L38: Origins of Mutations and DNA Repair

Question 1

Name three common DNA lesions

Answer 1

• Depurination: bond bw base and ribose hydrolyzed - Deamination: most often cytosine deamination generates uracil - Pyrimidine dimers: UV light dimerizes adjacent thymine bases on same DNA strand

Question 2

Distinguish bw somatic and germline mutations

Answer 2

• Somatic mutation: affects cell where mutation occurred, can cause CA development or mosaicism, no transfer to progeny - Germline mutation: transmitted to offspring of affect individual

Question 3

Distinguish bw gene mutations, cs mutations, and genome mutations. What are their approximate frequencies?

Answer 3

• Gene mutation: impact only single gene: 10^-10 per division - Chromosomal mutation: impact many genes: 10^-4 per division - Genomic mutations: surplus or loss of c/s: 10^-2 per division

Question 4

What are point mutations? How might point mutations arise? Distinguish bw silent, missense, and nonsense mutations.

Answer 4

• Exchange of one nt for another - Caused by replication errors, chemical mutagens, radiation, DNA damage repair - Silent: no changes to AA - Missense: change from one AA to another - Nonsense: change from AA to stop codon

Question 5

Describe three processes that can lead to small insertions or deletions.

Answer 5

1.) Alignment out of registration, unequal crossover 2.) Strand slippage in replication as a result of repetitive sequences 3.) Intercalating agents causes slip bw stacked base pairs of DNA that distort helix and cause insertions or deletions

Question 6

What is frameshift mutation?

Answer 6

• When deletion or insertion of 1 nucleotide occurs, resulting in shift of reading frame, causing nonsense or missense mutation

Question 7

Describe process of meiosis. Understand terms: crossing over, dysjunction, nondisjunction. What might the consequences of nondisjunction be?

Answer 7

1.) Chromosomes condense 2.) Maternal and paternal copies of c/s align: synapse, secured by synaptonemal complex 3.) C/s exchange homologous fragments (recombination of genetic material) = crossing over 4.) Synaptonemal complex disintegrates, held by chiasmata 5.) Disjunction = chromosomes pulled to each pole at anaphase 1 6.) Two haploid cells w/23 c/s at end of meiosis I 7.) Sister chromatids separate in anaphase II

Question 8

Trisomy is only observed for which autosomes?

Answer 8

• Most trisomies are not compatible with life - Trisomy 13 = Patau - Trisomy 18 = Edward syndrome - Trisomy 21 = Down syndrome

Question 9

In terms of chromosomal number abnormalities, what are compatible with life, what are not?

Answer 9

• Monosomy always fatal for autosomes - Trisomy 13, 18 and 21 viable - Sex c/s anomalies viable - Triploidy and tetraploidy always lethal

Question 10

What is meant by the term mosaicism? Give an example of a condition in which a significant number of affected individuals exhibit mosaicism.

Answer 10

• Mosaicism are cells within an individual with different genotypes - 2-4% of pts with DS are mosaics, c/s 21 lost during embryogenesis in some cells

Question 11

What is meant by the term proofreading activity as it relates to DNA polymerases?

Answer 11

• DNA synthesis proceeds in 5 to 3’ direction - After each nt added, DNA pol delta checks for correct base pairing - If not correct, 3 to 5’ exonuclease activity of DNA pol delta removes base and tries again

Question 12

Which polymerase has proofreading activity?

Answer 12

• DNA Pol delta

Question 13

Outline the process of strand-directed mismatch repair.

Answer 13

• Newly synthesized DNA contains nicks, which distinguishes parental and daughter strands - Base mismatch distort DNA helical structure - Repair enzymes recognize distortion and remove mismatched base, excise nucleotides back to nearest nick - Gap fill by DNA pol using parental DNA strand as template

Question 14

Describe base excision repair process using the example of the removal of a cytosine residue that has been deaminated to uracil.

Answer 14

• DNA glycosylases recognize diff types of altered bases in DNA and catalyze their hydrolytic removal - Cytosine spontaneously deaminated to uracil - Uracil DNA glycosylase removes uracil base creating AP site - AP endonuclease introduces nick - DNA phosphodiesterase removes remnant sugar and phosphate - DNA Pol and ligase fill in correct base

Question 15

Describe repair of a pyrimidine dimer by process of nucleotide excision repair.

Answer 15

• Helicases and nucleases work together to peel off region containing thymidine dimer lesion - DNA Pol and DNA ligase fill in gap

Question 16

What is xeroderma pigmentosum and what cause it?

Answer 16

• Extreme sensitivity of the skin to sunlight - Pigmentation changes, skin cancers, develop on areas exposed to sun - Result from any of several mutations that impair or abolish the nucleotide excision repair

Question 17

Why are double-strand DNA breaks particularly troublesome? What two avenues of repair are available? Explain what each does.

Answer 17

• No intact template / parental strand to direct repair - Nonhomologous end-joining: broken ends processed, loss of nucleotides at breakpoint, brought together, ligated, ends with permanent change in DNA sequence - Homologous end-joining: repair of damaged chromosome using information contained in undamaged, homologous chromosomes, allows repair of d-s breaks without loss of information

Question 18

How are double-stranded breaks formed?

Answer 18

• Ionizing radiation, oxidizing agents, certain replication errors

Question 19

Name 3 antineoplastic agents that are inhibitors of DNA replication and repair

Answer 19

• Cytarabine - Cyclophosphamide - Doxorubicin

Question 20

Outline mechanism of action of the chemotherapeutic agents cytarabine, cyclophosphamide, and doxorubicin.

Answer 20

1.) Cytarabine is an analogue of cytidine, contains arabinose in place of ribose. Converted to cytarabine triphosphate. Competes with dNTPs for binding to DNA polymerase 2.) Cyclophosphamide is converted to phosphoramide mustard by liver. It is a bi-functional alkylating agent that forms inter and intra strand DNA crosslinks that block replication and can trigger apoptosis 3.) Doxorubicin form tripartite complex with DNA and topo II. Topo II introduces D-S breaks in DNA. Binding doxorubicin prevents re-ligation of D-S breaks. Breaks accumulate and trigger apoptosis