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1.  Matching two independent cohorts validates DPH1 as a gene responsible for autosomal recessive intellectual disability with short stature, craniofacial and ectodermal anomalies 
Human mutation  2015;36(10):1015-1019.
Recently, Alazami and colleagues identified 33 putative candidate disease genes for neurogenetic disorders. One such gene was DPH1, in which a homozygous missense mutation was associated with a 3C syndrome-like phenotype in four patients from a single extended family. Here we report a second homozygous missense variant in DPH1, seen in four members of a founder population, and associated with a phenotype initially reminiscent of Sensenbrenner syndrome. This post-publication ‘match’ validates DPH1 as a gene underlying syndromic intellectual disability with short stature and craniofacial and ectodermal anomalies, reminiscent of, but distinct from, 3C and Sensenbrenner syndromes. This validation took several years after the independent discoveries due to the absence of effective methods for sharing both candidate phenotype and genotype data between investigators. Sharing of data via web-based anonymous data exchange servers will play an increasingly important role towards more efficient identification of the molecular basis for rare Mendelian disorders.
doi:10.1002/humu.22843
PMCID: PMC4575268  PMID: 26220823
DPH1; Sensenbrenner; intellectual disability; rare disorders; Matchmaker Exchange
2.  GeneMatcher Aids in the Identification of a New Malformation Syndrome with Intellectual Disability, Unique Facial Dysmorphisms, and Skeletal and Connective Tissue Abnormalities Caused by De Novo Variants in HNRNPK 
Human mutation  2015;36(10):1009-1014.
We report a new syndrome due to loss of function variants in the heterogeneous nuclear ribonucleoprotein K gene (HNRNPK). We describe two probands, one with a de novo frameshift (NM_002140.3: c.953+1dup), and the other with a de novo splice donor site variant (NM_002140.3: c.257G>A). Both probands have intellectual disability, a shared unique craniofacial phenotype, and connective tissue and skeletal abnormalities. The identification of this syndrome was made possible by a new online tool, GeneMatcher, which facilitates connections between clinicians and researchers based on shared interest in candidate genes. This report demonstrates that new web based approaches can be effective in helping investigators solve exome sequencing projects, and also highlights the newer paradigm of “reverse phenotyping”, where characterization of syndromic features follows the identification of genetic variants.
doi:10.1002/humu.22837
PMCID: PMC4589226  PMID: 26173930
HNRNPK; reverse phenotyping; GeneMatcher; Matchmaker Exchange; WES
3.  An siRNA-based functional genomics screen for the identification of regulators of ciliogenesis and ciliopathy genes 
Wheway, Gabrielle | Schmidts, Miriam | Mans, Dorus A. | Szymanska, Katarzyna | Nguyen, Thanh-Minh T. | Racher, Hilary | Phelps, Ian G. | Toedt, Grischa | Kennedy, Julie | Wunderlich, Kirsten A. | Sorusch, Nasrin | Abdelhamed, Zakia A. | Natarajan, Subaashini | Herridge, Warren | van Reeuwijk, Jeroen | Horn, Nicola | Boldt, Karsten | Parry, David A. | Letteboer, Stef J.F. | Roosing, Susanne | Adams, Matthew | Bell, Sandra M. | Bond, Jacquelyn | Higgins, Julie | Morrison, Ewan E. | Tomlinson, Darren C. | Slaats, Gisela G. | van Dam, Teunis J. P. | Huang, Lijia | Kessler, Kristin | Giessl, Andreas | Logan, Clare V. | Boyle, Evan A. | Shendure, Jay | Anazi, Shamsa | Aldahmesh, Mohammed | Al Hazzaa, Selwa | Hegele, Robert A. | Ober, Carole | Frosk, Patrick | Mhanni, Aizeddin A. | Chodirker, Bernard N. | Chudley, Albert E. | Lamont, Ryan | Bernier, Francois P. | Beaulieu, Chandree L. | Gordon, Paul | Pon, Richard T. | Donahue, Clem | Barkovich, A. James | Wolf, Louis | Toomes, Carmel | Thiel, Christian T. | Boycott, Kym M. | McKibbin, Martin | Inglehearn, Chris F. | Stewart, Fiona | Omran, Heymut | Huynen, Martijn A. | Sergouniotis, Panagiotis I. | Alkuraya, Fowzan S. | Parboosingh, Jillian S. | Innes, A Micheil | Willoughby, Colin E. | Giles, Rachel H. | Webster, Andrew R. | Ueffing, Marius | Blacque, Oliver | Gleeson, Joseph G. | Wolfrum, Uwe | Beales, Philip L. | Gibson, Toby | Doherty, Dan | Mitchison, Hannah M. | Roepman, Ronald | Johnson, Colin A.
