Biomedical Sciences I: Mutations and Polymorphisms (Lecture 4; Exam 1) Smozcer

Locus (loci)

DNA segment that occupies a specific position on a chromosome.

What are alleles?

Alternative versions of DNA sequence at a specific locus.

(2 alleles for each locus; maternal and paternal)

Polymorphic

Multiple versions of the allele that are very common in the population.

What is a wild-type allele?

The allele that encodes the most common phenotype or genotype among a particular population and is considered the standard or norm.

Mutant/Variant alleles

The original change in the DNA base sequence that creates a new or altered allele.

What are “private” alleles?

Rare variants that are confined/appear only in a specifically defined family.

Zygosity

Degree of allele similarity at one locus in an organism.

Homozygous

Two copies of same allele at one locus.

Heterozygous

Two different alleles at same locus.

Genotype

Genetic information at a locus.

Phenotype

Appearance of an organism based on their genotype; set of observable characteristics.

What is the source of genetic diversity?

Mutations.

How much sequence identity is shared among humans?

99.9%

What percentage of DNA is genetically determined variability?

0.1%

T/F: There is a standard genomic sequence.

False. There is no standard genomic sequence.

The most common sequence in a population is called:

Reference sequence.

Mutations Classifications:

Classified by:

• Size

• Function

• Heritability

Euploidy

The state of a cell or organism having a complete balanced set - or multiples of a set of chromosomes.

Aneuploidy

A genetic condition where a cell has an abnormal number of chromosomes.

(either missing a chromosome or having an extra one)

An example of euploidy:

Tetraploidy - rare chromosomal condition where cells contain 4 complete sets of chromosomes (92 total) instead of 46 total.

Examples of Aneuploidy:

• Trisomy

• Monosomy

Subchromosomal Mutations:

Changes in portions of the chromosomes; copy number of variations, structural rearrangements

(structural genetic changes smaller than a whole chromosome that involve the gain, loss or rearrangement o specific chromosomal segments)

Mutation rate:

The frequency of new genetic changes in a gene, nucleotide sequence, or organism over a specific unit of time or replication cycle.

Hot Spots: Mutations

Specific positions in a DNA sequence that show a much higher rate of mutation than other areas of the genome.

Mutation Frequency:

Number of mutations / locus / cell division dependent on:

• Frequency of spontaneous and induced nucleotide changes

• Probability of repair

• Probability of detection

Rate of disease-causing mutations:

Incidence of new cases of genetic disease NOT present in parents and caused by a single mutation.

How do chromosome mutations occur?

Result of chromosome mis-segregation during meiosis.

T/F: Chromosome mutations are generally severe.

True. They can result in spontaneously aborted fetuses.

How do regional mutations occur?

Result of homologous recombination between fragments with high homology at different sites or following repair of double-strand breaks.

Replication Mutation rate:

• Replication errors = <1 mutation/genome/cell division (Very low)

• DNA repairs ; occur frequently from spontaneous mutations that evade the repair machinery.

Types of Mutations:

• Nucleotide Substitutions

• Deletions, insertions, rearrangements

Nucleotide Substitutions:

• Synonymous mutations

• Missense mutations

• Nonsense mutation

• Mutation affecting mRNA processing

Synonymous mutations

Nucleotide change that specifies the same amino acid.

AAG → AAA = (Lys → Lys)

Missense Mutations

Single nucleotide change that specifies a new amino acid.

• Transitions

• Transversions

TCC → TGC = (Arg → Thr)

Nonsense Mutation:

Point mutations resulting in replacement of coding codon by a stop codon.

Mutation affecting mRNA processing:

Mutations that abolish or create alternative intron-exon junctions → alterations of the splicing pattern.

Frameshift Mutations

A genetic change caused by the insertion or deletion of a number of nucleotides that is NOT a multiple of three, shifting the way the genetic code is read.

What are the most common rearrangement muations?

Frameshit mutations.

Functional Consequences of Mutations:

• Gain-of-function

• Loss-of-function

• Dominant negative

• Lethality

Gain-of-function:

Overproduction / inappropriate production of protein.

Loss-of-function

Reduced production of protein.

