genetics. mutations point mutation –missense mutation –nonsense mutation frameshift mutation ...

GeneticsGenetics

MutationsMutations

Point mutationPoint mutation– Missense mutationMissense mutation– Nonsense mutationNonsense mutation

Frameshift mutationFrameshift mutation Trinucleotide repeat mutationTrinucleotide repeat mutation

MutationsMutations::

Decreased gene product or inactive protein:Decreased gene product or inactive protein: Enzymes:AREnzymes:AR Regulation of complex metabolic pathways: e.g., LDL receptorRegulation of complex metabolic pathways: e.g., LDL receptor Key structural proteins: dominant negativeKey structural proteins: dominant negative

Gain of function:Gain of function: Almost always AD, e.g., Huntington diseaseAlmost always AD, e.g., Huntington disease

Mendelian DisordersMendelian Disorders

Expressed mutations Expressed mutations in single genes of in single genes of large effectlarge effect

Gene expressionGene expression– DominantDominant– RecessiveRecessive– CodominantCodominant

Pleiotropism vs genetic Pleiotropism vs genetic heterogeneityheterogeneity

Autosomal Dominant DisordersAutosomal Dominant Disorders

Onset: older ageOnset: older age Reduced penetranceReduced penetrance Variable expressivityVariable expressivity New mutation:New mutation:

– Frequency depends on reproductive capabilityFrequency depends on reproductive capability– In egg or spermIn egg or sperm– Germ cells of older fathersGerm cells of older fathers– No increased risk in siblingsNo increased risk in siblings

Examples of AD inheritanceExamples of AD inheritance

Huntington diseaseHuntington disease NeurofibromatosisNeurofibromatosis Tuberous sclerosisTuberous sclerosis Polycystic kidney diseasePolycystic kidney disease Familial polyposis coliFamilial polyposis coli Hereditary spherocytosisHereditary spherocytosis Marfan syndromeMarfan syndrome Familial hypercholestrolemiaFamilial hypercholestrolemia

Autosomal Recessive DisordersAutosomal Recessive Disorders

The largest group in Mendelian disordersThe largest group in Mendelian disorders Almost all of the inborn errors of metabolismAlmost all of the inborn errors of metabolism EnzymesEnzymes Complete penetranceComplete penetrance More uniform expressionMore uniform expression Early onsetEarly onset New mutations: ?New mutations: ?

Examples of AR inheritanceExamples of AR inheritance

Cystic fibrosisCystic fibrosis PKUPKU Lysosomal storage diseaseLysosomal storage disease Sickle cell anemiaSickle cell anemia Congenital adrenal hyperplasiaCongenital adrenal hyperplasia Ehler-Danlos syndromeEhler-Danlos syndrome Spinal muscular atrophySpinal muscular atrophy

X- Linked DisordersX- Linked Disorders

No Y- linked inheritanceNo Y- linked inheritance Almost all recessiveAlmost all recessive Males are hemizygote for X-linked mutant Males are hemizygote for X-linked mutant

genesgenes Random inactivation of one of the X- Random inactivation of one of the X-

chromosomes; chromosomes; partialpartial symptoms,e.g., symptoms,e.g., G6PDG6PD

Examples of XLR inheritanceExamples of XLR inheritance

Duchenne muscular dystrophyDuchenne muscular dystrophy Hemophilia A and BHemophilia A and B G6PD deficiencyG6PD deficiency Wiskott-Aldrich syndromeWiskott-Aldrich syndrome Diabetes insipidusDiabetes insipidus Fragile X syndromeFragile X syndrome

X- Linked DisordersX- Linked Disorders

Rare X-linked DominantRare X-linked Dominant– How is the inheritance?How is the inheritance?– Such as Vitamin D resistant ricketsSuch as Vitamin D resistant rickets

Mendelian DisordersMendelian DisordersBiochemical & Molecular BasisBiochemical & Molecular Basis

Enzyme defectsEnzyme defects Defects receptors & transport systemsDefects receptors & transport systems Alterations in structure, function or quantity Alterations in structure, function or quantity

of nonenzyme proteinsof nonenzyme proteins Genetically determined adverse reaction to Genetically determined adverse reaction to

drugsdrugs

Enzyme DefectsEnzyme Defects

Enzyme:Enzyme:– Quantity.Quantity.– Quality.Quality.

