@article {708, title = {Genetic and molecular mechanism for distinct clinical phenotypes conveyed by allelic truncating mutations implicated in FBN1.}, journal = {Mol Genet Genomic Med}, volume = {8}, year = {2020}, month = {2020 01}, pages = {e1023}, abstract = {

BACKGROUND: The molecular and genetic mechanisms by which different single nucleotide variant alleles in specific genes, or at the same genetic locus, cause distinct disease phenotypes often remain unclear. Allelic truncating mutations of FBN1 could cause either classical Marfan syndrome (MFS) or a more complicated phenotype associated with Marfanoid-progeroid-lipodystrophy syndrome (MPLS).

METHODS: We investigated a small cohort, encompassing two classical MFS and one MPLS subjects~from China, whose clinical presentation included scoliosis potentially requiring surgical intervention. Targeted next generation sequencing was performed on all the participants. We analyzed the molecular diagnosis, clinical features, and the potential molecular mechanism involved in the MPLS subject in our cohort.

RESULTS: We report a novel de novo FBN1 mutation for the first Chinese subject~with MPLS, a more complicated fibrillinopathy, and two subjects~with more classical MFS. We further predict that the MPLS truncating mutation, and others previously reported, is prone to escape the nonsense-mediated decay (NMD), while MFS mutations are predicted to be subjected to NMD. Also, the MPLS mutation occurs within the glucogenic hormone asprosin domain of FBN1. In vitro experiments showed that the single MPLS mutation p.Glu2759Cysfs*9 appears to perturb proper FBN1 protein aggregation as compared with the~classical MFS mutation p.Tyr2596Thrfs*86. Both mutations appear to upregulate SMAD2 phosphorylation in vitro.

CONCLUSION: We provide direct evidence that a dominant-negative interaction of FBN1 potentially explains the complex MPLS phenotypes through genetic and functional analysis. Our study expands the mutation spectrum of FBN1 and highlights the potential molecular mechanism for MPLS.

}, keywords = {Adolescent, Adult, Child, Female, Fibrillin-1, HEK293 Cells, Humans, Lipodystrophy, Marfan Syndrome, Mutation, Phenotype, Progeria, Protein Domains, Smad2 Protein}, issn = {2324-9269}, doi = {10.1002/mgg3.1023}, author = {Lin, Mao and Liu, Zhenlei and Liu, Gang and Zhao, Sen and Li, Chao and Chen, Weisheng and Coban Akdemir, Zeynep and Lin, Jiachen and Song, Xiaofei and Wang, Shengru and Xu, Qiming and Zhao, Yanxue and Wang, Lianlei and Zhang, Yuanqiang and Yan, Zihui and Liu, Sen and Liu, Jiaqi and Chen, Yixin and Zuo, Yuzhi and Yang, Xu and Sun, Tianshu and Yang, Xin-Zhuang and Niu, Yuchen and Li, Xiaoxin and You, Wesley and Qiu, Bintao and Ding, Chen and Liu, Pengfei and Zhang, Shuyang and Carvalho, Claudia M B and Posey, Jennifer E and Qiu, Guixing and Lupski, James R and Wu, Zhihong and Zhang, Jianguo and Wu, Nan} } @article {720, title = {Xq22 deletions and correlation with distinct neurological disease traits in females: Further evidence for a contiguous gene syndrome.}, journal = {Hum Mutat}, volume = {41}, year = {2020}, month = {2020 01}, pages = {150-168}, abstract = {

Xq22 deletions that encompass PLP1 (Xq22-PLP1-DEL) are notable for variable expressivity of neurological disease traits in females ranging from a mild late-onset form of spastic paraplegia type 2 (MIM$\#$ 312920), sometimes associated with skewed X-inactivation, to an early-onset neurological disease trait (EONDT) of severe developmental delay, intellectual disability, and behavioral abnormalities. Size and gene content of Xq22-PLP1-DEL vary and were proposed as potential molecular etiologies underlying variable expressivity in carrier females where two smallest regions of overlap (SROs) were suggested to influence disease. We ascertained a cohort of eight unrelated patients harboring Xq22-PLP1-DEL and performed high-density array comparative genomic hybridization~and breakpoint-junction sequencing. Molecular characterization of Xq22-PLP1-DEL from 17 cases (eight herein and nine published) revealed an overrepresentation of breakpoints that reside within repeats (11/17, ~65\%) and the clustering of ~47\% of proximal breakpoints in a genomic instability hotspot with characteristic non-B DNA density. These findings implicate a potential role for genomic architecture in stimulating the formation of Xq22-PLP1-DEL. The correlation of Xq22-PLP1-DEL gene content with neurological disease trait in female cases enabled refinement of the associated SROs to a single genomic interval containing six genes. Our data support the hypothesis that genes contiguous to PLP1 contribute to EONDT.

