Genetics 11 - Genetic Diversity and Complex Genetic Diseases Flashcards
Composition of antibody or Ig molecule
composed of four chains: an identical pair of longer heavy chains and an identical pair of shorter light chains, which are linked together by disulfide bonds
Types of heavy chains
5
termed γ, μ, α, δ, and ε
Types of light chains
κ and λ
γ
IgG
μ
IgM
α
IgA
δ
IgD
ε
IgE
What do immature B lymphocytes produce
ONLY IgM
but as they mature, a rearrangement of heavy chain genes called class switching occurs
Class switching
Produces the other 4 major classes of immunoglobulins, each of which differs in AA composition, charge, size, CHO content
Each class tends to be localized in certain parts of the body, and each tends to respond to a different type of infection
The two types of light chains can be found in association with any of the five types of heavy chains
Relationship between light and heavy chains
2 types of light chains can be found in association with any of 5 types of heavy chains
What determines the major class to which an Ig belongs
Heavy chain constant region
What part of Ig recognises and binds antigens
Variable regions of the light and heavy chains
Production rate of humoral immune system
can generate 100 billion structurally distinct antibodies
Why is the 1 gene-1 antibody hypothesis incorrect
because the human genome has only 20,000 to 25,000 protein-coding genes
Multiple Germline Immunoglobulin Genes
Molecular genetic studies (cloning and DNA sequencing) have shown that for each heavy and light chain, an individual has more than 80 different V segments located continuously in his or her germline and 6 different J segments
There are at least 30 D segments in the heavy chain region
Somatic Recombination - VDJ Recombination
Specific combination of single V and J for light chain and separately, V, D and J for heavy chain
accomplished by deleting the DNA sequences separating the single V, J, and D segments before they are transcribed into mRNA
→ by recombinases (encoded // RAG1 and RAG2 genes) which intitiate 2x strand DNA breaks at specific DNA sequences that flank V and D segments
What does mutation of RAG1 and RAG2 genes lead to
severe combined immunodeficiency disease (SCID) as the recombination mechanisms are also involved in generation of T cell receptors (as well as recombinases)
When are Ig molecules formed
During B lymphocyte maturation
What happens after deletion of all but 1 V, D and J segment
Non-deleted segments are joined by ligases
This cutting-and-pasting process is known as somatic recombination (in contrast to the germline recombination that takes place during meiosis)
unlike most other cells of the body, whose DNA sequences are identical to one another, mature B lymphocytes vary in terms of their rearranged immunoglobulin DNA sequences
Many possible combinations of single V, J and D segments, somatic recombination can generate 100,000-1,000,000 different types of antibody molecules
Junctional diversity
As the V, D, and J regions are assembled, slight variations occur in the position at which they are joined, and small numbers of nucleotides may be deleted or inserted at the junctions joining the regions
This creates even more variation in antibody AA sequence
binding affinity between B cells and their cell surface receptors (Igs)
What happens upon binding
only a small subset of B cells has cell-surface receptors (immunoglobulins) that can bind to a specific foreign antigen, and their binding affinity is usually low
Once this subset of B cells is stimulated by a foreign antigen, they undergo an affinity maturation process characterized by somatic hypermutation of the V segments of immunoglobulin genes
Action of activation-induced deaminase
cytosine bases replaced by uracil
Errorprone DNA polymerases are recruited, and DNA repair processes are modified so that mutations can persist in the DNA sequence
consequences of persistence of mutations in DNA sequence
the mutation rate of these gene segments is approximately 103 per base pair per generation (recall that the mutation rate in the human genome is normally only 108 per base pair per generation)
This causes much additional variation in immunoglobulin-encoding DNA sequences and thus in the antigen-binding properties of the encoded immunoglobulins
Effect of somatic hypermutation on Igs
somatic hypermutation produces a subset of immunoglobulins that have high-affinity binding to the foreign antigen, and the B cells that harbor these immunoglobulins are selected to proliferate extensively
The end result is a population of mature plasma cells that secrete antibodies that are highly specific to the invading pathogen
Multiple combinations of Heavy and Light Chains
Further diversity is created by the random combination of different heavy and light chains in assembling the immunoglobulin molecule
