Genomics Flashcards
What are monogenic diseases?
Caused by mutations in a single gene; single mutation is both necessary and sufficient
What is genetic linkage?
DNA regions that are located in close proximity are more likely to be co-inherited than DNA regions originating further apart
What is the LOD score?
Detects presence of linkage; LOD = log10 (L(linkage)/L(no linkage)); more than 3 –> linkage; less than -2 no linkage
Which things can act as confounders in single pedigrees?
Non penetrance and phenocopies
What is Hajdu Cheney syndrome?
severe osteoporosis; NOTCH2 heterozygous mutation; autosomal dominant
What is a confounding factor in linkage analysis performed in multiple pedigrees?
Genetic heterogeneity
What is the difference between polymorphism and mutation?
Polymorphism: common DNA variant
Mutation: pathologic DNA variant
What is transition and transversion in single nucleotide substitutions?
Transition: substitution which conserves the base chemistry (G–>A; T–>C), twice as common as transversions
Transversion: substitution which changes the base chemistry (G–>T; A–>C)
What are the consequences of variants in splice sites?
Exon skipping, use of cryptic splice site (in exon or intron), intron retention, combination of these
What are the rarer types of functional variation?
Variants in promoters, untranslated regions (myotonic dystrophy and fukuyama muscular dystrophy), polyadenylation signals
What are the types of functional DNA variation defined by Muller?
Loss of function: amorph (complete loss of function), hypomorph (partial loss of gene function)
Gain of function: hypermorph (increase in normal gene function), antimorph (dominant alleles that act in opposition to normal gene activity), neomorphic (dominant gain in gene function that is different from the normal function)
What is NMD?
non-sense mediated decay
When do ‘loss of function’ alleles exhibit dominant forms of inheritance?
- Haploinsufficiency
- “Dominant negative” effect: homodimeric complex disrupts function of its normal counterpart
- Somatic second hits: cancer gene (recessive at cellular level but dominantly inherited in families)
What is infantile onset epilepsy in Amish?
Autosomal recessive due to nonsense mutation in SIAT9 encoding GM3 synthase –> less production of gangliosides (membrane stability); seizures starting at 6 months of age
What are the differences between phenotypic and genetic heterogeneity?
Phenotypic: one gene, mutations with different clinical outcomes (phenotypes)
Genetic: different genes but same clinical outcomes
What is penetrance?
The proportion of carriers who manifest phenotypic signs of the condition
What is Marfan syndrome?
Tall stature, long limbs, chest abnormalities, scoliosis, arachnodatcyly (long fingers), dislocated lens, risk of aortic aneurism
What is expression?
Gene carriers can be affected to different degrees; extent of clinical manifestation
What are examples of diseases with de novo mutations?
Achondroplasia, marfan syndrome, neurofibromatosis I and II
What is germline mosaicism?
A parent carries a small portion of gametes that harbour the same mutation (achondroplasia, osteogenesis imperfecta, duchenne muscular dystrophy)
How does cosanguinity affect the risk of recessive disorders in the offspring?
increases risk
What are compound heterozygosity and homozygosity?
Heterozygosity: someone who has different allelic mutations at the same locus, i.e. cystic fibrosis
Homozygosity: same allele on both chromosomes, i.e. sickle cell anemia
How can females be affected with X-linked recessive disorders?
- Non-random inactivation leading to chance expression in certain tissues
What is X inactivation?
One X in each cell is switched off before blastocyst implants in female embryos
What are lines of Blaschko?
Territories of clonal cell populations (incontinentia pigmenti, goltz focal dermal hypoplasia)
What are multifactorial disorders?
Polygenic, many different genes and environmental factors contribute to disease susceptibility; asthma, breast cancer, heart disease, depression, scizophrenia, autism, arthritis, migraine, obesity, diabetes, stroke…
What are the major genes in breast cancer?
BRCA1 and 2; mutations in these genes are rare with many different mutations in each gene; high penetrance, lower penetrance for ovarian cancer, BRCA2 has lower penetrance than BRCA1
What are the genes involved in Alzheimer’s disease?
Three rare, high risk genes (APP, PSEN1, PSEN2): young onset AD, high risk families
Moderate risk genes: APOE allele e4, TREM2 gene
Many common, low risk variants identified through association studies
What are the three alleles in APOE gene and what is their importance?
e4: increased risk of AD
e3: baseline risk
e2: protective against AD
Which statistical test is used in case-control association studies?
