Medical genetics II Microdeletion syndromes Standard karyotype 46,XX or 46, XY n Types of chromosomal aberrations https://readbiology.com/wp-content/uploads/2020/04/aberration.jpg strukutrní aber Structural chromosomal abnormalitites Weckselblatt et Rudd, 2015 n https://biologyboom.com/wp-content/uploads/2014/07/image001.jpg Structural variability of chromosomes nChanges in genome greater than 1000 bases (1 kb) n= submicroscopic changes, microscopic changes, changes in the number of individual chromosomes, changes in genomes... n na) Quantitative - copy number variation (CNVs) - deletion, duplication, insertion nb) Positional - translocations nc) Orientation - inversion n nCopy number variation (CNVs) - segments of DNA greater than 1 kb present in a variable copy number compared to the reference genome n Congenital chromozomal abberations (CHAs) nFor every child conceived, there is a general genetic risk of 3 - 5% of being born with a congential disorder !!! n nCongenital CHA: 50 - 60% of first trimester abortions n 0.56% of live births have CHA nof which 0.1% trisomy 21, 0.1% Robertsonian translocation or other balanced translocation, 0.1% XXX,XXY, or XYY, 0.05% unbalanced change, 0.01% trisomy 18 or 13 n n nNDD, psychomotor retardation, sterility, testicular hypoplasia, amenorrhea, obesity, autism, etc. n nCHAs - responsible for about 100 clinically defined syndromes n Designation According to: discoverer.....Down Syndrome by symptoms .....cat cry syndrome by location..... del 1p36 syndrome General origin of chromosomal aberations nA) spontaneous (DNA damage + replication, replication, recombination, segregation defects...) nB) induced by clastogens (ionizing radiation, UV radiation, chemicals, viruses...DNA damage) Structural aberration - the crucial lesion is a double-stranded DNA break (DSB)! unrepaired = lethal incorrectly repaired = aberration -Congenital aberrations -Acquired aberrations DSB effects on DNA Pandey and Raghavan 2018 Molecular mechanisms of creation of structural chromosomal aberrations DNA damage and lack of repair nformation of a double-strand break (DSB) + new telomeric sequences - terminal deletion nDSB + telomere loss - breakage-fusion-bridge cycle n DNA damage and repair failures: nDSBs and repair defects via homologous recombination (HR) nDSB and non-homologous end joining (NHEJ) and microhomology mediated end joining (MMEJ) n Replication disorders: nReplication fork stalling and jumping to a different template (FoSTeS) nReplication induced by DNA microhomology breaks (MMBIR) Recombination disorders: nnon-allelic homologous recombination (NAHR) - unequal crossing over Reparation of DSBs – mechanism of homologous / non-homomologous repairs (NHEJ, MMEJ) MMEJ 5-25 bp homologie NHEJ Examples of CHAs created by by non-homologous end joining (NHEJ) Chen at al. 2010 Failures of replication and chromosomal aberrations Hastings et al. 2010 MMBIR - replication induced by DNA microhomology breaks - initiated by a break in a template strand, skipping and restarting replication on another template - temporary replication fork on another chromosome or sister chromatid - complex changes - deletion, duplication FoSTeS - stopping the replication fork and jumping to a different template - during replication, the replication fork is blocked (single strand break or secondary structure formation), the lagging DNA strand is released and jumps to another replication fork that shares the microhomology region - deletion, duplication Duplications in genome – „hot spots“ for chromosomal aberrations nmore than 5.4% of the human genome is covered by duplications (>1 kb with >90% sequence homology), average 18.5 kb n n1%-14% of each chromosome contains SDs (segmental duplications) n3.4% to 10.7% of genes may be duplicated n nThe human genome consists nlow copy number repetitive sequences (LCRs) - intrachromosomal duplications of >10 kb with >97% sequence identity n nsegmental duplications (SDs) - segments >1 kb in size with >90% sequence identity n nretrotransposons LINE sequences (L1 6 kb - 516,000 copies), SINE (Alu sequences - 300 bp - 1 to 1.5 million copies) Zhang et al. 2004 Intrachromozomové oblasti SDs v lidském genomu Genome-wide view of segmental duplications. The positions of alignments are depicted in red for each of the 24 chromosomes. Panels separate alignments on the basis of similarity: (B) 98%–100% identity. Purple bars depict centromeric gaps as well as the p-arms of acrocentric chromosomes (13, 14, 15, 21, and 22).Because of scale constraints, only alignments >5 kb are visible. Views were generated with