}, keywords = {Child, Child, Preschool, Chromosome Breakpoints, Chromosome Deletion, Chromosome Mapping, Chromosomes, Human, X, Comparative Genomic Hybridization, Female, Genetic Association Studies, Genetic Predisposition to Disease, Humans, Male, Nervous System Diseases, Pedigree, Phenotype, Quantitative Trait, Heritable, Repetitive Sequences, Nucleic Acid, Sex Factors, Syndrome, X Chromosome Inactivation}, issn = {1098-1004}, doi = {10.1002/humu.23902}, author = {Hijazi, Hadia and Coelho, Fernanda S and Gonzaga-Jauregui, Claudia and Bernardini, Laura and Mar, Soe S and Manning, Melanie A and Hanson-Kahn, Andrea and Naidu, Sakkubai and Srivastava, Siddharth and Lee, Jennifer A and Jones, Julie R and Friez, Michael J and Alberico, Thomas and Torres, Barbara and Fang, Ping and Cheung, Sau Wai and Song, Xiaofei and Davis-Williams, Angelique and Jornlin, Carly and Wight, Patricia A and Patyal, Pankaj and Taube, Jennifer and Poretti, Andrea and Inoue, Ken and Zhang, Feng and Pehlivan, Davut and Carvalho, Claudia M B and Hobson, Grace M and Lupski, James R} } @article {446, title = {WNT Signaling Perturbations Underlie the Genetic Heterogeneity of Robinow Syndrome.}, journal = {Am J Hum Genet}, volume = {102}, year = {2018}, month = {2018 01 04}, pages = {27-43}, abstract = {

Locus heterogeneity characterizes a variety of skeletal dysplasias often due to interacting or overlapping signaling pathways. Robinow syndrome is a skeletal disorder historically refractory to molecular diagnosis, potentially stemming from substantial genetic heterogeneity. All current known pathogenic variants reside in genes within the noncanonical Wnt signaling pathway including ROR2, WNT5A, and more recently, DVL1 and DVL3. However, \~{}70\% of autosomal-dominant Robinow syndrome cases remain molecularly unsolved. To investigate this missing heritability, we recruited 21 families with at least one family member clinically diagnosed with Robinow or Robinow-like phenotypes and performed genetic and genomic studies. In total, four families with variants in FZD2 were identified as well as three individuals from two families with biallelic variants in NXN that co-segregate with the phenotype. Importantly, both FZD2 and NXN are relevant protein partners in the WNT5A interactome, supporting their role in skeletal development. In addition to confirming that clustered -1 frameshifting variants in DVL1 and DVL3 are the main contributors to dominant Robinow syndrome, we also found likely pathogenic variants in candidate genes GPC4 and RAC3, both linked to the Wnt signaling pathway. These data support an initial hypothesis that Robinow syndrome results from perturbation of the Wnt/PCP pathway, suggest specific relevant domains of the proteins involved, and reveal key contributors in this signaling cascade during human embryonic development. Contrary to the view that non-allelic genetic heterogeneity hampers gene discovery, this study demonstrates the utility of rare disease genomic studies to parse gene function in human developmental pathways.

}, keywords = {Adolescent, Adult, Base Sequence, Child, Child, Preschool, Chromosome Segregation, Craniofacial Abnormalities, Diagnosis, Differential, Dwarfism, Female, Genes, Dominant, Genetic Association Studies, Genetic Heterogeneity, Humans, Limb Deformities, Congenital, Male, Middle Aged, Mutation, Missense, Phenotype, Urogenital Abnormalities, Wnt Signaling Pathway}, issn = {1537-6605}, doi = {10.1016/j.ajhg.2017.10.002}, author = {White, Janson J and Mazzeu, Juliana F and Coban-Akdemir, Zeynep and Bayram, Yavuz and Bahrambeigi, Vahid and Hoischen, Alexander and van Bon, Bregje W M and Gezdirici, Alper and Gulec, Elif Yilmaz and Ramond, Francis and Touraine, Renaud and Thevenon, Julien and Shinawi, Marwan and Beaver, Erin and Heeley, Jennifer and Hoover-Fong, Julie and Durmaz, Ceren D and Karabulut, Halil Gurhan and Marzioglu-Ozdemir, Ebru and Cayir, Atilla and Duz, Mehmet B and Seven, Mehmet and Price, Susan and Ferreira, Barbara Merfort and Vianna-Morgante, Angela M and Ellard, Sian and Parrish, Andrew and Stals, Karen and Flores-Daboub, Josue and Jhangiani, Shalini N and Gibbs, Richard A and Brunner, Han G and Sutton, V Reid and Lupski, James R and Carvalho, Claudia M B} } @article {361, title = {Dual molecular diagnosis contributes to atypical Prader-Willi phenotype in monozygotic twins.}, journal = {Am J Med Genet A}, volume = {173}, year = {2017}, month = {2017 Sep}, pages = {2451-2455}, abstract = {

We describe monozygotic twin girls with genetic variation at two separate loci resulting in a blended phenotype of Prader-Willi syndrome and Pitt-Hopkins syndrome. These girls were diagnosed in early infancy with Prader-Willi syndrome, but developed an atypical phenotype, with apparent intellectual deficiency and lack of obesity. Array-comparative genomic hybridization confirmed a de novo paternal deletion of the 15q11.2q13 region and exome sequencing identified a second mutational event in both girls, which was a novel variant c.145+1G>A affecting a TCF4 canonical splicing site inherited from the mosaic mother. RNA studies showed that the variant abolished the donor splicing site, which was accompanied by activation of an alternative non-canonical splicing-site which then predicts a premature stop codon in the following exon. Clinical re-evaluation of the twins indicated that both variants are likely contributing to the more severe phenotypic presentation. Our data show that atypical clinical presentations may actually be the expression of blended clinical phenotypes arising from independent pathogenic events at two loci.