Each of these mechanisms contributes to antibody diversity
Considering all of them together, it has been estimated that as many as 1011 distinct antibodies can potentially be produced
5 mechanisms that produce antibody diversity
- multiple germline immunoglobulin gene segments
- somatic recombination of the immunoglobulin gene segments
- junctional diversity
- somatic hypermutation
- potential for multiple combinations of heavy and light chains
Similarities & differences between T cell receptors and Igs (B cell receptors)
T-cell receptors must be able to bind to a large variety of peptides derived from invading organisms
Unlike immunoglobulins, however, T-cell receptors are never secreted from the cell, and T-cell activation requires the presentation of foreign peptide along with an MHC molecule
Generation of T cell diversity by
Most of the mechanisms involved in generating immunoglobulin diversity—multiple germline gene segments, VDJ somatic recombination, and junctional diversity
What occurs with B cell receptor diversity but NOT t cell receptor diversity
somatic hypermutation does not occur in the genes that encode the T-cell receptors
This is thought to help avoid the generation of T cells that would react against the body’s own cells (an autoimmune response)
Copy Number Variants
CNVs
deletions and duplications of genomic sequence, ranging in length from a kilobase to multiple megabase pairs
They are known as major contributors to human genetic diversity
CNVs are known to influence both normal and disease variation
Molecular cytogenic techniques enabled
detection of submicroscopic deletions that encompass many genes and have typical characteristics of “chromosomal disorders,” even though specific features may be due to haploinsufficiency of specific genes
How have bridged and blurred the gap between chromosomal and monogenic conditions
recent advent of array technology allows for the detection of smaller and smaller copy number variants (CNVs)
prevalence of CNVs
most individuals (65% to 85% of the population) harbour a CNV of at least 100 kb of DNA
Much larger (exceeding 500 kb) variants occur in 5% to 10% of individuals
at least 1% of the population carries a CNV exceeding 1 Mb
While some of these CNVs are clearly pathogenic, others may represent risk modifiers for common diseases or may be completely benign
What do CNVs cause
Severity
CNVs can cause structural chromosome abnormalities via duplication or deletion
The clinical severity largely depends on the size of the respective chromosome segments
Smaller CNVs (< 10kb) - class of CNVs
What are they associated with
copy number polymorphisms (CNPs) because they are common in the general population (>1% frequency)
A subset of these CNPs are highly variable with respect to copy number
They often encode proteins involved in drug metabolism and immunity and are associated with susceptibility to complex inflammatory or immune disease such as psoriasis and Chron’s
2nd class of CNVs - larger (> 10kb but < 5 Mb)
Prevalence
also known as
How do they occur
how were they discovered
rare in the general population
They ae also known as microdeletions or microduplications, and usually have a much more recent origin within a family, sometimes occurring as a de novo mutation in the gamete
The era of molecular cytogenetics, with newer molecular methods of genomic quantification (e.g., fluorescence in situ hybridization [FISH] and array analysis), led to the molecular characterization of microdeletion and microduplication syndromes (genetic syndromes caused by a deletion that is not visible by light microscopy)
Microdeletions/duplications are associated with
susceptibility to neurocognitive diseases including autism spectrum disorders and schizophrenia
Examples of microdeletion syndromes
Williams Syndrome (Micro del 7q11.23)
Smith-Magenis Syndrome (Micro del. 17p11.2)
Potocki-Lupski Syndrome (Micro dup. 17p11.2)
Occasionally what might a microdeletion cause
Haploinsufficiency of several adjacent genes, of which 2+ are associated to distinct genetic disorders, including TSC2 (causing tuberous sclerosis) and PKD1 (causing autosomal dominant polycystic kidney disease) on chromosome 16p13
2 contiguous gene syndromes
tuberous sclerosis - TSC2 on chr 16p13
autosomal dominant polycystic kidney disease - PKD1 on chr 16p13
what is the phenotype of monogenic disorders influenced by
exogenous factors - otherwise treatment would not work
Phenylketonuria (PKU) - PAH, Chr 12q
what sort of disorder is it
how is it influenced
monogenic disorder
the clinical phenotype is fundamentally influenced by an exogenous factor (i.e., reduced intake of phenylalanine)
While untreated individuals with classic PKU develop a severe intellectual disability, a person with PKU who has been treated with an optimal diet is, clinically, not different from a “healthy” person
