Chapter 23

For each DNA repair process in column I, list all characteristics from column II that correctly describe that process. I (a) Nucleotide excision repair (b) Photoreactivation (c) Base excision repair (d) Recombinational repair (e) SOS-driven error-prone repair (f) Alkyltransferase repair (g) Mismatch repair (h) Double-strand break repair II 1\. RecA protein participates. 2\. Damaged nucleotides are removed by nick translation. 3\. A free radical mechanism is involved. 4\. The repair enzyme functions only once. 5\. The key enzyme contains a bound folate cofactor. 6\. No bases or nucleotides are removed from the DNA. 7\. Deficiency of this enzyme in humans greatly increases the risk of skin cancer. 8\. This system is chiefly responsible for the mutagenic effect of ultraviolet light. 9\. This process begins with cleavage of two phosphodiester bonds. 10\. This process begins up to \(1 \mathrm{kbp}\) away from the site to be repaired. 11\. DNA ligase catalyzes the final reaction. 12\. This process also occurs in meiotic recombination. 13\. Replication fork regression might occur during this process.

For each of the following characteristics, list all of the bases to which they apply. (a) A signal that identifies a parental DNA strand in the MutH,L,S mismatch correction system (b) Most likely to be involved in cyclobutane dimer formation after ultraviolet irradiation of DNA (c) A methylated base found immediately to the \(5^{\prime}\) side of dGMP residues in eukaryotic DNA (d) Created by treating DNA with alkylating agents that transfer methyl groups and repaired by an "enzyme" that functions only once in its lifetime (e) Created by AdoMet-dependent methylation of a nucleotide residue in DNA (f) A substrate for deamination at the DNA level, which would lead to a \(\mathrm{GC} \rightarrow \mathrm{AT}\) transition (g) A mutagenic base that can arise in DNA through ROS action.

Homologous recombination in \(E\). coli forms heteroduplex regions of DNA containing mismatched bases. Why are these mismatches not eliminated by the mismatch repair system?

Suppose that you want to study retroviral integration mechanisms by determining the nucleotide sequence at the integration site-several dozen nucleotides on each side of the viral-cellular DNA junction. Describe how to isolate DNA containing a junction site in amounts sufficient for sequence analysis.

Analysis of \(\mathrm{p} 53\) gene mutations in human tumors shows that a large proportion of these mutations involve \(\mathrm{GC} \rightarrow \mathrm{AT}\) transitions originating at sites of DNA methylation. Propose a model to explain preferential mutagenesis of this type at these sites.

Identify and briefly describe three of the processes by which deamination of DNA-cytosine residues by AID could lead to mutagenesis.

In what ways can insertion of a transposon affect the expression of genes in the neighborhood of the insertion site?

Briefly explain how integration of a retroviral genome could activate transcription of genes adjacent to the integration site.

Write a balanced equation for the hydrolytic deamination of a DNA5-methylcytosine residue.

There is evidence that some oxidative damage to DNA occurs at the nucleotide level, with oxidation of a nucleotide, followed by incorporation of the damaged nucleotide into DNA. (a) Describe a pathway by which this could occur. (b) Propose one or more experiments to test whether your proposed pathway does occur.

A mammalian cell line was cultured for many generations in the presence of methotrexate, whose concentration in the culture medium was steadily increased. After maximum resistance to methotrexate had been achieved, the cells were transferred to drug-free medium for many more generations. At the beginning of the experiment, dihydrofolate reductase represented \(0.1 \%\) of the soluble protein in the cell; at maximum resistance it was \(10.0 \%\); and at the conclusion of the experiment, \(1.5 \%\). Describe in qualitative terms the processes involved in generating maximum resistance.

Consider the two processes involved in repairing double-strand DNA breaks. Which process-homologous recombination or NHEJ-is more likely to restore the original gene sequences and why?