}, keywords = {Adolescent, Base Sequence, Child, Chromosome Deletion, Chromosomes, Human, Pair 15, Comparative Genomic Hybridization, Exome, Facies, Female, Humans, Hyperventilation, Intellectual Disability, Obesity, Pathology, Molecular, Phenotype, Prader-Willi Syndrome, Transcription Factor 4, Twins, Monozygotic}, issn = {1552-4833}, doi = {10.1002/ajmg.a.38315}, author = {Jehee, Fernanda S and de Oliveira, Valdirene T and Gurgel-Giannetti, Juliana and Pietra, Rafaella X and Rubatino, Fernando V M and Carobin, Nat{\'a}lia V and Vianna, Gabrielle S and de Freitas, Mariana L and Fernandes, Karla S and Ribeiro, Beatriz S V and Br{\"u}ggenwirth, Hennie T and Ali-Amin, Roza and White, Janson J and Akdemir, Zeynep C and Jhangiani, Shalini N and Gibbs, Richard A and Lupski, James R and Varela, Monica C and Koiffmann, C{\'e}lia and Rosenberg, Carla and Carvalho, Claudia M B} } @article {366, title = {Efficient CNV breakpoint analysis reveals unexpected structural complexity and correlation of dosage-sensitive genes with clinical severity in genomic disorders.}, journal = {Hum Mol Genet}, volume = {26}, year = {2017}, month = {2017 05 15}, pages = {1927-1941}, abstract = {

Genomic disorders are the clinical conditions manifested by submicroscopic genomic rearrangements including copy number variants (CNVs). The CNVs can be identified by array-based comparative genomic hybridization (aCGH), the most commonly used technology for molecular diagnostics of genomic disorders. However, clinical aCGH only informs CNVs in the probe-interrogated regions. Neither orientational information nor the resulting genomic rearrangement structure is provided, which is a key to uncovering mutational and pathogenic mechanisms underlying genomic disorders. Long-range polymerase chain reaction (PCR) is a traditional approach to obtain CNV breakpoint junction, but this method is inefficient when challenged by structural complexity such as often found at the PLP1 locus in association with Pelizaeus-Merzbacher disease (PMD). Here we introduced {\textquoteright}capture and single-molecule real-time sequencing{\textquoteright} (cap-SMRT-seq) and newly developed {\textquoteright}asymmetry linker-mediated nested PCR walking{\textquoteright} (ALN-walking) for CNV breakpoint sequencing in 49 subjects with PMD-associated CNVs. Remarkably, 29 (94\%) of the 31 CNV breakpoint junctions unobtainable by conventional long-range PCR were resolved by cap-SMRT-seq and ALN-walking. Notably, unexpected CNV complexities, including inter-chromosomal rearrangements that cannot be resolved by aCGH, were revealed by efficient breakpoint sequencing. These sequence-based structures of PMD-associated CNVs further support the role of DNA replicative mechanisms in CNV mutagenesis, and facilitate genotype-phenotype correlation studies. Intriguingly, the lengths of gained segments by CNVs are strongly correlated with clinical severity in PMD, potentially reflecting the functional contribution of other dosage-sensitive genes besides PLP1. Our study provides new efficient experimental approaches (especially ALN-walking) for CNV breakpoint sequencing and highlights their importance in uncovering CNV mutagenesis and pathogenesis in genomic disorders.

}, keywords = {Base Sequence, Comparative Genomic Hybridization, DNA Copy Number Variations, DNA Replication, Female, Gene Dosage, Gene Duplication, Gene Rearrangement, Genetic Association Studies, Genome, Human, Genomics, Humans, Male, Mutation, Pelizaeus-Merzbacher Disease, Sequence Analysis, DNA}, issn = {1460-2083}, doi = {10.1093/hmg/ddx102}, author = {Zhang, Ling and Wang, Jingmin and Zhang, Cheng and Li, Dongxiao and Carvalho, Claudia M B and Ji, Haoran and Xiao, Jianqiu and Wu, Ye and Zhou, Weichen and Wang, Hongyan and Jin, Li and Luo, Yang and Wu, Xiru and Lupski, James R and Zhang, Feng and Jiang, Yuwu} } @article {348, title = {An Organismal CNV Mutator Phenotype Restricted to Early Human Development.}, journal = {Cell}, volume = {168}, year = {2017}, month = {2017 02 23}, pages = {830-842.e7}, abstract = {

De novo copy number variants (dnCNVs) arising at~multiple loci in a personal genome have usually been considered to reflect cancer somatic genomic instabilities. We describe a multiple dnCNV (MdnCNV) phenomenon in which individuals with genomic disorders carry five to ten constitutional dnCNVs. These CNVs originate from independent formation incidences, are predominantly tandem duplications or complex gains, exhibit breakpoint junction features reminiscent of replicative repair, and show increased de novo point mutations flanking the rearrangement junctions. The active CNV mutation shower appears to be restricted to a transient perizygotic period. We~propose that a defect in the CNV formation process is responsible for the "CNV-mutator state," and this state is dampened after early embryogenesis. The constitutional MdnCNV phenomenon resembles chromosomal instability in various cancers. Investigations of this phenomenon may provide unique access to understanding genomic disorders, structural variant mutagenesis, human evolution, and cancer biology.

