17. Mutation, Repair, and Recombination

Types of Mutations

Open Question
Mutations in the IL2RG gene cause approximately 30 percent of severe combined immunodeficiency disorder (SCID) cases in humans. These mutations result in alterations to a protein component of cytokine receptors that are essential for proper development of the immune system. The IL2RG gene is composed of eight exons and contains upstream and downstream sequences that are necessary for proper transcription and translation. Below are some of the mutations observed. For each, explain its likely influence on the IL2RG gene product (assume its length to be 375 amino acids).

Insertion in Exon 1, causing frameshift
Open Question
Mutations in the IL2RG gene cause approximately 30 percent of severe combined immunodeficiency disorder (SCID) cases in humans. These mutations result in alterations to a protein component of cytokine receptors that are essential for proper development of the immune system. The IL2RG gene is composed of eight exons and contains upstream and downstream sequences that are necessary for proper transcription and translation. Below are some of the mutations observed. For each, explain its likely influence on the IL2RG gene product (assume its length to be 375 amino acids).

Nonsense mutation in a coding region
Open Question
Alkaptonuria is a human autosomal recessive disorder caused by mutation of the HAO gene that encodes the enzyme homogentisic acid oxidase. A map of the HAO gene region reveals four BamHI restriction sites (B1 to B4) in the wild-type allele and three BamHI restriction sites in the mutant allele. BamHI utilizes the restriction sequence 5′-GGATCC-3′. The BamHI restriction sequence identified as B3 is altered to 5′-GGAACC-3′ in the mutant allele. The mutation results in a Ser-to-Thr missense mutation. Restriction maps of the two alleles are shown below, and the binding sites of two molecular probes (probe A and probe B) are identified.DNA samples taken from a mother (M), father (F), and two children (C1 and C2) are analyzed by Southern blotting of BamHI-digested DNA. The gel electrophoresis results are illustrated.Explain how the DNA sequence change results in a Ser-to-Thr missense mutation.
Open Question
In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.Characterize the expected distribution of colony growth on the four plates. Defend your growth prediction for each plate.
Open Question
In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.What event is being detected by growth of a colony on any of the four plates?
Open Question
In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.Why is the S9 extract added to each of the plates?
Open Question
In an experiment employing the methods of the Ames test, two strains of Salmonella are used. Strain A contains a base-substitution mutation, and Strain B contains a frameshift mutation. Four plates are prepared to test the mutagenicity of the compound ethyl methanesulfonate (EMS). Plate 1 is a control plate with Strain A and S9 extract but no EMS. Plate 2 is also a control plate and contains Strain B and S9 extract but no EMS. Plate 3 contains Strain A along with S9 extract and EMS, and Plate 4 contains Strain B, S9 extract, and EMS.Suppose the compound being tested was proflavin instead of EMS. Would this change the Ames test results? Explain why or why not.
Open Question
Infantile cardiomyopathy is a devastating disorder that is fatal during the first year of life due to defects in the function of heart muscles resulting from mitochondrial dysfunction. A study, performed by Götz et al. [(2011). Am. J. Hum. Genet. 88:635–642), identified two different causative mutations in the gene for mitochondrial alanyl-tRNA synthetase (mtAlaRS). One mutation changes a leucine residue at amino acid position 155 to arginine (p.Leu155Arg). The other mutation changes arginine at position 592 to tryptophan (p.Arg592Trp). The mtAlaRS enzyme has an N-terminal domain (amino acids 36–481) that catalyzes tRNA aminoacylation and an internal editing domain (amino acids 484–782) that catalyzes deacylation in the case that the tRNA is charged with the wrong amino acid.Consider the position of the disease causing missense mutations in the mtAlaRS gene in the context of the known protein domains of this enzyme. What predictions can you make about how these mutations impair protein synthesis within mitochondria in different ways?
Open Question
Infantile cardiomyopathy is a devastating disorder that is fatal during the first year of life due to defects in the function of heart muscles resulting from mitochondrial dysfunction. A study, performed by Götz et al. [(2011). Am. J. Hum. Genet. 88:635–642), identified two different causative mutations in the gene for mitochondrial alanyl-tRNA synthetase (mtAlaRS). One mutation changes a leucine residue at amino acid position 155 to arginine (p.Leu155Arg). The other mutation changes arginine at position 592 to tryptophan (p.Arg592Trp). The mtAlaRS enzyme has an N-terminal domain (amino acids 36–481) that catalyzes tRNA aminoacylation and an internal editing domain (amino acids 484–782) that catalyzes deacylation in the case that the tRNA is charged with the wrong amino acid.

Which mutation would you predict has a more severe impairment of translation in mitochondria, and why?
Open Question
Base-substitution mutations often change the amino acid specified by a codon. For each of the amino acid changes listed, determine which ones can result from a one–base-pair substitution. For those that can result from a one–base-pair substitution, give the possible wild-type and mutant codons, listing multiple possibilities if there is more than one option. (Use either Figure 9.13 or the genetic code in Table A to help solve this problem).

Wild-type Mutant
a. Ser Ala
b. Cys Ser
c. Pro Glu
d. Lys Stop
e. Met His
f. Met Ile
Open Question
The two gels illustrated contain dideoxynucleotide DNA-sequencing information for a wild-type segment and mutant segment of DNA corresponding to the N-terminal end of a protein. The start codon and the next five codons are sequenced.

What is the cause of the mutation?