Chi-square test: test whether distribution of genotypes or alleles is the same in cases and controls
Which are the genes involved in rheumatoid arthritis?
PTPN22: encodes protein tyrosine phosphatase which interacts the negative regulatory kinase Csk to inhibit T cell signalling and activation
What are the measures used to communicate the contribution to disease in Mendelian and polygenic disorders?
Mendelian: absolute risk
Polygenic: relative risk (odds ratios)
What are the genetic tests used in diagnosis?
- Standard chromosome analysis
- Fluorescent in situ hybridisation (FISH)
- Comparative genomic hybridisation array (Array CGH)
- Single gene testing
- Next generation sequencing
- Methylation analysis (imprinting disorder)
What is VATER (VACTERL) association?
Vertebral Anal atresia Cardiac Trachea-oesophageal atresia/fistula Renal abnormalities Limb deformities (radial ray defect)
What is velocardiofacial syndrome?
CATCH 22; del22q11.2 syndrome; CHD (conotruncal anomaly, celft lip and/or palate, absent thymus, absent parathyroid glands, dysmorphic features, learning difficulties
What is comparative genomic hybridisation array (aCGH)?
Powerful tool for identify small genomic imbalances
What is del16p11.2?
Learning difficulties, ASD, seizures, schizophrenia, bipolar disorder, depression, obesity in young adults, unaffected carriers
What is achondroplasia?
FGFR3; most common skeletal dysplasia, disproportionally short limbs, large head, normal intelligence, usually new dominant mutation, unaffected parents 1% RR, affected parents 50% RR
How do you calculate the risk of a child being affected in incomplete penetrance?
probability of inheriting mutant allele*penetrance
In X-linked inheritance with a female carrier, what are the chances of having an affected offspring, having unaffected daughters that are carriers, having unaffected sons that are not carriers?
affected offspring: 1/4
unaffected daughters that are carriers: 1/2
unaffected sons, not carriers: 1/2
A consuldand has a mother that is a carrier for ocular albinism (X-linked recessive), what are the chances of her having an affected child with an unaffected husband?
chances of her being a carrier: 1/2
chances of her passing on the gene: 1/2
chances of her child being a boy: 1/2
chances of having an affected child: 1/21/21/2 = 1/8
What is the Hardy-Weinberg principle?
model which helps to explain gene frequencies in a population (useful for calculating risks of autosomal recessive disorder)
p + q = 1
p^2 + 2pq + q^2 = 1
Carrier frequency = 2pq
If cystic fibrosis has an incidence of 1 in 2500, what is the carrier frequency?
q^2 = 1/2500
q = 1/50
because q is small, carrier frequency = 2*(1/50) = 1/25
Knowing that the population carrier risk of cystic fibrosis is 1/25, what are the chances that a couple will have an affected offspring if the mother is unaffected, but has an affected sister and there is no family history about the father?
chances of mother being a carrier = 2/3
chances of father being a carrier = 1/25
chances of having an affected offspring = 2/31/251/4 = 1/150
What are the two main types of genotype inheritance models?
Additive genetic effect and dominant genetic effect
What are genome-wide association studies?
GWAS search for causal variants associated with phenotypes everywhere in the genome (not only genes); only prerequisite: trait is known to be heritable
What are the P values for polygenic risk scores vs significant SNPs?
polygenic risk scores: 10^-7/6/5
significant SNPs: 5*10^-8
What are polygenic risk scores?
A score that predicts an individual’s risk of disease, based on the combination of their genotypes and effect size estimates from GWAS results
What are the limitations of NGS?
- Short reads of NGS make accurate characterisation of large variants hard –> knowledge on normal structural variants is limited
- NGS accuracy is currently lower than older, more expensive sequencing technology: variants are verified independently using Sanger sequencing (use of primers to target variant
What are the strategies to identify causal variants?
- Filter variants that are frequently observed: gnomAD (genome aggregation database)
- Look for variants that are identified as pathogenic
- Look for variants in genes linked to the condition
- Look for variants that effect functional elements (protein coding sequence, splicing, regulatory elements)
- Look for variants that are normally conserved
How are WGS used differently in clinical diagnostics vs research on rare diseases?
Rare disease diagnostics: sequence of affected individual + family members (affected or unaffected)
Rare disease discovery science: sequence of groups of affected individuals, looking to identify disease sharing variants
Which are the restrictions of WGS use in clinical care?