}, keywords = {Chromosome Aberrations, Chromosome Breakpoints, Chromosome Duplication, DNA Copy Number Variations, DNA Replication, Embryonic Development, Female, Gametogenesis, Genetic Diseases, Inborn, Genomic Instability, Humans, Male, Mutation}, issn = {1097-4172}, doi = {10.1016/j.cell.2017.01.037}, author = {Liu, Pengfei and Yuan, Bo and Carvalho, Claudia M B and Wuster, Arthur and Walter, Klaudia and Zhang, Ling and Gambin, Tomasz and Chong, Zechen and Campbell, Ian M and Coban Akdemir, Zeynep and Gelowani, Violet and Writzl, Karin and Bacino, Carlos A and Lindsay, Sarah J and Withers, Marjorie and Gonzaga-Jauregui, Claudia and Wiszniewska, Joanna and Scull, Jennifer and Stankiewicz, Pawe{\l} and Jhangiani, Shalini N and Muzny, Donna M and Zhang, Feng and Chen, Ken and Gibbs, Richard A and Rautenstrauss, Bernd and Cheung, Sau Wai and Smith, Janice and Breman, Amy and Shaw, Chad A and Patel, Ankita and Hurles, Matthew E and Lupski, James R} } @article {390, title = {REST Final-Exon-Truncating Mutations Cause Hereditary Gingival Fibromatosis.}, journal = {Am J Hum Genet}, volume = {101}, year = {2017}, month = {2017 Jul 06}, pages = {149-156}, abstract = {

Hereditary gingival fibromatosis (HGF) is the most common genetic form of gingival fibromatosis that develops as a slowly progressive, benign, localized or generalized enlargement of keratinized gingiva. HGF is a genetically heterogeneous disorder and can be transmitted either as an autosomal-dominant or autosomal-recessive trait or appear sporadically. To date, four loci (2p22.1, 2p23.3-p22.3, 5q13-q22, and 11p15) have been mapped to autosomes and one gene (SOS1) has been associated with the HGF trait observed to segregate in a dominant inheritance pattern. Here we report 11 individuals with HGF from three unrelated families. Whole-exome sequencing (WES) revealed three different truncating mutations including two frameshifts and one nonsense variant in RE1-silencing transcription factor (REST) in the probands from all families and further genetic and genomic analyses confirmed the WES-identified findings. REST is a transcriptional repressor that is expressed throughout the body; it has different roles in different cellular contexts, such as oncogenic and tumor-suppressor functions and hematopoietic and cardiac differentiation. Here we show the consequences of germline final-exon-truncating mutations in REST for organismal development and the association with the HGF phenotype.

}, keywords = {Adolescent, Base Sequence, Chromosome Segregation, Exons, Family, Female, Fibromatosis, Gingival, Genetic Predisposition to Disease, Humans, Male, Middle Aged, Mutation, Pedigree, Repressor Proteins}, issn = {1537-6605}, doi = {10.1016/j.ajhg.2017.06.006}, author = {Bayram, Yavuz and White, Janson J and Elcioglu, Nursel and Cho, Megan T and Zadeh, Neda and Gedikbasi, Asuman and Palanduz, Sukru and Ozturk, Sukru and Cefle, Kivanc and Kasapcopur, Ozgur and Coban Akdemir, Zeynep and Pehlivan, Davut and Begtrup, Amber and Carvalho, Claudia M B and Paine, Ingrid Sophie and Mentes, Ali and Bektas-Kayhan, Kivanc and Karaca, Ender and Jhangiani, Shalini N and Muzny, Donna M and Gibbs, Richard A and Lupski, James R} } @article {7, title = {Whole-Genome Sequencing of Cytogenetically Balanced Chromosome Translocations Identifies Potentially Pathological Gene Disruptions and Highlights the Importance of Microhomology in the Mechanism of Formation.}, journal = {Hum Mutat}, volume = {38}, year = {2017}, month = {2017 Feb}, pages = {180-192}, abstract = {

Most balanced translocations are thought to result mechanistically from nonhomologous end joining or, in rare cases of recurrent events, by nonallelic homologous recombination. Here, we use low-coverage mate pair whole-genome sequencing to fine map rearrangement breakpoint junctions in both phenotypically normal and affected translocation carriers. In total, 46 junctions from 22 carriers of balanced translocations were characterized. Genes were disrupted in 48\% of the breakpoints; recessive genes in four normal carriers and known dominant intellectual disability genes in three affected carriers. Finally, seven candidate disease genes were disrupted in five carriers with neurocognitive disabilities (SVOPL, SUSD1, TOX, NCALD, SLC4A10) and one XX-male carrier with Tourette syndrome (LYPD6, GPC5). Breakpoint junction analyses revealed microhomology and small templated insertions in a substantive fraction of the analyzed translocations (17.4\%; n = 4); an observation that was substantiated by reanalysis of 37 previously published translocation junctions. Microhomology associated with templated insertions is a characteristic seen in the breakpoint junctions of rearrangements mediated by error-prone replication-based repair mechanisms. Our data implicate that a mechanism involving template switching might contribute to the formation of at least 15\% of the interchromosomal translocation events.

