Mutations

What is a mutation? Explain the difference between a gene (or point) mutation and a chromosome mutation, giving an example of each.

In the diagram, sequence A is a normal mRNA fragment and sequences B and C have undergone a mutation. State which type of mutation (substitution, insertion or deletion) has occurred in each of them and explain which one causes a frameshift and why.

What is a structural chromosome mutation? Give two examples of structural alterations of chromosomes (deletion, duplication, inversion, or translocation) and explain briefly what one of them consists of.

Classify the following point mutations according to their effect on the protein: a substitution that does not change the amino acid, a substitution that changes it to another one, and a substitution that turns a codon into a stop codon. State which of the three is usually the most serious and why.

A mutation changes the codon GAA to GAG, and both codons code for the same amino acid (glutamic acid). What is this type of mutation called, and why does it not alter the resulting protein?

A deletion of a single nucleotide occurs in a gene within the coding region. Explain what happens to the reading frame from that point onward, why all the subsequent amino acids can change, and reason whether a deletion of this type located at the beginning of the gene or at the end will be more damaging.

A mutation located in the regulatory region of a gene prevents it from being transcribed, even though its coding sequence is completely normal. Explain how this is possible and compare the effect of this mutation with that of a substitution within an exon of the same gene.

In the CFTR gene, the most common mutation causing cystic fibrosis (ΔF508) consists of the loss of three nucleotides: CTT. The sequence of the coding strand of DNA between codons 506 and 509 is:

5'-... ATC ATC TTT GGT ...-3' (Ile506 - Ile507 - Phe508 - Gly509).

After the deletion of the three central nucleotides CTT (the last base of ATC 507 and the first two of TTT 508), the resulting sequence is: 5'-... ATC ATT GGT ...-3'.

1. a) Knowing that both ATC and ATT code for isoleucine, justify why this mutation does not alter the downstream reading frame and what is the only alteration suffered by the primary structure of the CFTR protein.
2. b) In another patient, a single-nucleotide deletion is detected in that same region (the loss of the 'C' in triplet 507). Explain what consequences this mutation will have on the reading frame, on the downstream amino acid sequence, and on the probability of the protein being truncated.
3. c) There are drugs in development ('correctors') that manage to get the CFTR protein to the cell membrane if it retains its full length, even if it lacks one amino acid. Predict, with reasoning, which of the two patients could benefit from this therapy and why it will be completely ineffective in the other.

In the gene of an essential metabolic enzyme, two independent mutants are analyzed that display alterations at codon number 45, whose original triplet in mRNA is GGU (glycine):

• Mutant 1: has the codon AGU (serine).

• Mutant 2: has the codon UGU (cysteine).

1. a) If residue 45 is located on the surface of the enzyme, exposed to the aqueous medium and far from the catalytic center, reasonably predict which of the two substitutions is less likely to alter the active conformation of the protein.
2. b) It is observed that, upon mild oxidation of the medium, mutant 2 forms giant, inactive molecular aggregates composed of covalent enzyme dimers, a phenomenon absent in the wild type and in mutant 1. Justify this behavior at the chemical level.
3. c) If a third mutant strain displays the triplet GGA at that same codon, what phenotypic consequence will it have on enzymatic activity? Justify your answer by stating the property of the genetic code that explains this fact.

Unit challenge. As a genetic counselor, you must analyze the molecular impact of a real mutation in the human $HBB$ gene (responsible for sickle cell anemia) and propose a biotechnological solution.

1. Data collection: Access the public NCBI ClinVar or Ensembl database, locate the $HBB$ gene, and extract the original triplet of codon 6 versus the pathogenic hemoglobin S variant (c.20A>T, p.Glu7Val in ClinVar; Glu6Val in classical numbering).

2. Flow of genetic information: Prepare a comparative table showing the replication of the DNA fragment (template and complementary strands), its transcription into mRNA, and translation into the resulting peptide. Classify the mutation at the molecular level and predict how this amino acid change affects the three-dimensional conformation of hemoglobin.

3. Regulation and therapeutic report: Write a reasoned report (1.0-1.5 pages) explaining why the disease does not manifest during the fetal stage (gene silencing and regulation of $HBB$ versus fetal globins). Propose a well-founded hypothesis on how to reactivate fetal globin expression using gene editing to treat the disease.

