Differences from mendelian heredity Imprinting, dynamic mutations RNDr Z.Polívková Lecture No 438 - course: Heredity

Genomic imprinting Mendelian principle: autosomal genes have the same expression if transmitted from father or mother Imprinted genes: maternal and paternal alleles have different expression (activity) according to parental origin Gene imprinting = epigenetic form of regulation of gene expression

Imprinted genes: Monoallelic expression – expression of only one parental allele (parent-of-origin expression) Imprinted genes - functionally haploid Active allele – transcribed Inactive allele – imprinted – nontranscribed – silent Imprinting connected with DNA methylation and other changes of chromatin

Imprinted genes normally involved in embryonic growth, cell division Abnormality of imprinting = human pathologies

Gynogenesis and androgenesis: Examples of human pathologies : Human triploidies: Additional paternal set of chromosomes = partial mole = hyperplasia of trophoblast Additional maternal set of chromosomes = hypotrophic placenta Gynogenesis and androgenesis: Ovarial teratoma – division of ovum without fertilization Complete mole – division of only male pronucleus (without maternal contribution)

∑ : Role of the imprinted genes in early human embryogenesis: paternally expressed genes → placental proliferation and invasivness maternally expressed genes → development of embryo

Prader- Willi sy (PWS) MR, short stature ,obesity, hypotonia, characteristic facies, small feet and hands, hypogonadism Angelman sy (AS) MR, absence of speech, seizures, jerky gait, inappropriate laughter, dysmorphic features

PWS AS

UPD = both chromosomes 15 from one parent PWS AS deletion 15q11-13 on paternal chromosome on maternal chromosome UPD (uniparental disomy) maternal paternal mutation maternal active allele imprinting error maternal imprint paternal imprint On both chromosomes on both chromosomes in PWS region in AS region UPD = both chromosomes 15 from one parent

Deletion 15q11-13 Wysis

PWS AS PWS region – active paternal elleles in proximal region of chromosome 15 – two groups of reciprocally imprinted genes PWS region – active paternal elleles AS region - active maternal allele(s) loss of function of active alleles in PWS region (pat) loss of function of active allele(s) in AS region (mat) → functional nullisomy

Imprinted genes on chromosome No 15 normal situation pat mat } SNRPN PWS genes ZNF127 } AS gene UBE3A active paternal ellele active maternal allele „silent“= imprinted paternal allele „silent“=imprinted maternal allele

Deletion in PWS and AS pat mat pat mat Deletion of paternal active alleles in PWS Deletion of maternal active alleles in AS PWS AS

UPD - Uniparental disomy in PWS and AS mat mat pat pat Uniparental disomy maternal in PWS paternal in AS PWS AS UPD

Mutation in AS mutated active maternal allele in AS AS

Imprinting error pat mat pat mat maternal imprint of PWS genes on both chromosomes in PWS paternal imprint of AS gene on both chromosomes in AS PWS AS

Imprinting and Beckwith-Wiedeman syndrome (EMG – exomphalos - macroglossia – gigantism) growth abnormalities : macroglossia, gigantism, hemihypertrophy, visceromegaly, abdominal wall defects (omphalocele, umbilical hernia); hypoglycemia in neonatal period, renal dysplasia, skeletal anomalies predisposition to embryonal tumors (Wilms´ tumor,… ) Imprinted genes on 11p15: IGF2 – expressed from paternal allele H19 – expressed from maternal allele p57 - expressed from maternal allele ?