}, keywords = {Base Sequence, Chromosome Breakage, Chromosome Mapping, Comparative Genomic Hybridization, DNA Copy Number Variations, Female, Genetic Association Studies, Genomics, Genotype, Homologous Recombination, Humans, In Situ Hybridization, Fluorescence, Karyotype, Male, Phenotype, Translocation, Genetic, Whole Genome Sequencing}, issn = {1098-1004}, doi = {10.1002/humu.23146}, author = {Nilsson, Daniel and Pettersson, Maria and Gustavsson, Peter and F{\"o}rster, Alisa and Hofmeister, Wolfgang and Wincent, Josephine and Zachariadis, Vasilios and Anderlid, Britt-Marie and Nordgren, Ann and M{\"a}kitie, Outi and Wirta, Valtteri and K{\"a}ller, Max and Vezzi, Francesco and Lupski, James R and Nordenskj{\"o}ld, Magnus and Lundberg, Elisabeth Syk and Carvalho, Claudia M B and Lindstrand, Anna} } @article {30, title = {Copy-Number Variation Contributes to the Mutational Load of Bardet-Biedl Syndrome.}, journal = {Am J Hum Genet}, volume = {99}, year = {2016}, month = {2016 Aug 04}, pages = {318-36}, abstract = {

Bardet-Biedl syndrome (BBS) is a defining ciliopathy, notable for extensive allelic and genetic heterogeneity, almost all of which has been identified through sequencing. Recent data have suggested that copy-number variants (CNVs) also contribute to BBS. We used a custom oligonucleotide array comparative genomic hybridization (aCGH) covering 20 genes that encode intraflagellar transport (IFT) components and 74 ciliopathy loci to screen 92 unrelated individuals with BBS, irrespective of their known mutational burden. We identified 17 individuals with exon-disruptive CNVs (18.5\%), including 13 different deletions in eight BBS genes (BBS1, BBS2, ARL6/BBS3, BBS4, BBS5, BBS7, BBS9, and NPHP1) and a deletion and a duplication in other ciliopathy-associated genes (ALMS1 and NPHP4, respectively). By contrast, we found a single heterozygous exon-disruptive event in a BBS-associated gene (BBS9) in 229 control subjects. Superimposing these data with resequencing revealed CNVs to (1) be sufficient to cause disease, (2) Mendelize heterozygous deleterious alleles, and (3) contribute oligogenic alleles by combining point mutations and exonic CNVs in multiple genes. Finally, we report a deletion and a splice site mutation in IFT74, inherited under a recessive paradigm, defining a candidate BBS locus. Our data suggest that CNVs contribute pathogenic alleles to a substantial fraction of BBS-affected individuals and highlight how either deletions or point mutations in discrete splice isoforms can induce hypomorphic mutations in genes otherwise intolerant to deleterious variation. Our data also suggest that CNV analyses and resequencing studies unbiased for previous mutational burden is necessary to delineate the complexity of disease architecture.

}, keywords = {Adolescent, Adult, Alleles, Animals, Bardet-Biedl Syndrome, Child, Child, Preschool, Chromosome Breakpoints, Cytoskeletal Proteins, DNA Copy Number Variations, Exons, Female, Gastrulation, Genes, Recessive, Humans, Infant, Male, Mutation, Pedigree, Young Adult, Zebrafish, Zebrafish Proteins}, issn = {1537-6605}, doi = {10.1016/j.ajhg.2015.04.023}, author = {Lindstrand, Anna and Frangakis, Stephan and Carvalho, Claudia M B and Richardson, Ellen B and McFadden, Kelsey A and Willer, Jason R and Pehlivan, Davut and Liu, Pengfei and Pediaditakis, Igor L and Sabo, Aniko and Lewis, Richard Alan and Banin, Eyal and Lupski, James R and Davis, Erica E and Katsanis, Nicholas} } @article {79, title = {DVL3 Alleles Resulting in a -1 Frameshift of the Last Exon Mediate Autosomal-Dominant Robinow Syndrome.}, journal = {Am J Hum Genet}, volume = {98}, year = {2016}, month = {2016 Mar 03}, pages = {553-561}, abstract = {

Robinow syndrome is a rare congenital disorder characterized by mesomelic limb shortening, genital hypoplasia, and distinctive facial features. Recent reports have identified, in individuals with dominant Robinow syndrome, a specific type of variant characterized by being uniformly located in the penultimate exon of DVL1 and resulting in a -1 frameshift allele with a premature termination codon that escapes nonsense-mediated decay. Here, we studied a cohort of individuals who had been clinically diagnosed with Robinow syndrome but who had not received a molecular diagnosis from variant studies of DVL1, WNT5A, and ROR2. Because of the uniform location of frameshift variants in DVL1-mediated Robinow syndrome and the functional redundancy of DVL1, DVL2, and DVL3, we elected to pursue direct Sanger sequencing of the penultimate exon of DVL1 and its paralogs DVL2 and DVL3 to search for potential disease-associated variants. Remarkably, targeted sequencing identified five unrelated individuals harboring heterozygous, de novo frameshift variants in DVL3, including two splice acceptor mutations and three 1 bp deletions. Similar to the variants observed in DVL1-mediated Robinow syndrome, all variants in DVL3 result in a -1 frameshift, indicating that these highly specific alterations might be a common cause of dominant Robinow syndrome. Here, we review the current knowledge of these peculiar variant alleles in DVL1- and DVL3-mediated Robinow syndrome and further elucidate the phenotypic features present in subjects with DVL1 and DVL3 frameshift mutations.