- Limit application to: monogenic diseases, patients with a clear phenotype, patients who are ill
- Reporting currently limited to: variants in protein coding sequences
- Cannot currently be usefully applied to: diagnosis of complex diseases, prediction of risk, patients with unexplained condition
What does mitochondrial DNA encode and what is its structure?
13 polypeptides, 22 tRNAs, 2 rRNAs, no introns, 93% coding DNA, no histones but association with several proteins to form nucleoids, use of polymerase gamma
What is heteroplasmy in mtDNA?
More than one type of MT genomes is present (or variation at position)
What are examples of mitochondrial diseases caused by nuclear genome mutations?
Follow mendelian inheritance patterns; Friedreich ataxia: inherited neurodegenerative disorder, progressive gait and limb ataxia, autosomal recessive involved in iron trafficking
What are examples of mitochondrial diseases caused by nuclear genome mutations with mitochondrial dysfunction dependence
Defects in genes that monitor and regulate mitochondria, requires mitochondrial dysfunction, important in ageing; Parkinson’s disease: rigidity, postural instability and tremor, progressive loss of dopaminergic neurons in brain; Parkin and PINK1 genes: damaged mitochondria –> PINK1 accumulation in outer membrane –> parkin recruitment –> autophagy –> build up of dysfunctional mitochondria
What are examples of mitochondrial diseases caused by mitochondrial genome mutations?
Point mutations or re-arrangements, inherited from mother, high percentage in tRNAs; LHON: most common, predominantly affects men
What is the threshold model?
Mutations causing disease can be heteroplasmic and have a threshold effect; medium level heteroplasmy (NARP), high level heteroplasmy (MILS (Leigh))
What is the connection between mitochondria and cancer?
Theory supported by idea that impairment of OXPHOS function stimulates AKT cell survival pathway –> downregulation of apoptosis
What are the 4Ps in medical genetics?
Positive effects:
- Participation
- Personalisation
- Prevention
- Prediction
What are some key topics related to justice in medical genetics?
- Risk: people in lower socio-economic classes have higher health risks (health gap)
- Discrimination: fear of genetic discrimination
- Individualisation of duties and responsibilities: concerns about costly predictive tests with no or little clinical utility
How is personalised medicine defined?
Considering individual characteristics at every stage, prevention, diagnosis, treatment and monitoring
What are the main ethical/social challenges of PM?
- Autonomy (right to know, right to not know)
- Rights and interests of family members
- Data protection
- Avoiding discrimination
- Effect on identities
Where does modification of core histones and DNA methylation take place?
Histone modification: H3, N-termini
DNA methylation: CG dinucleotides, addition of methyl group to 5th carbon of cytosine by methyltransferases (really favourable; to break down –> add hydroxyl group by TET proteins)
What are the roles of the different DNA methyltransferases?
Dnmt1: maintenance methylation of hemi-methylated strands
Dnmt2: tRNA methylase
Dnmt3a/b: de novo methylation (methylation of unmodified DNA during development)
Dnmt3L: oocyte and sperm specific roles, required to establish imprinting in oocytes
What are the two methylation-mediated transcriptional repression methods?
- Direct: methylation on promoter prevents binding of transcription factors
- Indirect: MeCP2 complex bind to methylated dinucleotide –> repressive chromatin
What is used to predict regulatory regions?
Histone code: post-translational modification of histone N-terminal tails
What is genomic imprinting?
Imposes biparental reproduction in mammals; an epigenetic phenomenon that causes genes to be expressed in a parent-of-origin-specific manner; around 100 genes in humans –> non-Mendelian inheritance
What are uniparental disomies?
Both chromosomes are derived from the same parent, or balanced translocation chromosome –> portion of chromosome is uniparental
What are Prader-Willi syndrome and Angelman’s syndrome?
Both 15q11 - q13
AS mutations in a single imprinted gene from mother (UBE3A): happy disposition, inappropriate laughter, widely spaced teeth, mental retardation
PWS deletions of multiple paternally expressed genes (or mother uniparental disomy of Ch 15): central obesity, short stature, small hands and feet, mild facial dysmorphism, always hungry
How are AS and PWS diagnosed?
Methylation-PCR method identifies patients with deletion, UPD or imprinting mutation at AS/PWS region
What is Beckwith-Wiedeman syndrome?