Decreased product.Decreased product.– Albinism.Albinism.

Increased substrate or Increased substrate or intermediates.intermediates.– PKU.PKU.

Ipmaired inactivation of Ipmaired inactivation of toxic substrate.toxic substrate.– Alpha1 Antitrypsin D.Alpha1 Antitrypsin D.

Genetically Determined Adverse Genetically Determined Adverse Reaction to DrugsReaction to Drugs

PharmacogeneticsPharmacogenetics– Enzyme deficiency unmasked by drug Enzyme deficiency unmasked by drug

administrationadministration

G6PD and Primaquine G6PD and Primaquine

Disorders Associated With Disorders Associated With Defects in Structural ProteinDefects in Structural Protein

Fibrillin: Marfan syndromeFibrillin: Marfan syndrome Collagen: Ehler Danlos syndromeCollagen: Ehler Danlos syndrome Dystrophin: Duchene/BeckerDystrophin: Duchene/Becker Spectrin/Ankyrin/Protein 4,1: SpherocytosisSpectrin/Ankyrin/Protein 4,1: Spherocytosis

Marfan SyndromeMarfan Syndrome

Definition: Definition: – Connective tissue (elastic fiber) disorderConnective tissue (elastic fiber) disorder– Major involved organsMajor involved organs

SkeletonSkeleton EyeEye Cardiovascular systemCardiovascular system

Prevalence: 1/10,000 – 1/20,000Prevalence: 1/10,000 – 1/20,000


Autosomal dominant inheritanceAutosomal dominant inheritance 70-80% familial vs 20-30% new mutations70-80% familial vs 20-30% new mutations Variable expression: genetically heterogeneous Variable expression: genetically heterogeneous MutationMutation

– Almost allAlmost all– Negative dominantNegative dominant– Chromosome 15q21.1Chromosome 15q21.1– FNB1 geneFNB1 gene

Elastic fibersElastic fibers

Central coreCentral core– Predominantly ellastinPredominantly ellastin

Peripheral Peripheral microfibrillary networkmicrofibrillary network– Predominantly Predominantly fibrillinfibrillin

FibrillinFibrillin

Particularly abundant inParticularly abundant in– AortaAorta– LigamentsLigaments– Ciliary zonules of lensCiliary zonules of lens


PathogenesisPathogenesis– Inherited defect in fibrillin, an extracellular Inherited defect in fibrillin, an extracellular

glycoproteinglycoprotein– FBN1 gene mutationFBN1 gene mutation

70 different mutation70 different mutation Mostly nonsense mutationsMostly nonsense mutations


Skeletal abnormalitiesSkeletal abnormalities– Most strikingMost striking– Usually tallUsually tall– Upper segment/lower segment: lowUpper segment/lower segment: low– Long extremitiesLong extremities– Pectus excavatumPectus excavatum– Long tapering fingers and toesLong tapering fingers and toes

Marfan SyndromeMarfan Syndrome…Skeletal Abnormalities…Skeletal Abnormalities

Hyperflexibility of jointsHyperflexibility of joints ScoliosisScoliosis KyphosisKyphosis Rotation or slipping of thoracic vertebraeRotation or slipping of thoracic vertebrae Dolichocephalic (long-headed)Dolichocephalic (long-headed)

– Cranial index less than 75%Cranial index less than 75%– Cranial endex: width of skull/length of skullCranial endex: width of skull/length of skull

Bossing of frontal & supraorbital ridgesBossing of frontal & supraorbital ridges

Marfan SyndromeMarfan SyndromeOcular ChangesOcular Changes

CharacteristicCharacteristic– Very rare in those without this diseaseVery rare in those without this disease

Bilateral subluxation or dislocation of lensBilateral subluxation or dislocation of lens– Ectopia lentisEctopia lentis

Marfan SyndromeMarfan SyndromeCardiovascular ChangesCardiovascular Changes

Aortic neurysmAortic neurysm– Cystic medionecrosisCystic medionecrosis

Intimal tearIntimal tear– DissectionDissection

Towards root of aorta or iliacTowards root of aorta or iliac Ruptured dissection: cause of 30-45% of deathsRuptured dissection: cause of 30-45% of deaths