}, keywords = {Adaptor Proteins, Signal Transducing, Alleles, Base Sequence, Codon, Nonsense, Craniofacial Abnormalities, Dishevelled Proteins, Dwarfism, Exons, Female, Frameshift Mutation, Genetic Variation, Humans, Limb Deformities, Congenital, Male, Molecular Sequence Data, Phosphoproteins, Proto-Oncogene Proteins, Receptor Tyrosine Kinase-like Orphan Receptors, Sequence Analysis, DNA, Sequence Deletion, Urogenital Abnormalities, Wnt Proteins, Wnt-5a Protein}, issn = {1537-6605}, doi = {10.1016/j.ajhg.2016.01.005}, author = {White, Janson J and Mazzeu, Juliana F and Hoischen, Alexander and Bayram, Yavuz and Withers, Marjorie and Gezdirici, Alper and Kimonis, Virginia and Steehouwer, Marloes and Jhangiani, Shalini N and Muzny, Donna M and Gibbs, Richard A and van Bon, Bregje W M and Sutton, V Reid and Lupski, James R and Brunner, Han G and Carvalho, Claudia M B} } @article {119, title = {Mechanisms for the Generation of Two Quadruplications Associated with Split-Hand Malformation.}, journal = {Hum Mutat}, volume = {37}, year = {2016}, month = {2016 Feb}, pages = {160-4}, abstract = {

Germline copy-number variants (CNVs) involving quadruplications are rare and the mechanisms generating them are largely unknown. Previously, we reported a 20-week gestation fetus with split-hand malformation; clinical microarray detected two maternally inherited triplications separated by a copy-number neutral region at 17p13.3, involving BHLHA9 and part of YWHAE. Here, we describe an 18-month-old male sibling of the previously described fetus with split-hand malformation. Custom high-density microarray and digital droplet PCR revealed the copy-number gains were actually quadruplications in the mother, the fetus, and her later born son. This quadruplication-normal-quadruplication pattern was shown to be expanded from the triplication-normal-triplication CNV at the same loci in the maternal grandmother. We mapped two breakpoint junctions and demonstrated that both are mediated by Alu repetitive elements and identical in these four individuals. We propose a three-step process combining Alu-mediated replicative-repair-based mechanism(s) and intergenerational, intrachromosomal nonallelic homologous recombination to generate the quadruplications in this family.

}, keywords = {14-3-3 Proteins, Adult, Aged, Alu Elements, Base Sequence, Basic Helix-Loop-Helix Transcription Factors, Chromosome Duplication, Chromosomes, Human, Pair 17, DNA Copy Number Variations, Female, Genetic Loci, Genome, Human, Hand Deformities, Congenital, Humans, Infant, Male, Molecular Sequence Data, Pedigree, Sequence Alignment, Sequence Analysis, DNA}, issn = {1098-1004}, doi = {10.1002/humu.22929}, author = {Gu, Shen and Posey, Jennifer E and Yuan, Bo and Carvalho, Claudia M B and Luk, H M and Erikson, Kelly and Lo, Ivan F M and Leung, Gordon K C and Pickering, Curtis R and Chung, Brian H Y and Lupski, James R} } @article {345, title = {The role of combined SNV and CNV burden in patients with distal symmetric polyneuropathy.}, journal = {Genet Med}, volume = {18}, year = {2016}, month = {2016 05}, pages = {443-51}, abstract = {

PURPOSE: Charcot-Marie-Tooth (CMT) disease is a heterogeneous group of genetic disorders of the peripheral nervous system. Copy-number variants (CNVs) contribute significantly to CMT, as duplication of PMP22 underlies the majority of CMT1 cases. We hypothesized that CNVs and/or single-nucleotide variants (SNVs) might exist in patients with CMT with an unknown molecular genetic etiology.

METHODS: Two hundred patients with CMT, negative for both SNV mutations in several CMT genes and for CNVs involving PMP22, were screened for CNVs by high-resolution oligonucleotide array comparative genomic hybridization. Whole-exome sequencing was conducted on individuals with rare, potentially pathogenic CNVs.

RESULTS: Putatively causative CNVs were identified in five subjects (~2.5\%); four of the five map to known neuropathy genes. Breakpoint sequencing revealed Alu-Alu-mediated junctions as a predominant contributor. Exome sequencing identified MFN2 SNVs in two of the individuals.

CONCLUSION: Neuropathy-associated CNV outside of the PMP22 locus is rare in CMT. Nevertheless, there is potential clinical utility in testing for CNVs and exome sequencing in CMT cases negative for the CMT1A duplication. These findings suggest that complex phenotypes including neuropathy can potentially be caused by a combination of SNVs and CNVs affecting more than one disease-associated locus and contributing to a mutational burden.Genet Med 18 5, 443-451.

}, keywords = {Adult, Age of Onset, Charcot-Marie-Tooth Disease, Child, Preschool, Comparative Genomic Hybridization, DNA Copy Number Variations, Exome, Female, Genetic Predisposition to Disease, GTP Phosphohydrolases, High-Throughput Nucleotide Sequencing, Humans, Male, Mitochondrial Proteins, Motor Neurons, Myelin P0 Protein, Myelin Proteins, Neural Conduction, Polymorphism, Single Nucleotide, Polyneuropathies}, issn = {1530-0366}, doi = {10.1038/gim.2015.124}, author = {Pehlivan, Davut and Beck, Christine R and Okamoto, Yuji and Harel, Tamar and Akdemir, Zeynep H C and Jhangiani, Shalini N and Withers, Marjorie A and Goksungur, Meryem Tuba and Carvalho, Claudia M B and Czesnik, Dirk and Gonzaga-Jauregui, Claudia and Wiszniewski, Wojciech and Muzny, Donna M and Gibbs, Richard A and Rautenstrauss, Bernd and Sereda, Michael W and Lupski, James R} } @article {170, title = {Alu-mediated diverse and complex pathogenic copy-number variants within human chromosome 17 at p13.3.}, journal = {Hum Mol Genet}, volume = {24}, year = {2015}, month = {2015 Jul 15}, pages = {4061-77}, abstract = {