(The product of the normal allele is generally sufficient for function)

Haploinsufficiency

When 50% of the protein is insufficient for function.

Dominant negative

Mutate protein inhibits the product of the normal allele in heterozygosity.

(A mutated gene makes a faulty protein that actively blocks or ruins the work of the normal protein made by the healthy gene.)

Germline Mutations:

Mutations inherited from parents or de novo mutations that are transmitted to offspring.

(De novo mutations are VERY rare.

Somatic Mutations

An acquired change in the DNA sequence of a body cell that happens after conception and is never passed on to children.

**Are not transmitted to the next generations.

What mutations are frequent in cancers?

Somatic mutations.

What are genetic polymorphisms?

Mutations with frequency exceeding 1% of all alleles in a population (regardless of the type, size, effect, or location of mutation).

Examples of Genetic polymorphisms:

1. Single nucleotide polymorphisms (SNPs)

2. Insertion-deletion polymorphisms (indels)

3. Copy number variants (CNVs)

4. Inversion polymorphisms

Single Nucleotide Polymorphisms (SNPs):

Change of 1 base pair.

• Average 1 for every 1000bp (approx 5-10million SNPs/genome)

• Generally do not produce phenotypic differences

• 25x higher rate at adjacent CG (CpG) (hot spots**)

• Approx. 100,000 SNPs in protein-coding genes

SNPs Mutations:

• Synonymous

• Nonsynonymous → leads to protein variants

Insertion-deletion polymorphisms (indels):

Up to 1000 base pairs.

• Simple - Presence or absence of a short fragment → 2 alleles

• Microsatellites - Variable number of repeated short segments (2, 3, or 4 nt) → multiple alleles

Copy number variants (CNVs)

Up to hundreds of kb.

• Can include dozens of genes → altered gene dosage

Inversion polymorphisms

Few bp-mb.

• Sequence homology at edges (homologous recombination)

• Balanced - no loss / gain of DNA

Detection Process:

1. Discovery

2. Validation

3. Screening

Detection Process: Discovery

• Initial identification

• Whole genome / whole exome sequencing + comparison to reference sequence

Detection process: Validation

Replication assay to exclude sequencing errors

Larger population to get statistical occurrence

Detection process: Screening

• Sequencing / analysis of thousands of SNPs from same individual and multiple individuals

• High - density DNA arrays (SNP arrays)

Methods for mapping human disease variants:

• Linkage analysis (family-based)

• Association analysis (population-based)

• genome sequencing

Genome-wide association studies:

Molecular technique that analyzes simultaneously hundreds of thousands (even millions) of variations in genomic DNA to determine if a genetic locus is associated with a certain phenotype.

What is a candidate gene?

A specific gene suspected of playing a direct tole in determining a particular trait, physical attribute or disease.

(Have greater power but rely on previous knowledge.)

What is GWAS?

A research method used to scan entire genomes and find genetic differences linked to specific diseases or traits.

What is SNP imputation?

A statistical method used to predict or fill in missing and unmeasured genetic variants in a dataset.

Linkage Disequilibirum:

Non-random association of alleles at linked loci.

• Measure of tendency of some alleles to be inherited

• Haplotypes: Sets of closely linked SNPs present on the same chromosome which tend to be inherited together.

Manhattan Plot

Scatter plot of association between statistical significance as p-value on the y-axis against chromosomes on the x-axis.

Association is considered statistically significant if:

p = 5 × 10^-8

If a SNP is significantly associated with the phenotype:

• Casual relationship between SNP and disease

• Marker in linkage disequilibrium with casual locus

• False positive

Clinically functional variants;

• Risk variants - increase risk of disease

• Protective variants - lower risk of disease

• Risk of disease - diagnosis of disease

23andme

Testing of genome-wide polymorphisms:

• Problems: privacy, lack of regulation

• Traits

• Carrier Status

• Ancestry **

• Drug response **

Clinical Utility / Limitations:

• Gene-disease associations not relevant for all patients

• Environmental role unknown and hard to predict and estimate

• Relevance only for prognosis (cofounding factors can change risk)

• Combined risk from multiple SNPs is hard to calculate