On 11p15; parent-of-origin associated prenatal overgrowth and cancer pre-disposition; large tongue (macroglassia), large organs (visceromegaly), large body size (macrosomia), omphalocele and microcephaly; most cases result from epimutations at either of the two 11p15.5 imprinting centres (IC1 and IC2); some germline mutations of NLRP2 gene
What is Silver-Russell syndrome?
Low birth weight, decreased birth length, triangular shaped face, postnatal growth retardation, poor appetite, fifth-finger clinodactyly ; 7% of cases have maternal UPD 7, paternal UPD 7 is normal –> SRS exhibits imprinting effects
What is Albright’s hereditary osteodystrophy?
Imprinted GNAS locus on Ch20; abnormal renal response to PTH; short metatarsals/carpals and phalanges and wide bones, thickening of calvaria
What is transient neonatal diabetes mellitus?
Epimutation in imprinted genes + genetic mutations in ZFP57 gene; heritable imprinting disorders compatible with life; diabetic, cardiac and & sometimes development delay; Zfp57 –> role in maintenance of DNA methylation in development
What are the effects of histone tail methylation?
Lysine 4 (K4): activation
K9: repression
K27: repression
K36: activation
What are the genes associated with cancer predisposition?
- Tumor suppressor genes: control rate of cell growth, typically diploid, i.e. APC in sporadic colon cancer and familial adenomatous polyposis
- Oncogenes: accelerate cell division, KRAS and BRAF
- DNA damage response/repair genes: constantly repairing DNA, cancer arises due to accumulation of mutations across genome, BRCA1 and 2
What are the DNA repair mechanisms and the diseases associated with them?
- Mismatch repair: lynch syndrome, colon, ovarian, endometrial cancer
- Double strand break repair: BRCA1 and 2
- Nucleotide excision repair: XP
What is Lynch syndrome?
Results from mismatch repair deficiency (MLH1, MSH2, MSH6, PMS2); colorectal, endometrial, ovarian, urothelial, brain, gastric, biliary tract, skin cancer
Why is low dose aspirin used for prevention of Lynch syndrome?
anti-platelet activity improves immune response to tumour cells; induces apoptosis of tumour cells
What is familial adenomatous polyposis (FAP)?
autosomal dominant; APC 100% penetrant; 100s or 1000s adenomatous polyps through out the colon
How do the cancer risks in BRCA1 and 2 differ?
BRCA1: high breast and ovarian cancer, 15% prostate cancer
BRCA2: high breast, lower ovarian, pancreatic, 25% prostate, high risk for men as well
What is Cowden syndrome?
Germline PTEN mutations, endometrial cancer, benign and malignant tumours of breast and thyroid, hamartomatous bowel polyps, skin and mucosal lesions, macrocephaly, developmental delay, targeted therapies: PARP inhibitors, mTOR inhibitors
What is Peutz-Jeghers syndrome?
Germline mutations in STK11; GI polyposis disorder, pigmented lesions on lips and buccal mucosa, increased risk of breast, GI and gynaecological tumours
What is Li-Fraumeni syndrome?
Germline TP53 mutations, sarcomas, breast cancer, adenocortical tumours, leukaemia, brain and spinal cord tumours, general increase in cancer risk
What are the two types of genetic tests?
Diagnostic: full screen of gene, normally undertaken in individual affected with the condition
Predictive: targeted test for specific mutation identified in another family member, normally undertaken in individual unaffected with condition
What are the techniques used to investigate chromosomes?
Traditional cytogenetics (cell cultures required): G banding, FISH, breakage Molecular cytogenetics (tests carried out on DNA): QF-PCR, MLPA, array CGH
What are the types of chromosomal abnormalities?
Chromosome rearrangements: recurrent miscarriage, infertility
Copy number imbalance: dysmorphism, developmental delay, learning difficulties, specific phenotypes (epilepsy, diabetes, cardiac manifestations)
Chromosome breakage syndromes: fanconi anemia, ataxia telangiectasia
Whole chromosome aneuploidy: non-disjunction at mitosis or meiosis
How can mosaicism arise in an initially normal conceptus?
non-disjunction or anaphase lag
What is CPM mosaicism?
mosaicism confined to extraembryonic tissue, may go undetected, may result in UPD following trisomy rescue
What are the types of chromosome rearrangements?
