Unit 3 - Human Genome Variation

constitutional mutations

present in every cell body, example is cystic fibrosis, its a loss of function mutation where you need 2 defective copies to develop the disease, its constitutional cause each cell has to have the defective CFTR allele

genetic mosaicism

results from mutations occurring during development, genetic change causes some cells to be mutated while others stay normal creating a mosaic, example is retinoblastoma (RB1 is an important tumor suppressor gene, when its mutated and you get loss of function it causes retinoblastoma in early childhood and more cancers early in life, has to be in eye tissue to get the disease)

germline vs somatic mutations

variants in the germline are heritable while somatic ones are not, so will adults with the RB1 mutation result in their child getting the mutation, if its in the person’s germline so the gametes it gets passed down so the kid will get the mutation, this is important for family planning

single nucleotide variants (SNVs)

they are the most common mutation mechanisms, also often result in silent changes/polymorphisms (they have no clinical consequence so not the same as a mutation), different types includes substitutions (one base pair for another), deletion (loss of one or a small number of nucleotides) or insertions (gain of one or a small number of nucleotides)’ this results in either change of AA (missense mutation affecting protein function), introduction of a stop codon (nonsense mutation), alteration of a splice site or alteration of a splice enhancer, true mutations are those affecting translation or processing of RNA or function or protein

deletions and insertions

alters all codons following the change, for deletion everything shifts over which may change amino acid, could silence it or have some other effect, for insertion this shifts the reading frame which could introduce nonsense codon or make new AA or no change at all, if there is no change its a silent mutation

missense mutations

depends on impact of the amino acid change, can predict likelihood that change will be deleterious, if it alters amino acid size or charge it might affect protein folding or stability of certain protein domains, if it occurs in a highly conserved AA it likely has a more important function and if it occurs in a functionally important part of the protein it has the most potential for affecting the protein, different proteins have different tolerance for AA substitutions (ex: all missense changes in ACTA1 being a protein that polymerizes to form actin filament in muscles are pathogenic)

dominant negative missense mutation example

hypertrophic cardiomyopathy, a healthy 30 yr old man presents with chest pain and dyspnea (difficulty breathing), physical exam showed double apical sign and laterally displaced split second heart sound indicating its a cardiac issue, family history notable for brother who died of sudden death during football practice and father who died of congestive heart failure, gene panel revealed the disease is due to Arg403Gln mutation in MYH7 gene (interferes with binding globular head domain), dominant negative cause it interferes with dimerization of MYH7 protein so affects mutated copy and normal one, the disease results in left ventricle and atrium enlargement so ventricle wall expansion

myosin head group

if one copy abnormal, affects function of both

disease heterogeneity

47 yr old female presents with slow progressive distal weakness (muscles of hand + feet are affected) so she was clumsy as a child and now needs a cane to walk, first physical exam feature was a dropped toe (trying to lift your toe and you cant), also with marked hand weakness, muscle biopsy non specific myopathic muscle dysfunction, gene panel revealed p.Ala1636Pro mutation in MYH7, she cant really spread out or lift her fingers

MYH7

mutation location determines if it causes cardiac myopathy or skeletal myopathy (example of variable expressivity of disease)

recessive missense mutations

this means both alleles have to be mutated to cause disease, 2yr old boy with failure to grow properly and frequent infections, also had chronic diarrhea + colic, physical exam only remarkable for weight and height in the 2nd percentile, family history was unremarkable, Cl- sweat test was positive indicating clinical diagnosis of CF was made, CFTR genetic testing revealed homozygous G551D mutation (not considered dominant negative cause need other mutated allele to cause disease), mutation impairs CFTR, CF testing now part of newborn screening, mutation specific therapy available

nonsense mutations

SNV causing introduction of a stop codon, most commonly stop codons initiate nonsense mediated decay + RNA degradation resulting in loss of protein expression, if mutation is in the last 2 exons then instead a truncated protein results (invades the nonsense mediated decay machinery)

mutation breakdown

stop codon in the untranslated region would have no effect on gene processing, if a ribosome in the nonsense mediated decay zone encounters a STOP codon, RNA gets degraded producing very little protein, but in the nmd freezone ribosome leaps off only to make protein at the point of STOP codon so protein is stable just shorter

example of a nonsense mutation resulting in loss of protein

newborn female infant with respiratory distress, low muscle tone, inability to move arms or legs against gravity, also with joint contractures (restrictions in the movement of a joint to passive motion), no improvements of symptoms in first month of life, muscle biopsy shows nemaline myopathy (known genetic congenital muscle disease), whole exome sequencing reveals homozygous mutations in LMOD3 (substitution of G for A in c.1231 changing tryptophan to missense mutation), mutation in the same gene can result in different disease presentation, presence of rods are characteristic of nemaline myopathy, almost all patients have biallelic missense mutations resulting in very little protein production

LMOD3 WT vs LMOD3 mutation

if you have 2 nonsense mutations you make no normal LMOD3, affects protein coding sequence but not expression cause its nonsense

splice site mutations

interferes with processing of pre-mRNA, can result in exon removal (lack of inclusion of an exon, can happen when splice site mutation affects processing so RNA processing machinery skips a specific exon, this can produce either dominant negative effects if inframe or frameshift and stop codon (if reading frame is preserved it can have a truncated protein with dominant negative effect or remove exon altering reading frame), can also result in addition of intronic sequence but rare (typically introduces a stop codon (stop/gain), example is skipping exon 2 makes you retain intron 1 cause find novel splice site somewhere and use that to introduce intronic sequence which usually adds in a stop codon

ullrich congenital muscular dystrophy (UCMD)

characterized by muscle weakness and joint contractness, makes you hyperflexible

bethlem myopathy (BM)

presents with elbow contractions but more mild than congenital muscular dystrophy