Alu repetitive elements are known to be major contributors to genome instability by generating Alu-mediated copy-number variants (CNVs). Most of the reported Alu-mediated CNVs are simple deletions and duplications, and the mechanism underlying Alu-Alu-mediated rearrangement has been attributed to non-allelic homologous recombination (NAHR). Chromosome 17 at the p13.3 genomic region lacks extensive low-copy repeat architecture; however, it is highly enriched for Alu repetitive elements, with a fraction of 30\% of total sequence annotated in the human reference genome, compared with the 10\% genome-wide and 18\% on chromosome 17. We conducted mechanistic studies of the 17p13.3 CNVs by performing high-density oligonucleotide array comparative genomic hybridization, specifically interrogating the 17p13.3 region with \~{}150 bp per probe density; CNV breakpoint junctions were mapped to nucleotide resolution by polymerase chain reaction and Sanger sequencing. Studied rearrangements include 5 interstitial deletions, 14 tandem duplications, 7 terminal deletions and 13 complex genomic rearrangements (CGRs). Within the 17p13.3 region, Alu-Alu-mediated rearrangements were identified in 80\% of the interstitial deletions, 46\% of the tandem duplications and 50\% of the CGRs, indicating that this mechanism was a major contributor for formation of breakpoint junctions. Our studies suggest that Alu repetitive elements facilitate formation of non-recurrent CNVs, CGRs and other structural aberrations of chromosome 17 at p13.3. The common observation of Alu-mediated rearrangement in CGRs and breakpoint junction sequences analysis further demonstrates that this type of mechanism is unlikely attributed to NAHR, but rather may be due to a recombination-coupled DNA replicative repair process.

}, keywords = {Alleles, Alu Elements, Base Sequence, Chromosomes, Human, Pair 17, Comparative Genomic Hybridization, DNA Copy Number Variations, Female, Gene Duplication, Gene Rearrangement, Genome, Human, Genomic Instability, Genomics, Homologous Recombination, Humans, Male, Molecular Sequence Data, Segmental Duplications, Genomic, Sequence Deletion}, issn = {1460-2083}, doi = {10.1093/hmg/ddv146}, author = {Gu, Shen and Yuan, Bo and Campbell, Ian M and Beck, Christine R and Carvalho, Claudia M B and Nagamani, Sandesh C S and Erez, Ayelet and Patel, Ankita and Bacino, Carlos A and Shaw, Chad A and Stankiewicz, Pawe{\l} and Cheung, Sau Wai and Bi, Weimin and Lupski, James R} } @article {120, title = {Nonrecurrent 17p11.2p12 Rearrangement Events that Result in Two Concomitant Genomic Disorders: The PMP22-RAI1 Contiguous Gene Duplication Syndrome.}, journal = {Am J Hum Genet}, volume = {97}, year = {2015}, month = {2015 Nov 05}, pages = {691-707}, abstract = {

The genomic duplication associated with Potocki-Lupski syndrome (PTLS) maps in close proximity to the duplication associated with Charcot-Marie-Tooth disease type 1A (CMT1A). PTLS is characterized by hypotonia, failure to thrive, reduced body weight, intellectual disability, and autistic features. CMT1A is a common autosomal dominant distal symmetric peripheral polyneuropathy. The key dosage-sensitive genes RAI1 and PMP22 are respectively associated with PTLS and CMT1A. Recurrent duplications accounting for the majority of subjects with these conditions are mediated by nonallelic homologous recombination between distinct low-copy repeat (LCR) substrates. The LCRs flanking a contiguous genomic interval encompassing both RAI1 and PMP22 do not share extensive homology; thus, duplications encompassing both loci are rare and potentially generated by a different mutational mechanism. We characterized genomic rearrangements that simultaneously duplicate PMP22 and RAI1, including nine potential complex genomic rearrangements, in 23 subjects by high-resolution array comparative genomic hybridization and breakpoint junction sequencing. Insertions and microhomologies were found at the breakpoint junctions, suggesting potential replicative mechanisms for rearrangement formation. At the breakpoint junctions of these nonrecurrent rearrangements, enrichment of repetitive DNA sequences was observed, indicating that they might predispose to genomic instability and rearrangement. Clinical evaluation revealed blended PTLS and CMT1A phenotypes with a potential earlier onset of neuropathy. Moreover, additional clinical findings might be observed due to the extra duplicated material included in the rearrangements. Our genomic analysis suggests replicative mechanisms as a predominant mechanism underlying PMP22-RAI1 contiguous gene duplications and provides further evidence supporting the role of complex genomic architecture in genomic instability.