Changes in BWS: paternal duplication of 11p (2 x IGF2) paternal UPD (2 x IGF2) deletion or translocation of maternal active allele H19→ activation of maternal IGF2 allele („enhancer“ competition model for expression control of IGF2 and H19) abnormal imprint = biallelic expression of IGF2 Pathogenesis of disease – increased dose of IGF2 (growth factor) role of H19 and other genes = ?

pat mat pat mat 1. paternal duplication IGF2 IGF2 IGF2 H19 H19 H19 IGF2 H19 normal situation BWS region on 11p15 1. paternal duplication active and silent paternal alleles active and silent maternal alleles

biallelic expression of IGF2 pat pat pat mat IGF2 IGF2 H19 3. del,, transl. of maternal allele H19, expression of paternal IGF2 2. paternal UPD pat mat IGF2 H19 biallelic expression of IGF2 4. imprinting error

Imprinting and cancer Genetic changes in cancer: inherited or spontaneous mutations that are not corrected by repair mechanisms – irreversible changes in protooncogenes, tumour suppressor genes Epigenetic changes: (do not affect the primary sequences of genome) – changes in methylation (imprinting) of these genes → activation of protooncogenes, silencing of tumour suppressor genes by aberrant methylation

But! in some persons, in some tissues – WT1 is imprinted Wilms´ tumor (WT) Locus 11p13 – connected with WAGR syndrome (Wilms´tumor, aniridia, urogenital anomaly, mental retardation) WT1 gene- transcription factor (tumour suppressor) – biallelic expression in kidney and other organs But! in some persons, in some tissues – WT1 is imprinted = polymorphism of imprinting = predisposition to cancer

Locus 11p15 – connected with BWS IGF2 – growth factor – monallelic expression in kidney biallelic expression in Wilms tumor and other tumors (LOI = loss of imprinting) oncogen H19 – oncofetal RNA – expressed in embryogenesis- function = ?, loss of H19 expression in WT with biallelic IGF2 expression H19 - expressed in some tumors (lung, oesophageal, bladder carcinoma) p57 – inhibitor of cyclin dependent kinase reduced expression in WT and other tumors (lungs)

Imprinting connected with methylation Imprinted allele = methylated Imprinted protooncogenes – loss of imprinting (LOI) = activation of imprinted allele = biallelic expression = oncogenes Imprinted tumor supressor genes = predisposition to tumors – loss of only one allele = loss of gene function

Knudson two-hit hypothesis of inactivation of tumor suppressor genes : 1st step: germ mutation or somatic mutation, or imprinting of one allele 2nd step: loss of heterozygosity (LOH) –by mutation in somatic cell

Polymorphism of imprinting of some genes in population: tumour supressor genes WT1 (11p13), IGF2R (=IGF2 receptor on 6q26 - inactive in different tumours, role of IGF2R in extracellular IGF2 degradation) In population – biallelic expression of these genes in some people monoallelic expression (imprinted) = predisposition to cancers methylation = reversible process – possibility of therapy of tumours caused by aberrant methylation ???

Imprinting stage-, tissue-, species- and strain-specific polymorphism of imprinting (inter-individual differences) imprinted genes – function in positive or negative regulation of embryonal growth, cell division and differentiation imprinted genes: receptors, growth factors, regulation proteins, transcription factors, proteins in splicing = protooncogenes, tumour suppressor genes role in embryonal development biallelic expression in some stages of ontogenesis ? imprinting is reversible

Imprinting connected with methylation, histone deacetylation and with remodelation of chromatine structure to inactive state

Abnormality of imprinting = growth abnormalities, abnormality of development, behaviour and cancers Lack of expression (because of mutation, deletion, deficiency) of a gene usually expressed monoallelically from a specific parent or overexpression (because of duplication, relaxation of imprinting or loss of control) of a normally monoallelically expressed gene

Uniparentalní disomy=inheritace of both homologs from the same parent Mechanisms of UPD origin: Loss of one chromosome from trisomic zygote =„correction“ of initial trisomy (trisomy rescue) gametic complementation = fertilization between nullisomic and disomic gametes (for the same chromosome) duplication of the single chromosome in monosomic zygote postfertilisation error – nondisjunction and reduplication of the single chromosome or mitotic recombination

Origin of uniparental disomy from trisomic zygote trizomic zygote loss of chromosome loss of chromosome uniparental disomy normal

Origin of uniparental disomy by gamete complemetation Fertilization of disomic and nullisomic gametes

duplication of chromosome Origin of uniparental disomy by duplication of chromosome of monosomic zygote monozomic zygote duplication of chromosome