UCMD5 18 aa deletion

loss of exons causing a dominant negative effect, when parts of individual polymer missing not only effects that one but also the following ones too so you cant form tetramers efficiently and mostly retained within the cell, this leads to no collagen VI matrix being formed

genomic organization of collagen IV

UCMD gets full in frame deletions that cause disease, and BM you get milder disease and see loss of function but need 2 affected alleles to get disease

duplications/insertions/deletions

can range in size greatly (example of a very small deletion is the 3 base pair in frame deletion F508del in CFTR, most common recessive mutation), this can cause in frame insertion/deletion so when insertion/deletion is in a multiple of 3 bases, the reading frame is not altered, frameshift with stop codon, gene duplication or deletion, deletion results in no gene product so you delete the whole gene (both alleles or deleted or haplosufficient so still get function with one allele left), duplication increases the amount of gene production and amplfication increases it even more so you can get deleterious effrects or cause improvement in disease phenotype

PMP22 neuropathy example

patients with this either present with charcot marie tooth disease type IA which is most common, presents in early childhood, causes slowly progressive distal to proximal weakness and sensory abnormalities (first sign is usually frequent trips and falls, it rarely progresses to wheelchair dependence), dominant inheritance, its the complete duplication of the PMP22 gene (3 copies), it can also be caused by point mutations, another example is hereditary neuropathy with liability to pressure palsies (HNPP), where single limb can become weak or numb for days at a time, episodes or weakness and numbness, often restricted to one limb, example would be inability to use arm for 2-3 days after sleeping on it, due to deletion of the PMP22 gene (single allele deletion)

X + Y chromosomes and gene dosage

the dosage is the amount of gene copies in genome + amount of it created, PAR1 + PAR2 is where pairing of X and Y happens, 1 chromosome has 824 coding and 769 non coding which is important for organismal function, the y chromosome is important for gonad and genital development, the X chromosome is much larger than the Y one

X inactivation in maintaining gene dosage

this is when one of the X chromosome is turned off in each cell by random process (creation of Barr body persisting through all somatic cell division), this occurs in the late blastocyst so either the maternal or paternal X is randomly inactivated resulting in mosaicism, if its the paternal once all descendant cells have paternal X inactivation and opposite if its maternal, with each mitotic division inactivation of X is transmitted, XIST encodes long non coding RNA that coats inactive X (has methylation of heterochromatin), for turner syndrome causes mild phenotypes but this would not show if females needed only one X, 50% of genes can escape X inactivation cause we have counter parts of Y chromosome so no need for gene dosage adjustment

monosomies

only viable one is turner syndrome

unbalanced and balanced rearrangements

if there is no imbalance in genomic content it has no deleterious effect on individual, balanced ones may put affected persons offspring at risk

unbalanced rearrangement

terminal deletion, interstitial deletion, duplication (could create gene dosage imbalances), ring and isochromosome

balanced rearrangement

robertsonian and reciprocal translocations

large scale deletions

caused by chromosomal breaking, the terminal deletion will occur at the end of a chromosome while the interstitial one happens within a chromosome arm

Cri du chat syndrome

caused by deletion of part of the short arm of chromosome 5, symptoms include reduced head size, distinctive facial features and intellectual impairment, its haploinsufficient so remaining allele cant compensate for mutated copy

tandem repeats

can lead to unequal crossover and unequal sister chromatic exchange, basically there’s a misalignment that happens during crossover but this only causes disease phenotypes when gene function is affected

ring chromosomal abnormality

this is when 2 breaks occur and the ends fuse together creating a ring, difficulty in anaphase results in more breakage + increased ring formation

isochromosome

1 arm missing causes the other arm to duplicate to compensate, this means you get 3 copies of duplicated arm and single copy from missing arm (frequently seen in cancer)

robertsonian translocation

karyotype only has 45 chromosomes, its a balanced rearrangement because short arms of an acrocentric chromosome consist of non coding DNA so no gene dosage imbalance so phenotypically normal but it might affect offspring, it involves the fusion of 2 acrocentric chromosomes (chromosomes where centromeres are located at the very end of one chromosome), not reciprocal translocation as short one is lost

reciprocal translocation

no genetic material lost or change so not typically associated with genetic abnormality, generates unbalanced gametes so offspring might develop phenotype

philadelphia chromsome

its derived from translocation between chromosome 9 and 22, increased activity leads to CML, basically you have normal chromosome 9 with abl gene and chromosome 22 with bcr , they break and those portions fuse together creating the philadelphia chromosome

autosome abnormalities

no monosomies are viable, only trisomy of 3 chromosomes are viable (chromosomes 13, 18 or 21 down syndrome

sex chromosome abnormalities

affects 1 out of 400 live births, less severe than autosomal aneuploidies because gene dosage imbalance is minimized by Y chromosome being relatively gene poor + X inactivation, of these XXY, XXX and XYY are most common, turner syndrome (45, X) results in puffy feet and webbed neck

nondisjunction

failure of chromosomes to separate from each other during either round of meiosis, 1 sister chromatid = chromosome

non disjunction (meiosis 1)

the first division homologous chromosomes fail to separate so at the second one you get two cells with 2 copies of chromosomes and 2 with nothing resulting in ½ trisomy and ½ monosomy

non disjunction (meiosis 2)

you get normal 1st division so 2 pairs of sister chromatids, one per division, but at second division sister chromatids in 1 cell fail to separate so one has 2 copies, 1 with none and 2 normal haploid cells, these means you get 2 normal diploid cells, 1 trisomic and 1 monosomic