– Aortic regurgitationAortic regurgitation

Marfan SyndromeMarfan Syndrome...Cardiovascular Changes...Cardiovascular Changes

Mitral prolapseMitral prolapse– Loss of connective tissue supportLoss of connective tissue support– More commonMore common– Less seriousLess serious– Floppy valveFloppy valve– Elongated chordae tendineaeElongated chordae tendineae– Similar changes in tricuspid and rarely aortaSimilar changes in tricuspid and rarely aorta


DiagnosisDiagnosis– Presymptomatic Dx:Presymptomatic Dx:

RFLPRFLP 70 different mutations70 different mutations

– Direct gene diagnosis impossibleDirect gene diagnosis impossible

Defects in Collagen Synthesis or Defects in Collagen Synthesis or StructureStructure

Osteogenesis imperfectaOsteogenesis imperfecta Alport syndromeAlport syndrome Epidermolysis bullosaEpidermolysis bullosa Ehler Danlos syndrome (EDS)Ehler Danlos syndrome (EDS)

CollagenCollagen

Most abundant protein in animal worldMost abundant protein in animal world At least 14 distinct collagen typesAt least 14 distinct collagen types General formulaGeneral formula

– (gly-x-y)(gly-x-y)nn

– Triple helixTriple helix Three Three chains: about 30 chains: about 30 chains chains

Collagen SynthesisCollagen Synthesis

Ehler Danlos Syndrome (EDS)Ehler Danlos Syndrome (EDS)

Genetically heterogeneousGenetically heterogeneous At least 10 variantAt least 10 variant Clinical manifestationsClinical manifestations

– SkinSkin HyperextensibleHyperextensible Extremely fragileExtremely fragile

– JointsJoints Prone to dislocationProne to dislocation Hypermobile Hypermobile


Type VIType VI– Most commom AR form of EDSMost commom AR form of EDS– Mutation in lysyl hydroxylase geneMutation in lysyl hydroxylase gene

Only collagen I and IIIOnly collagen I and III

– Ocular fragility with rupture of cornea and retinal Ocular fragility with rupture of cornea and retinal detachmentdetachment


Type IVType IV– AD inheritanceAD inheritance– Collagen type IIICollagen type III– At least 3 different mutation:At least 3 different mutation:

Abnormal collagenAbnormal collagen Decreased synthesisDecreased synthesis Decreased excretionDecreased excretion

– Some negative dominantSome negative dominant– Rupture of colon and large arteriesRupture of colon and large arteries


Type VIIType VII– AD inheritanceAD inheritance– Abnormal procollagen type IAbnormal procollagen type I– Peptidase can not cleave the N terminalPeptidase can not cleave the N terminal– GenesGenes

1[I]1[I] 2[I]2[I]


Type IXType IX– XLR inheritanceXLR inheritance– Mutation in copper binding proteinMutation in copper binding protein

Decreased activity of lysyl hydroxylaseDecreased activity of lysyl hydroxylase– Cross-linking of collagen & elasticCross-linking of collagen & elastic

– High level of copper within the cellHigh level of copper within the cell– Low serum copper & ceruloplasmin levelsLow serum copper & ceruloplasmin levels

Familial HypercholestrolemiaFamilial Hypercholestrolemia

A receptor diseaseA receptor disease The most frequent mendelial disorderThe most frequent mendelial disorder

– 3-6% of survivors of MI3-6% of survivors of MI

Mutation in the gene encoding LDL receptorMutation in the gene encoding LDL receptor– HypercholestrolemiaHypercholestrolemia

Premature atherosclerosis: MIPremature atherosclerosis: MI XanthomaXanthoma


HeterozygotesHeterozygotes– 1/5001/500– 2-3 times higher plasma cholestrol2-3 times higher plasma cholestrol

HomozygotesHomozygotes– 5-6 times higher plasma cholestrol5-6 times higher plasma cholestrol– MI before 20 years of ageMI before 20 years of age


PathogenesisPathogenesis– Decreased LDL clearance (uptake)Decreased LDL clearance (uptake)– Increased LDL productionIncreased LDL production

More IDL coverts to LDLMore IDL coverts to LDL In both heterozygotes and homozygotesIn both heterozygotes and homozygotes

