E1: Mutation basics

Question 1

Germline vs Somatic Mutation

Answer 1

Mutation: a change in the DNA sequence of a cell. Often associated with
changes in DNA sequence that cause disease.

Germline mutation: those mutations affecting cells that produce
gametes (sperm and ova). These mutations are passed on from
generation to generation.

Somatic mutation: those mutations affecting all cells other than
germline cells. These mutations only affect the individual who has it.

Question 2

What is a polymorphic locus?

Answer 2

Polymorphic locus: DNA sequence variants for a specific gene that are common
in the population. There are usually at least 2 or more alleles for this locus
which have frequencies > 1%. These loci are called polymorphisms.

Question 3

Single-Gene Mutation types (7)

Answer 3

Base-pair substitutions

• Silent: no change in the amino acid sequence
• Missense: change in a single amino acid
• Nonsense: change to a translational stop codon (TAA, TAG, or TGA)

Deletions/Insertions – deletion or insertion of base pairs.

• Results in missing or additional amino acids if multiples of 3 bases are changed

Frameshift mutation:

• Type of deletion/insertion where the changed number of bases is not a multiple of 3.
• This alters the translational reading frame which often results in a premature stop codon being encountered downstream of the mutation site.

Duplications – entire gene(s) is duplicated.

• Leads to a increased gene dosage effect.

Promoter mutations – alters the regulation of transcription or translation.

Splice-site mutations – alters the 5’ or 3’ splice site selection; may activate cryptic splice sites to be generated.

Trinucleotide repeats – insertion of 3 bases into the gene to alter function.

Question 4

Mutation Consequences

Answer 4

Gain-of-Function:

• Over-expression of the normal product
• Inappropriate expression (i.e. in the wrong tissue or stage of development).

Loss-of-Function:

• Decreased amounts of the gene or protein product.
• Sometimes, the situation ( i.e. only 50% protein production) is fine phenotypically; other times, 50% production is problematic for normal function – this is known as haploinsufficiency, which can result in a dominant disorder.

Dominant negative:

• When the mutation causes the protein product not only be nonfunctional, but also inhibits the function of the normal protein produced by the normal allele in the heterozygote.
• Most often occurs in multimeric proteins.

Question 5

Causes of Mutations

Answer 5

Spontaneous mutations:

• Arise naturally during the DNA replication

Induced mutations:

• Caused by mutagens, such as radiation, chemical agents, toxins, nucleotide base analogs (i.e. ara-Cwhich is used in the treatment of AIDS).

Ionizing radiation:

• Produced by x-rays, radioactive nuclear material
• Produces electrically charged ions. When in contact with DNA, the ions can promote changes in DNA chemical bonds, including bonds of double-stranded DNA.

Nonionizing radiation (UV radiation):

• Causes covalent bonds between adjacent pyrimidine bases (cystosine or thymine), known as pyrimidine dimers.
• These dimers are unable to properly base pair with purines during DNA replication, leading to basepair substitution.
• UV radiation is absorbed by the skin and leads to skin cancer, but does not reach germ-line cells.

Question 6

Mutation Rate

Answer 6

At the nucleotide level, the mutation rate is ~ 10-9 per base pair per cell
division (this is the number of mutations that have escaped the DNA repair
process).
This rate varies among genes (10-4 to 10-7 per locus per cell division). This
rate varies for 2 primary reasons:

1. Genes vary in size (i.e. 1000 bp to 2 million bp).
2. Mutation hot spots – nucleotide sequences that are more susceptible to change (i.e. CG dinucleotides).
3. Mutation rates increase with age of the parents.

Question 7

Types of Polymorphisms in a Genome

Answer 7

RFLP – Restriction fragment length polymorphism.

• Changes in endonuclease restriction sites caused by mutations result in different sized fragments when the DNA is cut by that enzyme. Fragments are sized and separated by gel electrophoresis.

VNTR – Variable number of tandem repeats.

• Due to short (10 - 100 bp) sequences repeated numerous times in the genome. Also referred to as minisatellite polymorphisms.

STRPs – Short tandem repeat polymorphisms.

• Similar to VNTRs, but the repeat sequences are 2-6 bp long. Also referred to as microsatellite polymorphisms.

SNPs – Single nucleotide polymorphisms.

• Single bp differences within a population. In humans, there are ~ 3 million SNPs between 2 individuals. This corresponds to SNPs occurring at a rate of ~ 1 in 1000 single bp. These are the most common type of variation in the human genome.

CNVs – Copy number variants.

• Differences in the number of repeat DNA segments that are > 1000 bp, They can be present in some people, but absent in others. It is estimated that CNVs account for at least 4 million bps of differences between individuals.