}, keywords = {Abnormalities, Multiple, Charcot-Marie-Tooth Disease, Child, Child, Preschool, Chromosome Disorders, Chromosome Duplication, Chromosomes, Human, Pair 17, Comparative Genomic Hybridization, Female, Follow-Up Studies, Gene Duplication, Gene Rearrangement, Genome, Human, Genomics, Humans, Infant, Male, Models, Genetic, Myelin Proteins, Phenotype, Prognosis, Recombination, Genetic, Transcription Factors}, issn = {1537-6605}, doi = {10.1016/j.ajhg.2015.10.003}, author = {Yuan, Bo and Harel, Tamar and Gu, Shen and Liu, Pengfei and Burglen, Lydie and Chantot-Bastaraud, Sandra and Gelowani, Violet and Beck, Christine R and Carvalho, Claudia M B and Cheung, Sau Wai and Coe, Andrew and Malan, Val{\'e}rie and Munnich, Arnold and Magoulas, Pilar L and Potocki, Lorraine and Lupski, James R} } @article {285, title = {Exonic duplication CNV of NDRG1 associated with autosomal-recessive HMSN-Lom/CMT4D.}, journal = {Genet Med}, volume = {16}, year = {2014}, month = {2014 May}, pages = {386-394}, abstract = {

PURPOSE: Copy-number variations as a mutational mechanism contribute significantly to human disease. Approximately one-half of the patients with Charcot-Marie-Tooth (CMT) disease have a 1.4 Mb duplication copy-number variation as the cause of their neuropathy. However, non-CMT1A neuropathy patients rarely have causative copy-number variations, and to date, autosomal-recessive disease has not been associated with copy-number variation as a mutational mechanism.

METHODS: We performed Agilent 8 {\texttimes} 60 K array comparative genomic hybridization on DNA from 12 recessive Turkish families with CMT disease. Additional molecular studies were conducted to detect breakpoint junctions and to evaluate gene expression levels in a family in which we detected an intragenic duplication copy-number variation.

RESULTS: We detected an ~6.25 kb homozygous intragenic duplication in NDRG1, a gene known to be causative for recessive HMSNL/CMT4D, in three individuals from a Turkish family with CMT neuropathy. Further studies showed that this intragenic copy-number variation resulted in a homozygous duplication of exons 6-8 that caused decreased mRNA expression of NDRG1.

CONCLUSION: Exon-focused high-resolution array comparative genomic hybridization enables the detection of copy-number variation carrier states in recessive genes, particularly small copy-number variations encompassing or disrupting single genes. In families for whom a molecular diagnosis has not been elucidated by conventional clinical assays, an assessment for copy-number variations in known CMT genes might be considered.

}, keywords = {Adult, Base Sequence, Cell Cycle Proteins, Charcot-Marie-Tooth Disease, Comparative Genomic Hybridization, DNA Copy Number Variations, Female, Gene Duplication, Gene Expression, Genes, Recessive, Humans, Intracellular Signaling Peptides and Proteins, Male, Mutation, Refsum Disease, Sequence Analysis, DNA, Turkey, Young Adult}, issn = {1530-0366}, doi = {10.1038/gim.2013.155}, author = {Okamoto, Yuji and Goksungur, Meryem Tuba and Pehlivan, Davut and Beck, Christine R and Gonzaga-Jauregui, Claudia and Muzny, Donna M and Atik, Mehmed M and Carvalho, Claudia M B and Matur, Zeliha and Bayraktar, Serife and Boone, Philip M and Akyuz, Kaya and Gibbs, Richard A and Battaloglu, Esra and Parman, Yesim and Lupski, James R} } @article {245, title = {Structural variation and missense mutation in SBDS associated with Shwachman-Diamond syndrome.}, journal = {BMC Med Genet}, volume = {15}, year = {2014}, month = {2014 Jun 04}, pages = {64}, abstract = {

BACKGROUND: Shwachman-Diamond syndrome (SDS) is an autosomal recessive ribosomopathy caused mainly by compound heterozygous mutations in SBDS. Structural variation (SV) involving the SBDS locus has been rarely reported in association with the disease. We aimed to determine whether an SV contributed to the pathogenesis of a case lacking biallelic SBDS point mutations.

CASE PRESENTATION: Whole exome sequencing was performed in a patient with SDS lacking biallelic SBDS point mutations. Array comparative genomic hybridization and Southern blotting were used to seek SVs across the SBDS locus. Locus-specific polymerase chain reaction (PCR) encompassing flanking intronic sequence was also performed to investigate mutation within the locus. RNA expression and Western blotting were performed to analyze allele and protein expression. We found the child harbored a single missense mutation in SBDS (c.98A > C; p.K33T), inherited from the mother, and an SV in the SBDS locus, inherited from the father. The missense allele and SV segregated in accordance with Mendelian expectations for autosomal recessive SDS. Complementary DNA and western blotting analysis and locus specific PCR support the contention that the SV perturbed SBDS protein expression in the father and child.

CONCLUSION: Our findings implicate genomic rearrangements in the pathogenesis of some cases of SDS and support patients lacking biallelic SBDS point mutations be tested for SV within the SBDS locus.

}, keywords = {Abdomen, Alleles, Bone Marrow Diseases, Cell Line, Transformed, Child, Preschool, Comparative Genomic Hybridization, DNA Mutational Analysis, Exocrine Pancreatic Insufficiency, Female, Gene Order, Humans, Lipomatosis, Mutagenesis, Insertional, Mutation, Mutation, Missense, Pedigree, Proteins, Radiography, Abdominal, Ultrasonography}, issn = {1471-2350}, doi = {10.1186/1471-2350-15-64}, author = {Carvalho, Claudia M B and Zuccherato, Luciana W and Williams, Christopher L and Neill, Nicholas J and Murdock, David R and Bainbridge, Matthew and Jhangiani, Shalini N and Muzny, Donna M and Gibbs, Richard A and Ip, Wan and Guillerman, Robert Paul and Lupski, James R and Bertuch, Alison A} }