– Increased LDL uptake by Increased LDL uptake by macrophage/monocyte (scavenger receptor)macrophage/monocyte (scavenger receptor) Acetylated or oxidized LDLAcetylated or oxidized LDL


LDL receptor geneLDL receptor gene– Extremely largeExtremely large

18 exons18 exons 5 domains5 domains 45 kb45 kb


LDL receptor geneLDL receptor gene– More than 150 different mutationsMore than 150 different mutations

InsertionInsertion DeletionDeletion MissenseMissense NonsenseNonsense

– Mutations categorized in 5 groupsMutations categorized in 5 groups

ManagementManagement

StatinsStatins– HMG-CoA reductase inhibitionHMG-CoA reductase inhibition

Decreased synthesis of cholestrolDecreased synthesis of cholestrol Increased synthesis of LDL receptorIncreased synthesis of LDL receptor

Gene therapyGene therapy

Lysosomal Storage DiseasesLysosomal Storage Diseases

DefinitionDefinition– Lack of any protein essential for the normal Lack of any protein essential for the normal

function of lysosomesfunction of lysosomes


Involved organs depend onInvolved organs depend on– The site where most of the material to be The site where most of the material to be

degraded is found.degraded is found. GM1 & GM2 gangliosidosesGM1 & GM2 gangliosidoses

– BrainBrain MucopolysaccharidosesMucopolysaccharidoses

– All of the bodyAll of the body

– The location where most of the degradation The location where most of the degradation normally occursnormally occurs Mononuclear phagocytesMononuclear phagocytes

Tay-Sachs DiseaseTay-Sachs Disease

Most common form of GM2 gangliosidosesMost common form of GM2 gangliosidoses Ashkenazi jewsAshkenazi jews

– 1/30 carrier rate1/30 carrier rate

All tissues lack hexosaminidase AAll tissues lack hexosaminidase A– Including leukocytes and plasmaIncluding leukocytes and plasma

GM2 accumulation in many organsGM2 accumulation in many organs– Heart, liver, spleen,Heart, liver, spleen,CNS, autonomous nervous CNS, autonomous nervous

system, retinasystem, retina, .., ..

Niemann-Pick DiseaseNiemann-Pick Disease

Rare lysosomal storage diseaseRare lysosomal storage disease Lysosomal accumulation of sphingomyelinLysosomal accumulation of sphingomyelin

– Sphingomyelinase deficiencySphingomyelinase deficiency

Common in Ashkenazi jewsCommon in Ashkenazi jews Types A & BTypes A & B Previously type CPreviously type C

– Defect in intracellular cholestrol esterification & transportDefect in intracellular cholestrol esterification & transport

SphingomyelinSphingomyelin

Present in cell membranes


Type AType A– Severe infantile typeSevere infantile type

Extensive neurologic involvementExtensive neurologic involvement Severe visceral accumulation of sphingomyelinSevere visceral accumulation of sphingomyelin

– 75-80% of cases75-80% of cases– Survival: less than 3 yearsSurvival: less than 3 years


……Type AType A Missense mutationMissense mutation Complete deficiency of sphingomyelinaseComplete deficiency of sphingomyelinase


Type BType B– OrganomegalyOrganomegaly– No CNS involvementNo CNS involvement– Survive adulthoodSurvive adulthood

Niemann-Pick DiseaseNiemann-Pick DiseaseDiagnosisDiagnosis

Biochemical studiesBiochemical studies– Sphingomyelinase activity in leukocytes and Sphingomyelinase activity in leukocytes and

cultured fibroblastscultured fibroblasts

DNA probes:DNA probes:– Both patients and carriersBoth patients and carriers

Gaucher DiseaseGaucher Disease

Glucocerebrosidase gene mutationGlucocerebrosidase gene mutation Accumulation of glucocerebroside in Accumulation of glucocerebroside in

phagocytes and sometimes CNSphagocytes and sometimes CNS


Most common lysosomal storage diseaseMost common lysosomal storage disease TypesTypes

– I (chronic non-neuropathic): 99%I (chronic non-neuropathic): 99% Decreased enzyme activityDecreased enzyme activity Without CNS involvementWithout CNS involvement Predominantly spleen & skeletonPredominantly spleen & skeleton Pancytopenia or thrombocytopenia Pancytopenia or thrombocytopenia Pathologic Fx and bone painPathologic Fx and bone pain Progressive but compatible with long lifeProgressive but compatible with long life European JewsEuropean Jews