Nature cell biology  2015;17(8):1074-1087.
Defects in primary cilium biogenesis underlie the ciliopathies, a growing group of genetic disorders. We describe a whole genome siRNA-based reverse genetics screen for defects in biogenesis and/or maintenance of the primary cilium, obtaining a global resource. We identify 112 candidate ciliogenesis and ciliopathy genes, including 44 components of the ubiquitin-proteasome system, 12 G-protein-coupled receptors, and three pre-mRNA processing factors (PRPF6, PRPF8 and PRPF31) mutated in autosomal dominant retinitis pigmentosa. The PRPFs localise to the connecting cilium, and PRPF8- and PRPF31-mutated cells have ciliary defects. Combining the screen with exome sequencing data identified recessive mutations in PIBF1/CEP90 and C21orf2/LRRC76 as causes of the ciliopathies Joubert and Jeune syndromes. Biochemical approaches place C21orf2 within key ciliopathy-associated protein modules, offering an explanation for the skeletal and retinal involvement observed in individuals with C21orf2-variants. Our global, unbiased approaches provide insights into ciliogenesis complexity and identify roles for unanticipated pathways in human genetic disease.
doi:10.1038/ncb3201
PMCID: PMC4536769  PMID: 26167768
cilia; ciliopathies; reverse genetics; whole-genome siRNA screen; Jeune syndrome; Joubert syndrome
4.  Development of a diagnostic DNA chip to screen for 30 autosomal recessive disorders in the Hutterite population 
Abstract
Background
The Hutterites are a religious isolate living in colonies across the North American prairies. This population originated from approximately 90 founders, resulting in a number of genetic diseases that are overrepresented, underrepresented, or unique. The founder effect in this population increases the likelihood that Hutterite couples carry the same recessive mutations. We have designed a diagnostic chip on a fee‐for‐service basis with Asper Biotech to provide Hutterites with the option of comprehensive carrier screening.
Methods
A total of 32 disease‐causing mutations in 30 genes were selected and primers were designed for array primer extension‐based testing. Selected mutations were limited to those leading to autosomal recessive disorders, maintaining its primary use as a test for determining carrier status.
Results
The DNA chip was developed and validated using 59 DNA controls for all but one of the mutations, for which a synthetic control was used. All mutations were readily detected except for a duplication causing restrictive dermopathy where heterozygotes and homozygotes could only be distinguished by sequencing. Blinded testing of 12 additional samples from healthy Hutterites was performed by Asper Biotech using chip testing. All known mutations from previous molecular testing were detected on the chip. As well, additional mutations identified by the chip in these 12 samples were subsequently verified by a second method.
Conclusions
Our analysis indicates that the chip is a sensitive and specific means of carrier testing in the Hutterite population and can serve as a model for other founder populations.
doi:10.1002/mgg3.206
PMCID: PMC4867565  PMID: 27247959
chip; Hutterite; diagnostic; mutation; APEX array; carrier screening
5.  An Algorithm Measuring Donor Cell-Free DNA in Plasma of Cellular and Solid Organ Transplant Recipients That Does Not Require Donor or Recipient Genotyping 
Cell-free DNA (cfDNA) has significant potential in the diagnosis and monitoring of clinical conditions. However, accurately and easily distinguishing the relative proportion of DNA molecules in a mixture derived from two different sources (i.e., donor and recipient tissues after transplantation) is challenging. In human cellular transplantation, there is currently no useable method to detect in vivo engraftment, and blood-based non-invasive tests for allograft rejection in solid organ transplantation are either non-specific or absent. Elevated levels of donor cfDNA have been shown to correlate with solid organ rejection, but complex methodology limits implementation of this promising biomarker. We describe a cost-effective method to quantify donor cfDNA in recipient plasma using a panel of high-frequency single nucleotide polymorphisms, next-generation (semiconductor) sequencing, and a novel mixture model algorithm. In vitro, our method accurately and rapidly determined donor:recipient DNA admixture. For in vivo testing, donor cfDNA was serially quantified in an infant with a urea cycle disorder after receiving six daily infusions of donor liver cells. Donor cfDNA isolated from 1 to 2 ml of recipient plasma was detected as late as 24 weeks after infusion suggesting engraftment. The percentage of circulating donor cfDNA was also assessed in pediatric and adult heart transplant recipients undergoing routine endomyocardial biopsy with levels observed to be stable over time and generally measuring <1% in cases without moderate or severe cellular rejection. Unlike existing non-invasive methods used to define the proportion of donor cfDNA in solid organ transplant patients, our assay does not require sex mismatch, donor genotyping, or whole-genome sequencing and potentially has broad application to detect cellular engraftment or allograft injury after transplantation.