In this way PKU is similar to a risk factor for a disorder; however, in the absence of treatment the risk of disease is far larger than with classic multifactorial disorders
Genetic disorder
disorder = clinical symptoms of a person, not to his or her genome
asymptomatic mutation carrier and reduced penetrance
A carrier for myotonic dystrophy falls ill when the first symptoms occur; before that he or she is an asymptomatic mutation carrier
Some carriers of dominant disease-causing mutations never fall ill (reduced penetrance)
What does detection of a mutation imply
especially during predictive testing, may only imply an increased risk or probability for the disorder
Multifactorial disorders
examples
= complex disorders
those disorders that do not follow a clear inheritance pattern and whose clinical symptoms are suspected to have been caused by the interplay between several genes (polygenic) and exogenous (environmental) factors
T2D
Coronary artery disease
Inflammatory bowel disease
cancers
Polygenic theory
useful framework for considering the inheritance patterns of traits/disorders that rely on the interaction of a large number of genetic factors, each of which makes a small contribution to the overall phenotype
2 main concepts of polygenic theory
Heritability – The proportion of the total phenotypic variance that is attributable to genetic variance in a population
Estimates how much of the differences between people in a social group are down to genetic differences between them, and how much is down to differences in their environments (i.e. nature vs nurture)
relates to population > individuals
estimated by comparing the incidence of a condition in relatives of affected people with the incidence of the condition in the general population - relevance of genetic factors to a condition in specific place/time
if a phenotypic trait has a heritability of 0.25, that means that 25% of the variability in that trait in the population is down to genetic differences among the people e.g. PKU - depends on intake of phenylalanine
Threshold effect as a principle of polygenic theory
explains how dichotomous characters can be polygenic
susceptibility to a disease is a continuous character that depends on combined effect of many genes
According to this concept, a certain polygenic/complex trait (for example, a congenital malformation) becomes phenotypically evident only once the threshold of genetic predisposition is exceeded; when this happens, the effect is complete (i.e., the all-or-nothing principle)
What happens when you’re susceptibility exceeds a threshold
Complex diseases and prevalence in one’s family
you will manifest the disease
relatives will therefore be more likely to also manifest the disease than the general population as they are more likely to share your high risk alleles (probability)
Therefore complex diseases tend to run in families, but this tendency is much weaker than with Mendelian diseases
Parents with multiple affected children
Higher risk of future recurrence
Close relatives of more severely affected individuals
also at greater risk of manifesting the phenotype. Likewise, closer relatives of the index case are more at risk than more distant relatives
gender dependence
In the case of some conditions, the threshold of genetic predisposition seems to differ between males and females
e.g. Hypertrophic Pyloric Stenosis - males have a lower threshold of genetic predisposition and as such are more likely to manifest the clinical phenotype
herefore, if a female is affected it is likely that she was dealt a particularly band genetic “hand” from her parents and the recurrence risk will be higher in her descendants
Carter effect
: Multifactorial diseases with a gender-dependent threshold effect have a higher recurrence risk if the index patient is of the less commonly affected sex
Incidence of hypertrophic pyloric stenosis
t its incidence is highest in sons of affected women and lowest in daughters of affected men
Hypertrophic pyloric stenosis
postnatal hypertrophy of the circular muscle of the pylorus that results in functional obstruction of the pylorus, projectile vomiting, and failure to thrive
Cumulative freq = 1 in 1,000
boys are affected 5x more frequently than girls
In most cases, the disorder is diagnosed in infants between 3 and 12 weeks of life
Sometimes the pylorus can be felt as a cylindrical tumor in the upper right quadrant of the abdomen
Water and salt deficiency can result in a hypochloremic alkalosis
The treatment of choice is pyloromyotomy (i.e., Ramstedt procedure)
Why don’t monozygotic/’identical’ twins share 100% of their genes
(originate from same fertilised zygote)
CNVs and somatic mutation of the immune receptor genes