……typestypes– II (acute neuropathic)II (acute neuropathic)

No enzyme activityNo enzyme activity No predilection for jewsNo predilection for jews InfantileInfantile Progressive involvement of CNS & early deathProgressive involvement of CNS & early death HepatosplenomegalyHepatosplenomegaly


DiagnosisDiagnosis– HomozygotesHomozygotes

Enzyme activityEnzyme activity– Peripheral blood leukocytesPeripheral blood leukocytes– Cultured skin fibroblastsCultured skin fibroblasts

– HeterozygotesHeterozygotes Enzymatic methods not reliableEnzymatic methods not reliable Detection of mutationDetection of mutation

– More than 30 different mutationsMore than 30 different mutations


ManagementManagement– DifficultDifficult– Replacement therapyReplacement therapy

Recombinant enzyme: extremely expensiveRecombinant enzyme: extremely expensive Bone marrow transplantationBone marrow transplantation Gene therapy: futureGene therapy: future

Glycogen Storage DiseasesGlycogen Storage Diseases

AKA: GlycogenosesAKA: Glycogenoses Genetic disease with metabolic defect in Genetic disease with metabolic defect in

synthesis or catabolism of glycogensynthesis or catabolism of glycogen


Hepatic typeHepatic type– HepatomegalyHepatomegaly– HypoglycemiaHypoglycemia– ExamplesExamples

Von Gierke: Glucose-6-phosphatase (I)Von Gierke: Glucose-6-phosphatase (I) Liver phosphorylase (VI)Liver phosphorylase (VI) Debranching enzyme(III)Debranching enzyme(III)


Myopathic typeMyopathic type– Muscle weaknessMuscle weakness– Cramps following exerciseCramps following exercise– Following exercise lactate does not increaseFollowing exercise lactate does not increase– ExamplesExamples

McArdle: muscle phosphorylase(V)McArdle: muscle phosphorylase(V) Muscle phosphofructokinase (VII)Muscle phosphofructokinase (VII)


MiscellaneousMiscellaneous– Pompe (acid maltase, Pompe (acid maltase, -glucosidase)-glucosidase)

Lysosomal accumulation of glycogenLysosomal accumulation of glycogen Predominantly heart involvementPredominantly heart involvement Early deathEarly death

Pompe DiseasePompe Disease

Disorders With multifactorial Disorders With multifactorial InheritanceInheritance

Some normal phenotypesSome normal phenotypes– HeightHeight– IntelligenceIntelligence– Eye & hair colorEye & hair color

Normal DistributionNormal DistributionNormal DistributionNormal Distribution

90% Samples90% Samples



+1.65+1.65x x +2.58+2.58xx

xx__

XX

+1.96+1.96xx

-2.58-2.58xx -1.65-1.65xx

-1.96-1.96xx


Different diseasesDifferent diseases– Cleft lip & palateCleft lip & palate– Congenital heart diseaseCongenital heart disease– Coronary heart diseaseCoronary heart disease– HTNHTN– GoutGout– DMDM– Pyloric stenosisPyloric stenosis


Both environment and two or more mutant Both environment and two or more mutant genes (dosage effect)genes (dosage effect)

Not polygenic inheritanceNot polygenic inheritance Variable expressibilityVariable expressibility Reduced penetranceReduced penetrance First rule out mendelian & chromosomal First rule out mendelian & chromosomal

inheritanceinheritance


Risk of expression: # of mutant genes inheritedRisk of expression: # of mutant genes inherited– Severity of diseaseSeverity of disease– # of diseased individuals# of diseased individuals

The rate of recurrence of the disorder (in range of The rate of recurrence of the disorder (in range of 2-7%) is the same for all first-degree relatives of 2-7%) is the same for all first-degree relatives of affected individualsaffected individuals

Identical twins: concordance 20-40%Identical twins: concordance 20-40% Expression of multifactorial traitExpression of multifactorial trait

– Continuous: heightContinuous: height– Discontinuous: DMDiscontinuous: DM

genetics. mutations point mutation –missense mutation –nonsense mutation frameshift mutation ...

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