doi:10.3389/fcvm.2016.00033
PMCID: PMC5031701  PMID: 27713880
cell-free DNA; transplantation; hepatocyte transplantation; heart; solid organ transplantation; biomarker
6.  A Shared Founder Mutation Underlies Restrictive Dermopathy in Old Colony (Dutch-German) Mennonite and Hutterite Patients in North America 
doi:10.1002/ajmg.a.35302
PMCID: PMC4247856  PMID: 22495976
restrictive dermopathy; tight skin contracture syndrome; laminopathy; lethal; Hutterite; Mennonite; ZMPSTE24
7.  Disrupted auto-regulation of the spliceosomal gene SNRPB causes cerebro–costo–mandibular syndrome 
Nature Communications  2014;5:4483.
Elucidating the function of highly conserved regulatory sequences is a significant challenge in genomics today. Certain intragenic highly conserved elements have been associated with regulating levels of core components of the spliceosome and alternative splicing of downstream genes. Here we identify mutations in one such element, a regulatory alternative exon of SNRPB as the cause of cerebro–costo–mandibular syndrome. This exon contains a premature termination codon that triggers nonsense-mediated mRNA decay when included in the transcript. These mutations cause increased inclusion of the alternative exon and decreased overall expression of SNRPB. We provide evidence for the functional importance of this conserved intragenic element in the regulation of alternative splicing and development, and suggest that the evolution of such a regulatory mechanism has contributed to the complexity of mammalian development.
Cerebro–costo–mandibular syndrome, CCMS, is a severe human multiple malformation disorder. Here, the authors report that mutations in SNRPB disrupt the normal regulation of alternative splicing at this gene, and in so doing, may be responsible for the development of CCMS.
doi:10.1038/ncomms5483
PMCID: PMC4109005  PMID: 25047197
8.  An international effort towards developing standards for best practices in analysis, interpretation and reporting of clinical genome sequencing results in the CLARITY Challenge 
Brownstein, Catherine A | Beggs, Alan H | Homer, Nils | Merriman, Barry | Yu, Timothy W | Flannery, Katherine C | DeChene, Elizabeth T | Towne, Meghan C | Savage, Sarah K | Price, Emily N | Holm, Ingrid A | Luquette, Lovelace J | Lyon, Elaine | Majzoub, Joseph | Neupert, Peter | McCallie Jr, David | Szolovits, Peter | Willard, Huntington F | Mendelsohn, Nancy J | Temme, Renee | Finkel, Richard S | Yum, Sabrina W | Medne, Livija | Sunyaev, Shamil R | Adzhubey, Ivan | Cassa, Christopher A | de Bakker, Paul IW | Duzkale, Hatice | Dworzyński, Piotr | Fairbrother, William | Francioli, Laurent | Funke, Birgit H | Giovanni, Monica A | Handsaker, Robert E | Lage, Kasper | Lebo, Matthew S | Lek, Monkol | Leshchiner, Ignaty | MacArthur, Daniel G | McLaughlin, Heather M | Murray, Michael F | Pers, Tune H | Polak, Paz P | Raychaudhuri, Soumya | Rehm, Heidi L | Soemedi, Rachel | Stitziel, Nathan O | Vestecka, Sara | Supper, Jochen | Gugenmus, Claudia | Klocke, Bernward | Hahn, Alexander | Schubach, Max | Menzel, Mortiz | Biskup, Saskia | Freisinger, Peter | Deng, Mario | Braun, Martin | Perner, Sven | Smith, Richard JH | Andorf, Janeen L | Huang, Jian | Ryckman, Kelli | Sheffield, Val C | Stone, Edwin M | Bair, Thomas | Black-Ziegelbein, E Ann | Braun, Terry A | Darbro, Benjamin | DeLuca, Adam P | Kolbe, Diana L | Scheetz, Todd E | Shearer, Aiden E | Sompallae, Rama | Wang, Kai | Bassuk, Alexander G | Edens, Erik | Mathews, Katherine | Moore, Steven A | Shchelochkov, Oleg A | Trapane, Pamela | Bossler, Aaron | Campbell, Colleen A | Heusel, Jonathan W | Kwitek, Anne | Maga, Tara | Panzer, Karin | Wassink, Thomas | Van Daele, Douglas | Azaiez, Hela | Booth, Kevin | Meyer, Nic | Segal, Michael M | Williams, Marc S | Tromp, Gerard | White, Peter | Corsmeier, Donald | Fitzgerald-Butt, Sara | Herman, Gail | Lamb-Thrush, Devon | McBride, Kim L | Newsom, David | Pierson, Christopher R | Rakowsky, Alexander T | Maver, Aleš | Lovrečić, Luca | Palandačić, Anja | Peterlin, Borut | Torkamani, Ali | Wedell, Anna | Huss, Mikael | Alexeyenko, Andrey | Lindvall, Jessica M | Magnusson, Måns | Nilsson, Daniel | Stranneheim, Henrik | Taylan, Fulya | Gilissen, Christian | Hoischen, Alexander | van Bon, Bregje | Yntema, Helger | Nelen, Marcel | Zhang, Weidong | Sager, Jason | Zhang, Lu | Blair, Kathryn | Kural, Deniz | Cariaso, Michael | Lennon, Greg G | Javed, Asif | Agrawal, Saloni | Ng, Pauline C | Sandhu, Komal S | Krishna, Shuba | Veeramachaneni, Vamsi | Isakov, Ofer | Halperin, Eran | Friedman, Eitan | Shomron, Noam | Glusman, Gustavo | Roach, Jared C | Caballero, Juan | Cox, Hannah C | Mauldin, Denise | Ament, Seth A | Rowen, Lee | Richards, Daniel R | Lucas, F Anthony San | Gonzalez-Garay, Manuel L | Caskey, C Thomas | Bai, Yu | Huang, Ying | Fang, Fang | Zhang, Yan | Wang, Zhengyuan | Barrera, Jorge | Garcia-Lobo, Juan M | González-Lamuño, Domingo | Llorca, Javier | Rodriguez, Maria C | Varela, Ignacio | Reese, Martin G | De La Vega, Francisco M | Kiruluta, Edward | Cargill, Michele | Hart, Reece K | Sorenson, Jon M | Lyon, Gholson J | Stevenson, David A | Bray, Bruce E | Moore, Barry M | Eilbeck, Karen | Yandell, Mark | Zhao, Hongyu | Hou, Lin | Chen, Xiaowei | Yan, Xiting | Chen, Mengjie | Li, Cong | Yang, Can | Gunel, Murat | Li, Peining | Kong, Yong | Alexander, Austin C | Albertyn, Zayed I | Boycott, Kym M | Bulman, Dennis E | Gordon, Paul MK | Innes, A Micheil | Knoppers, Bartha M | Majewski, Jacek | Marshall, Christian R | Parboosingh, Jillian S | Sawyer, Sarah L | Samuels, Mark E | Schwartzentruber, Jeremy | Kohane, Isaac S | Margulies, David M
Genome Biology  2014;15(3):R53.
Background
There is tremendous potential for genome sequencing to improve clinical diagnosis and care once it becomes routinely accessible, but this will require formalizing research methods into clinical best practices in the areas of sequence data generation, analysis, interpretation and reporting. The CLARITY Challenge was designed to spur convergence in methods for diagnosing genetic disease starting from clinical case history and genome sequencing data. DNA samples were obtained from three families with heritable genetic disorders and genomic sequence data were donated by sequencing platform vendors. The challenge was to analyze and interpret these data with the goals of identifying disease-causing variants and reporting the findings in a clinically useful format. Participating contestant groups were solicited broadly, and an independent panel of judges evaluated their performance.
Results
A total of 30 international groups were engaged. The entries reveal a general convergence of practices on most elements of the analysis and interpretation process. However, even given this commonality of approach, only two groups identified the consensus candidate variants in all disease cases, demonstrating a need for consistent fine-tuning of the generally accepted methods. There was greater diversity of the final clinical report content and in the patient consenting process, demonstrating that these areas require additional exploration and standardization.
Conclusions
The CLARITY Challenge provides a comprehensive assessment of current practices for using genome sequencing to diagnose and report genetic diseases. There is remarkable convergence in bioinformatic techniques, but medical interpretation and reporting are areas that require further development by many groups.
doi:10.1186/gb-2014-15-3-r53
PMCID: PMC4073084  PMID: 24667040
9.  Intellectual disability associated with a homozygous missense mutation in THOC6 
Background
We recently described a novel autosomal recessive neurodevelopmental disorder with intellectual disability in four patients from two related Hutterite families. Identity-by-descent mapping localized the gene to a 5.1 Mb region at chromosome 16p13.3 containing more than 170 known or predicted genes. The objective of this study was to identify the causative gene for this rare disorder.
