cancers; caused by mutations abolishing/impairing proteins required for nucleotide excision repair

Question 28

Why are double-stranded DNA breaks troublesome?

Answer 28

Makes repair difficult due to no intact strand remaining

Question 29

What mechanisms can fix double-stranded DNA breaks?

Answer 29

1. Nonhomologous end-joining (results in loss of nucleotide at break point, permanently changing nucleotide sequence)
2. Homologous end-joining (recombination processes used to repair broken DNA so that info needed to repair lesion without loss of sequence can be used)

Question 30

Cytaburine

Answer 30

Analogue of cytidine that contains arabinose instead of ribose; converts to cytarabine triphosphate inside cells which then competes with deoxyribonucleotides for binding to DNA polymerases

Question 31

Cyclophosphamide

Answer 31

Converted to active drug phosphoramide mustard in liver, which is a bi-functional alkylating agent that forms inter- and intra-strand DNA crosslinks

Question 32

Doxorubicin

Answer 32

Forms tripartite copmlex with DNA and topoisomerase II and causes double-strand breaks to accumulate