Origin of partial isodisomy by postfertilisation error normal zygote-dizomic nondisjunction and duplication mitotic recombination

Evidence for UPD: trisomy 15 in CVS, normal karyotype in fetal blood  child with PWS increased parental age in UPD transmission of hemphilia from father to son (zygote XXY and loss of maternal X chromosome transmission od balanced translocation 22/22 in balanced form to healthy child (trisomic zygote and loss of single chromosome 22) pericentric inversion was present on one homologue in mother and on both homologues in one offspring maternal UPD in PWS, paternal UPD in AS maternal UPD of chromosome No 7 in a patient with cystic fibrosis and growth retardation (first detection of UPD)

UPD  abnormal development if imprinted genes are present

Dynamic mutations

mothers od affected males = carriers Fragile X syndrome = X-linked mental retardation - 1:1500 of males cytogenetic manifestation – fragile site Xq27.3 = FRAXA Clinical signs: mental retardation, macroorchidism (large testicles), long face, large mandible, large everted ears mothers od affected males = carriers but: 30% of women = carriers - mentally retarded 20% fraX men mentally normal deterioration of manifestation through generation = (Sherman paradox)

Unstable triplet repeats (CCG)n in FMR1 gene in normal population 6-50 copies premutation (without MR) 50-200 copies full mutation (with MR) 200-2000 copies DNA methylation (promoter region)  FMR1 is not transcribedabsence of proteinMR

Premutation=unstable premutationfull mutation = only through mother carrier (in oogenesis or early in embryonal life) man with premutation length of element is not increased in the next generation length of amplification in correlation with cytogenetic expression gene function ?? – protein expressed in tissues, higer levels in brain and testes

gradual origin of mutation = dynamic mutation Dynamic mutations = initial change of DNA produce another change = expansion of triplet repeats

Main features od dynamic mutations: homogenity – no more alleles somatic variability- different numbers of copies in different tissues effect of parental origin on manifestation difference from mendelian principles (low penetrance) no new mutations – gradual arise through premutation, familiar expresivity depends on number of copies anticipation=deterioration of clinical signs through generations

Two groups of dynamic mutations: amplification in noncoding (nontranslated) region of gene (promoters, introns) loss of function fra X(CCG/GGC), myotonic dystrophy (CTG), Friedreich ataxia (GAA) amplification in exons (usually CAG repeats)  genes are transcribed abnormal protein Huntington disease-HD- (abnormal protein huntingtin inactivates associated proteins), spinocerebellar ataxia type 1

Expansions depends on the sex of transmitting parent Fra X, myotonic dystrophy – expansion if disease is inherited from mother HD - expansion- if inherited from father Postzygotic origin of amplification on chromosome of specific parental origin - determined in gametogenesis

Dynamic mutation Fragile X Myotonic dystrophy Huntington disease FRAXA DM HD heredity XD with reduced AD with different age AD with different age penetrance of onset of onset parental origin maternal=full mutation maternal –congenital forms paternal – early onset mechanism abnormal DNA metylation different mRNA level abnormal protein transcription of FMR1 toxic for neurons? is stopped amplification (CCG)n (CTG)n (CAG)n normal number 10-50 5-35 9-34 abnormal number 50-200 premutation 50-80 30-1000 200-2000 full mutation 80-2000 lower number=reduced penetrance gene FMR1 IT15

http://dl1.cuni.cz/course/view.php?id=324 presentation http://dl1.cuni.cz/course/view.php?id=324 supplementary text to cytogenetics Thompson &Thompson: Genetics in medicine, 7th ed. Chapter 5: Principles of clinical cytogenetics: Parent-of-origin effects: Genomic imprinting Chapter 7: Patterns of single gene i heritance: Imprinting in pedigrees, Unstable repeat expansions Chapter 12: The molecular, biochemical and cellular basis of genetic disease: Diseases due to the expansion of unstable repeat sequences: Biochemical and cellular mechanisms