Methods and results
Candidate gene sequencing followed by exome sequencing identified a homozygous missense mutation p.Gly46Arg, in THOC6. No other potentially causative coding variants were present within the critical region on chromosome 16. THOC6 is a member of the THO/TREX complex which is involved in coordinating mRNA processing with mRNA export from the nucleus. In situ hybridization showed that thoc6 is highly expressed in the midbrain and eyes. Cellular localization studies demonstrated that wild-type THOC6 is present within the nucleus as is the case for other THO complex proteins. However, mutant THOC6 was predominantly localized to the cytoplasm, suggesting that the mutant protein is unable to carry out its normal function. siRNA knockdown of THOC6 revealed increased apoptosis in cultured cells.
Conclusion
Our findings associate a missense mutation in THOC6 with intellectual disability, suggesting the THO/TREX complex plays an important role in neurodevelopment.
doi:10.1186/1750-1172-8-62
PMCID: PMC3644499  PMID: 23621916
Intellectual disability; THOC6; THO/TREX complex; mRNA export; Hutterite
11.  Mutations in VLDLR as a Cause for Autosomal Recessive Cerebellar Ataxia with Mental Retardation (Dysequilibrium Syndrome) 
Journal of child neurology  2009;24(10):1310-1315.
Dysequilibrium syndrome (DES) is a genetically heterogeneous condition that combines autosomal recessive, non-progressive cerebellar ataxia with mental retardation. Here we report the first patient heterozygous for two novel mutations in VLDLR. An 18-month old girl presented with significant hypotonia, global developmental delay, and truncal and peripheral ataxia. MR imaging of the brain demonstrated hypoplasia of the inferior cerebellar vermis and hemispheres, small pons, and a simplified cortical sulcation pattern. Sequence analysis of the VLDLR gene identified a nonsense and missense mutation. Six mutations in VLDLR have now been identified in five families with a phenotype characterized by moderate-to-profound mental retardation, delayed ambulation, truncal and peripheral ataxia and occasional seizures. Neuroanatomically, the loss-of-function effect of the different mutations is indistinguishable. VLDLR-associated cerebellar hypoplasia is emerging as a panethnic, clinically and molecularly well-defined genetic syndrome.
doi:10.1177/0883073809332696
PMCID: PMC2849979  PMID: 19332571
VLDLR; Cerebellar hypoplasia; Dysequilibrium syndrome
12.  Evaluation of BRCA1 and BRCA2 mutation prevalence, risk prediction models and a multistep testing approach in French‐Canadian families with high risk of breast and ovarian cancer 
Journal of Medical Genetics  2006;44(2):107-121.
Background and objective
In clinical settings with fixed resources allocated to predictive genetic testing for high‐risk cancer predisposition genes, optimal strategies for mutation screening programmes are critically important. These depend on the mutation spectrum found in the population under consideration and the frequency of mutations detected as a function of the personal and family history of cancer, which are both affected by the presence of founder mutations and demographic characteristics of the underlying population. The results of multistep genetic testing for mutations in BRCA1 or BRCA2 in a large series of families with breast cancer in the French‐Canadian population of Quebec, Canada are reported.
Methods
A total of 256 high‐risk families were ascertained from regional familial cancer clinics throughout the province of Quebec. Initially, families were tested for a panel of specific mutations known to occur in this population. Families in which no mutation was identified were then comprehensively tested. Three algorithms to predict the presence of mutations were evaluated, including the prevalence tables provided by Myriad Genetics Laboratories, the Manchester Scoring System and a logistic regression approach based on the data from this study.
Results
8 of the 15 distinct mutations found in 62 BRCA1/BRCA2‐positive families had never been previously reported in this population, whereas 82% carried 1 of the 4 mutations currently observed in ⩾2 families. In the subset of 191 families in which at least 1 affected individual was tested, 29% carried a mutation. Of these 27 BRCA1‐positive and 29 BRCA2‐positive families, 48 (86%) were found to harbour a mutation detected by the initial test. Among the remaining 143 inconclusive families, all 8 families found to have a mutation after complete sequencing had Manchester Scores ⩾18. The logistic regression and Manchester Scores provided equal predictive power, and both were significantly better than the Myriad Genetics Laboratories prevalence tables (p<0.001). A threshold of Manchester Score ⩾18 provided an overall sensitivity of 86% and a specificity of 82%, with a positive predictive value of 66% in this population.
Conclusion
In this population, a testing strategy with an initial test using a panel of reported recurrent mutations, followed by full sequencing in families with Manchester Scores ⩾18, represents an efficient test in terms of overall cost and sensitivity.
doi:10.1136/jmg.2006.044388
PMCID: PMC2598057  PMID: 16905680

Results 1-13 (13)