(Nonetheless, MZ twins do share homology at the majority of loci, and by studying differences in MZ twin genomes and relating it to phenotype, we can identify genetic changes implicated in disease onset)
Discordance
presence of a given genetic or phenotypic trait in only one member of a pair of MZ twins (called discordance) suggests environment strongly effects such a trait (low heritability)
⇒ end result of many twin studies is an estimate of the heritability of the condition
A classic twin study design compares many pairs of MZ and fraternal (dizygotic - DZ) twins, where one twin in each pair is known to be affected
The frequency that the co-twin in each pair is also affected is investigated, with the assumption being that both MZ and DZ twins share their environment to the same degree
Higher concordance (both display the phenotype) in MZ twins than DZ twins is evidence for genetic factors – i.e. heritability in the trait
Linkage
relationship between loci (not alleles)
Linkage analysis
looks at physical chunks of the genome of related individuals with the phenotype and associates them with given traits
Principle: if we find a common genetic marker (e.g. microsatellite, SNP), we assume that the gene that causes the disease is somewhere in the same area
What is linkage useful for
associating large sections of the genome with highly penetrant disorders/traits shared between relatives, but it is also useful for identifying chromosome sections to target for association analysis in unrelated or more distantly related groups of people
Resolution of linkage
Low
we can link a haplotype or section of chromosome to a trait, but cannot zero in on the exact gene without more information
Genetic Association Studies
purely statistical phenomenon and not specifically genetic
Association is when one or more genotypes (alleles) within a population co-occur with a particular phenotypic trait more often than would be expected by chance
It is important to stress that association is not causation
Association studies seek to identify particular allele/marker variants that are associated with the phenotype at the population level
Principle of genetic association studies
What are they used for
gather some people with a disease (cases) and some people without a disease (controls) and look to see what alleles are present more in cases than controls
Association is used to find common low penetrance alleles associated with a disease
causes of association between a genetic variant and a disease may have several causes
- The variant may directly confer disease susceptibility
- The variant may be on the same shared ancestral Chr segment as the variant that directly causes disease
- The variant may be more frequent in the population subgroup in which the disease is also more frequent
Positional cloning
process of identifying a disease gene based on its location on a chromosome
Linkage and association are complimentary methods to describe the aetiology of complex traits -
By using linkage analysis we can define large candidate genome regions - Usually by analysis of a genome wide panel of a few hundred markers in affected sibling pairs - Positional cloning can then be used to zero in on the position of interest using high density SNP genotyping
dbSNP database
has 10 million SNPs – one for every 300 bp of human DNA
In this way, we can associate short shared segments of a few Kb with a disease phenotype
Next candidate genes in this section are analysed for mutations (candidate gene analysis)
So by knowing only a genes’ location on a chromosome, researchers can identify or “clone” a gene using positional cloning
Functional cloning
Cloning a gene required knowledge of the disease pathology
Prior knowledge of what the gene function is was essential in order to make an educated guess
Genome Wide Association Studies (GWAS)
GWAS typically test hundreds or thousands of individuals and scans all genes in the genome to look for associations between variants (alleles) and phenotypic traits
No prior knowledge is needed, just a large number of cases and controls
High resolution SNP chips are usually used - 1 for every 3000 bps of human genome
Gold standard for linkage/GWAS
replication of linkage/association in different populations
What is GWAS used for
calculate individual genome-wide polygenic risk scores (GPSs) for complex polygenic diseases such as coronary artery disease, type 2 diabetes, inflammatory bowel disease and cancer
Sophisticated algorithms were used to integrate pervariant risk scores from hundreds of thousands of individuals into one GPS associated risk, which was found for many individuals to be as good a predictor of disease as some single mutations causing rare Mendelian diseases already routinely considered in clinical settings