the program PARASIGHT (J.A. Bailey, unpubl.), a graphical pairwise alignment viewer. Bailey et al 2001 duplications (90%-98%; ≥1 kb) cover 3.6% of all sequences Accumulation in pericentric and subtelomeric regions of chromosomes interstitial regions chromosomes "hot-spot" for NAHR (169 ?) Non-allelic homologous recombination (NAHR) recurent deletions / duplications / inversions ndefective meiotic recombination between repetitive sequences (SDs or LCRs) Genomic disorders Genomic disorders - diseases resulting from genomic rearrangements causing gene gain or loss (CNVs) The genome architecture creates suitable conditions for non-allelic homologous recombination, which can result in chromosomal rearrangements (NAHR) Microdeletion/duplication syndromes Genomic rearrangements resulting from recombination between repeated sequences. Mikrodeletion syndromes - pathological CNVs na group of genetically determined diseases caused by small microdeletions of DNA segments (2-4 Mb) that are not detectable by classical cytogenetic methods npatients share specific clinical symptoms...previously described by phenotype ("phenotype first"...) nnow "genotype first" approach ...first finding, size comparison, genes - influence on phenotype nsubset of CNVs....pathogenic CNVs ! nrecurrent - arise repeatedly at the same location on the chromosome ...e.g. del 22q11....areas with LCRs... nnon-recurrent - can arise anywhere in the genome ... Incidence of microdeletions / microduplications in human genome Weise et al. 2012 del2 Recurent microdeletions Examples of LCRs in known syndromes Genetic effects of microdeletions nAD heritability - haploinsufficiency - inability of one copy of the genetic material to maintain the functions that are provided by both copies n nheterozygote - monosomy for that part of the genetic information that is located on the corresponding segment of the normal chromosome n nloss of heterozygosity - LOH (loss of the dominant allele.....) n ndeletion of multiple genes - „contiguous gene syndromes“ nPathological effects of the patient are connected to the size of the microdeletion..... nincomplete penetrance and variable expressivity..... Mikrodeletion 22q11 DiGeorge (VCFS) syndrome nautosomal dominant with variable expressivity ndiscovered in 1965 by pediatric endocrinologist Angelo DiGeorge nfrequency 1:4000 to 1:6000 live births nalso referred to as Velo-Cardio-Facial Syndrome (VCFS) or CATCH 22 ntypical congenital heart defects (also detected n prenatally !) nFacial dysmorphia nhypoplasia - thymus aplasia (Tbx1 gene) n or parathyroid glands - ncalcium deficiency, cramps nimmunodefects n(absence of T-lymphocytes) Fallotova tetralogie: A: stenóza plicnice B: dextropozice aorty C: defekt komorového septa D: hypertrofie pravé komory Cri-du-chat (cat's cry) syndrome n Mainardi CP, 2006 Comparison of sizes of 5p deletions - phenotypy - candidate genes 1: 5000 Microdeletion syndrome 1p36 - patient OLG FN Brno nProband - 4 year old boy n nsevere PMR nfacial dysmorphia nAutism ntendency to self-harm nkaryotype normal n nmicrodeletion 1p36 - 3.2 Mb Microdeletion 22q11 DiGeorge (VCFS) syndrome nautosomal dominant with variable expressivity ndiscovered in 1965 by pediatric endocrinologist Angelo DiGeorge nfrequency 1:4000 to 1:6000 live births nalso referred to as Velo-Cardio-Facial Syndrome (VCFS) or CATCH 22 ntypical congenital heart defects (also detected n prenatally !) nFacial dysmorphia nhypoplasia - thymus aplasia (Tbx1 gene) n or parathyroid glands - ncalcium deficiency, cramps nimmunodefects n(absence of T-lymphocytes) final-22q-infograph-pano CATCH 22 - del(22)(q11) nCardiac defect - typical heart defects nAbnormal faces - chin nThymic hypoplasia - immune disorders, infections nCleft palate - cleft palate nHypocalcemia - convulsions n n nmicrodeletion size 22q11 1,5 - 3 Mb nrarely - deletion 10p n(DiGeorge II) n 10 % 80% of patients Photos of DG/VCFS patients prominent chin, low set ears, prominent nose.... CHRIS ...developmental delay - mild mental retardation, schizophrenia, autism.... ...speech disorders, especially slurring of speech, articulation disorders, speech fluency.... DG/VCFS and detection techniques (general approach foe all microeletion syndromes) nApproximately 90% of probands have de novo deletion of 22q11, about 6% have familial transmission ncytogenetic analysis of karyotype detects only 10-20% of 22q11 microdeletions na small percentage of patients may have a translocation or monosomy of chromosome 22 nmicrodeletion testing is performed by FISH, MLPA or aCGH nalways starting with the karyotype ! DiGeorgův syndrom – Wikipedie Velocardiofacial Syndrome: Background, Pathophysiology, Etiology DiGeorge DIGPOZ~1 I-FISH Array-CGH profile of patient with microdeletion 22q11 of size 2,72 Mb ISCN: arr[GRCh37] 22q11.21(18818376_21540347)x1 Array –CGH – size of microdeletion Case report - child with DG/VCFS nmicrodeletions can also arise secondarily n n nresult of abnormal chromosome segregation with balanced translocations or inversions n nE.g. healthy parent with balanced translocation - affected child with deletion or duplication nPhoto of the proband at 2 months of age showing micrognathia, hypertelorism, n prominent tubular nose, dysplastic lowset nears, thin lips, carp-shaped mouth, and na short neck. Translocation and two different microdeletions in one patient! n Moleclary-cytogenetic investigation of proband -Translocation t(14;22) -Deletion 22q11 -Deletion 14q n ..but mother was healthy..? Penetrance ? Prader-Willi Angelman syndrome (microdeletion + uniparental disomy) n n u abnormalities in n chromozome 15 n in 15q11-q13 n u clinically different n syndromes!! u Juan Carreño de Miranda (1680) „The nude monster“ PW syndrome? Genetické příčiny vzniku PWS a AS nPrader-Willi syndrome n n1. Deletion on paternal n chromosome 15 n (70%) n2. Maternal uniparental n disomy on chromosome 15 (20-25%) n3. Change in imprinting n (2 - 4 %) n4. Different chromosomal n rearrangements n ( less than 5%) n n Angelman syndrome n n 1.Deletion on the maternal n chromosome 15 (70 %) n n2. Paternal uniparental n disomy on chromosome n 15 (4 %) n n3. Change in imprinting 1 %) n n4. Various chromosomal n rearrangements (2 %) n n5. Mutations in the UBE3A n gene (3-5 %) GENOMIC IMPRINTING Prader –Willi/Angelman syndrome nepigenetic form of gene regulation that leads to functional haploidy: parent-specific monoallelic expression of ! nloss of expression of paternal (PWS) or maternal (AS) genes causes disease •a reversible process whereby sex-specific modification of genes in the parental generation leads to functional differences between the paternal and maternal genomes (alleles) in the offspring n n Smith and Hung, 2016 Critical region for PW / AS in chromozome 15 Prader-Willi syndrome (del 15q11-q13) npaternal deletion nlow fetal activity nhypotonia nexcessive weight gain, hyperphagia nshort stature nhypogonadism nmental retardation nhypopigmentation nskeletal development delay (acromicria) Chaloupková Angelman syndrom ( happy puppet syndrome) n nHarry Angelman (1965) – first description nOccurrence: frequency not exactly known -estimate about 1:15,000- 1:30,000 -in both sexes and all races Angelman syndrome (del 15q11-q13) „Happy Puppet“ nmaternal deletion nhard mental retardation nhypotonia nepilepsia, seizures nhypopigmentation nhyperactivity nspeech absence nprominent scull shape (mandibul, microcephaly, flat back of head..) n„happy character“ nmovement or balance disorder n Kubíková 2 4 FISH probe - Vysis Locus SNRPN PW syndrom Control probe 922-99upr William-Beuren syndrome nautosomal dominant disease with variable expressivity, usually de novo nrisk of the same disability is 50% for children of probands nincidence 1:20 000 live births ncause: del (7)(q11.23), the deletion region of about 1.5 Mb includes at least 17 genes, the most important being the ELN gene encoding elastin ndetekce: FISH sonda Vysis – LSI Elastin gene 7q11.23 Spectrum Orange/ 7q31 Spectrum Green control probe DNA nMLPA, aCGH n n Williams Beuren syndrome (del 7q11) ndevelopmental delay nmental disability nfailure to thrive nheart defects (heart murmur, narrowing of main blood vessels) nflattened nasal bridge nwidely spaced teeth nhypercalcemia ngastrointestinal problems nurinary difficulties n n n http://www.thespeciallife.com/images/Williams-Syndrome.jpg http://geneticsf.labanca.net/wp-content/uploads/2010/11/Williams-syndrome.jpg https://www.google.cz/search?q=williams+beuren+syndrome&source=lnms&tbm=isch&sa=X&ei=MimGUvDpC4GctQ aJhYGwCg&ved=0CAcQ_AUoAQ&biw=1920&bih=989#facrc=_&imgdii=_&imgrc=HTjyFEuSnZo4JM%3A%3BkqK81uaGRtKPIM %3Bhttp%253A%252F%252Fgeneticsf.laba https://www.google.cz/search?q=williams+beuren+syndrome&source=lnms&tbm=isch&sa=X&ei=MimGUvDpC4GctQ aJhYGwCg&ved=0CAcQ_AUoAQ&biw=1920&bih=989#facrc=_&imgdii=_&imgrc=MdTDkcoWBwg-WM%3A%3BTKKZzTKDfniYlM %3Bhttp%253A%252F%252Fwww.thespeciallife.com%252Fimages%252F Williams – Beurenův syndrome: case report nBirth: 2010 nPhenotype: NDD, stigmatizatio nCause: del 7q11.23 (1,4 Mb) G:\Koláčková_A_10_1.JPG G:\Koláčková_A_10_3.JPG G:\Koláčková_A_10_4.JPG obr2a Výsledek vyšetření – profil array-CGH s mikrodelecí 7q11.23