Robertsonian translocations, reciprocal translocations, inversions, intrachromosomal insertions
What are Robertsonian translocations?
Fusion of two acrocentric chromosomes (13, 14, 15, 21, 22); most common der(13;14) and der(14;21); balanced carriers phenotypically normal but have reproductive risks (recurrent miscarriages, Patau or Down (15% for female carriers) syndrome, male infertility); alternate segregation is fine, adjacent –> trisomies
What are reciprocal translocations?
Between any segments of any non-homologous chromosomes; balanced are phenotypically normal but with reproductive risk (infertility, miscarriage, child with congenital abnormalities); alternate segregation (normal), adjacent segregation (unbalanced), 3:1 segregation (unbalanced)
What are the two types of inversions?
Pericentric: includes centromere
Paracentric: does not include centromere, quite benign
What are examples of polygenic autoinflammatory diseases, mixed pattern diseases and classic polygenic autoimmune diseases?
Polygenic autoinflammatory diseases: Crohn’s disease and ulcerative colitis
Mixed pattern disease: psoriasis/psoriatic arthritis
Classic autoimmune diseases: systemic lupus erythematosus
What are the triggers for psoriasis?
Infection (streptococcal, viral), skin trauma, psychological stress, drugs (beta blockers, lithium), sunburn, metabolic factors (calcium deficiency), hormonal factors (pregnancy), unknown skin antigens stimulate immune response –> impaired differentiation and hyperproliferation of keratinocytes
What is inflammatory bowel disease (IBD)?
Ulcerative colitis: continuous inflammation/ulcers only in the mucosa of the colon, extra-intestinal manifestation, risk of cancer
Crohn’s disease: patchy inflammation of all layers of the GI tract (Ilietis, ileocolitis, colitis), mouth to anus involvement, fistulas and strictures, risk of cancer, extra-intestinal manifestations
What are the four etiologic hypotheses for IBD?
Persistent infection, dysbiosis, defective mucosal integrity, dysregulated immune response (loss of tolerance, aggressive cellular activation, defective apoptosis)
What are the different types of CYPs?
CYPs 1-3: high degree of polymorphism, 22 different isoforms
CYPs 4-51: metabolism of mostly endogenous compounds, low degree of polymorphism
What is the relation between tacrolimus and and CYP3A5*3 variation?
CYP3A53 is an intronic noncoding SNP which creates a splice site resulting in a mRNA insertion and frame shift. Some normally spliced transcripts still formed, more common in caucasians –> reduced expression of CYP3A in liver and jejunum –> transplant patients require lower doses of tacrolimus compared to CYP3A1 allele
How does warfarin dosage vary in different genotypes?
Vitamin K epoxide reductase subunit 1 (VKORC1) polymorphisms –> explain 30% of dose variation
CYP2C9 poor metabolism –> explains 10% of dose variation
What is the importance of pharmacogenetics for allopurinol?
Active metabolite: oxypurinol –> potent xanthine oxidase inhibitor preventing the formation of uric acid, used to treat gout
HLA-B*5801: associated with toxic epidermal necrolysis in Han Chinese populations
What is azathioprine and what is the importance of pharmacogenetics for this drug?
Standard 1st line immunosuppressant in IBD, used for remission, induction and maintenance, variable response 40-65%
TPMT deficiency –> increased drug-activation and overdosing from thioguanine nucleotides
What is the importance of pharmacogenetics in fluoropyrimidines?
dihydropyrimidine dehydrogenase deficiency –> toxicity in cancer patients treate with 5 fluorouracil
What are the components of a gene therapy protocol?
- Target tissue: accessible and manipulable
- Efficiency gene delivery: viral vs non-viral vectors
- Transcriptional control: sufficiently high, sustained
What is an ideal cell target in gene therapy?
Pluripotent, self-regenerating stem cells from the patient being treated
What are the four different viral gene therapy vectors?
Adenovirus: non-integrating
Adeno-associated virus: non-integrating
Retrovirus: integrating
Lentivirus: integrating
Which are the most important human gene therapy trial successes via ex vivo procedure?
Severe combined immune deficiency (SCID) X-linked chronic granulomatous disease Junctional epidermolysis bullosa X-linked adrenoleukodystrophy Metachromatic leukodystrophy beta-thalassemia/ SCD
Which are the most important human gene therapy trial successes via in vivo procedure?
Haemophilia B
Parkinson’s disease
Leber’s congenital amaurosis