Evaluation criteria:

• Rigor and accuracy in deducing the processes of replication, transcription, and translation from real NCBI data.

• Proper classification of the mutation and its correlation with protein structure.

• Critical integration of gene expression regulation mechanisms into the therapeutic proposal.

Name two mutagenic agents, one physical and one chemical, and briefly explain how they damage DNA.

Mutations can be spontaneous or induced. Explain the difference between them and give an example of a cause of each type.

Cells possess DNA repair systems that correct many of the lesions that escape DNA polymerase. Explain what happens when a lesion is not repaired and explain why people with xeroderma pigmentosum, who have a defective system for repairing the thymine dimers caused by ultraviolet light, show such a high risk of skin cancer.

Retroviruses, such as HIV, copy their genome using reverse transcriptase, an enzyme with no proofreading activity. Explain why these viruses show very high mutation rates, and describe what consequences this fact has for the treatment of the infections they cause and for the development of vaccines.

Explain the difference, in terms of their consequences and their transmission to offspring, between a mutation occurring in a somatic cell and one occurring in a germ cell.

Down syndrome is due to the presence of an extra copy of chromosome 21 (trisomy 21). Explain what type of mutation it is, how it can originate through nondisjunction during meiosis, and explain why mutations of this type affect many traits simultaneously.

Are all mutations harmful? Explain your reasoning and give an example of a mutation that has turned out to be beneficial for a species.

Explain what the mutation rate is and reason why living things maintain low but non-zero mutation rates, relating it to the balance between genetic stability and variability.

Mutations are said to be the primary source of genetic variability. Explain the relationship between mutations, the evolution of species, and biodiversity.

In a region where malaria is endemic, the frequency of the mutant allele $Hb^S$ (responsible for sickle cell anemia in homozygosity) remains stable at around 0.12 due to the selective advantage it confers on heterozygotes against infection by Plasmodium.

1. a) Following an intensive health campaign, malaria is completely eradicated in that region. Reasonably predict how the frequency of the $Hb^S$ allele will evolve over the following generations.
2. b) Justify why the $Hb^S$ allele will not immediately disappear from the population despite being lethal or severely deleterious in homozygosity.
3. c) A person states: "Now that malaria has disappeared, the $Hb^S$ mutation has ceased to exist in the population." Refute this statement by clearly distinguishing between the origin of a mutation and its adaptive value depending on the environment.

To study the origin of antibiotic resistance, an experiment inspired by the Luria-Delbrück fluctuation test was designed: 20 independent tubes were inoculated with an identical and very low number of ampicillin-sensitive bacteria, allowing them to grow to saturation. Simultaneously, a large flask with a volume equivalent to that of the 20 tubes combined was cultured. Equal aliquots were then plated on ampicillin-containing plates:

• Samples from the large flask yielded a very similar number of resistant colonies to one another (mean of 24 colonies, with very low variance).

• Samples from the 20 independent tubes showed enormous fluctuation: most tubes yielded 0 or 1 resistant colony, while a few yielded more than 300 colonies.

1. a) Interpret these results: do they demonstrate that resistance is induced by exposure to the antibiotic or that it is due to pre-existing spontaneous mutations? Justify your answer by explaining the cause of the high variance among the independent tubes.
2. b) Predict what result would have been observed on the plates from the 20 tubes if the presence of the antibiotic were the direct stimulus triggering the mutation in a small, constant fraction of bacteria.

In an experimental evolution assay in the lab, two bacterial populations (standard strain with a normal mutation rate and mutator strain with a 150-fold higher mutation rate due to a defect in DNA repair) are subjected to increasing lethal doses of a novel toxic drug.

1. a) During the initial cycles of exposure, the mutator strain generates viable resistant clones significantly faster than the standard strain. Interpret this result in terms of genetic variability and natural selection.
2. b) After one thousand generations in an optimal and constant medium, without the presence of any toxic compound, the mutator strain experiences a marked decrease in its biological fitness and a population decline compared to the standard strain. Justify this decline based on the nature of the majority of mutations and the concept of mutational load.
3. c) Based on the above results, formulate a hypothesis explaining why natural selection has favored very low mutation rates, but not strictly equal to zero, in living organisms.