Publications by authors named "Sanjeev S Bhaskar"

24 Publications

  • Page 1 of 1

A deep intronic SMARCB1 variant associated with schwannomatosis.

Clin Genet 2020 02 9;97(2):376-377. Epub 2019 Sep 9.

NW Genomic Laboratory Hub, Manchester Centre for Genomic Medicine, St Mary's Hospital, Manchester University Hospitals NHS Foundation Trust, Manchester, UK.

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http://dx.doi.org/10.1111/cge.13637DOI Listing
February 2020

A homozygous missense variant in CHRM3 associated with familial urinary bladder disease.

Clin Genet 2019 12 11;96(6):515-520. Epub 2019 Sep 11.

Manchester Centre for Genomic Medicine, Manchester University NHS Foundation Trust, Health Innovation Manchester, Manchester, UK.

CHRM3 codes for the M3 muscarinic acetylcholine receptor that is located on the surface of smooth muscle cells of the detrusor, the muscle that effects urinary voiding. Previously, we reported brothers in a family affected by a congenital prune belly-like syndrome with mydriasis due to homozygous CHRM3 frameshift variants. In this study, we describe two sisters with bladders that failed to empty completely and pupils that failed to constrict fully in response to light, who are homozygous for the missense CHRM3 variant c.352G > A; p.(Gly118Arg). Samples were not available for genotyping from their brother, who had a history of multiple urinary tract infections and underwent surgical bladder draining in the first year of life. He died at the age of 6 years. This is the first independent report of biallelic variants in CHRM3 in a family with a rare serious bladder disorder associated with mydriasis and provides important evidence of this association.
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http://dx.doi.org/10.1111/cge.13631DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6899476PMC
December 2019

Whole Exome Sequencing Reveals the Major Genetic Contributors to Nonsyndromic Tetralogy of Fallot.

Circ Res 2019 02;124(4):553-563

From the Division of Cardiovascular Sciences, School of Medical Sciences, Faculty of Biology, Medicine, and Health, Manchester Academic Health Science Centre, University of Manchester, United Kingdom (D.J.P., S.G.W., R.M.M., E.F., B.D.K.).

Rationale: Familial recurrence studies provide strong evidence for a genetic component to the predisposition to sporadic, nonsyndromic Tetralogy of Fallot (TOF), the most common cyanotic congenital heart disease phenotype. Rare genetic variants have been identified as important contributors to the risk of congenital heart disease, but relatively small numbers of TOF cases have been studied to date.

Objective: We used whole exome sequencing to assess the prevalence of unique, deleterious variants in the largest cohort of nonsyndromic TOF patients reported to date.

Methods And Results: Eight hundred twenty-nine TOF patients underwent whole exome sequencing. The presence of unique, deleterious variants was determined; defined by their absence in the Genome Aggregation Database and a scaled combined annotation-dependent depletion score of ≥20. The clustering of variants in 2 genes, NOTCH1 and FLT4, surpassed thresholds for genome-wide significance (assigned as P<5×10) after correction for multiple comparisons. NOTCH1 was most frequently found to harbor unique, deleterious variants. Thirty-one changes were observed in 37 probands (4.5%; 95% CI, 3.2%-6.1%) and included 7 loss-of-function variants 22 missense variants and 2 in-frame indels. Sanger sequencing of the unaffected parents of 7 cases identified 5 de novo variants. Three NOTCH1 variants (p.G200R, p.C607Y, and p.N1875S) were subjected to functional evaluation, and 2 showed a reduction in Jagged1-induced NOTCH signaling. FLT4 variants were found in 2.4% (95% CI, 1.6%-3.8%) of TOF patients, with 21 patients harboring 22 unique, deleterious variants. The variants identified were distinct to those that cause the congenital lymphoedema syndrome Milroy disease. In addition to NOTCH1, FLT4 and the well-established TOF gene, TBX1, we identified potential association with variants in several other candidates, including RYR1, ZFPM1, CAMTA2, DLX6, and PCM1.

Conclusions: The NOTCH1 locus is the most frequent site of genetic variants predisposing to nonsyndromic TOF, followed by FLT4. Together, variants in these genes are found in almost 7% of TOF patients.
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http://dx.doi.org/10.1161/CIRCRESAHA.118.313250DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6377791PMC
February 2019

Diagnosing childhood-onset inborn errors of metabolism by next-generation sequencing.

Arch Dis Child 2017 11 3;102(11):1019-1029. Epub 2017 May 3.

Manchester Centre for Genomic Medicine, St. Mary's Hospital, Central Manchester NHS Trust, Manchester Academic Health Science Centre, Manchester, UK.

Background: Inborn errors of metabolism (IEMs) underlie a substantial proportion of paediatric disease burden but their genetic diagnosis can be challenging using the traditional approaches.

Methods: We designed and validated a next-generation sequencing (NGS) panel of 226 IEM genes, created six overlapping phenotype-based subpanels and tested 102 individuals, who presented clinically with suspected childhood-onset IEMs.

Results: In 51/102 individuals, NGS fully or partially established the molecular cause or identified other actionable diagnoses. Causal mutations were identified significantly more frequently when the biochemical phenotype suggested a specific IEM or a group of IEMs (p<0.0001), demonstrating the pivotal role of prior biochemical testing in guiding NGS analysis. The NGS panel helped to avoid further invasive, hazardous, lengthy or expensive investigations in 69% individuals (p<0.0001). Additional functional testing due to novel or unexpected findings had to be undertaken in only 3% of subjects, demonstrating that the use of NGS does not significantly increase the burden of subsequent follow-up testing. Even where a molecular diagnosis could not be achieved, NGS-based approach assisted in the management and counselling by reducing the likelihood of a high-penetrant genetic cause.

Conclusion: NGS has significant clinical utility for the diagnosis of IEMs. Biochemical testing and NGS analysis play complementary roles in the diagnosis of IEMs. Incorporating NGS into the diagnostic algorithm of IEMs can improve the accuracy of diagnosis.
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http://dx.doi.org/10.1136/archdischild-2017-312738DOI Listing
November 2017

Homozygous mutation in PTRH2 gene causes progressive sensorineural deafness and peripheral neuropathy.

Am J Med Genet A 2017 Apr;173(4):1051-1055

Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.

PTRH2 is an evolutionarily highly conserved mitochondrial protein that belongs to a family of peptidyl-tRNA hydrolases. Recently, patients from two consanguineous families with mutations in the PTRH2 gene were reported. Global developmental delay associated with microcephaly, growth retardation, progressive ataxia, distal muscle weakness with ankle contractures, demyelinating sensorimotor neuropathy, and sensorineural hearing loss were present in all patients, while facial dysmorphism with widely spaced eyes, exotropia, thin upper lip, proximally placed thumbs, and deformities of the fingers and toes were present in some individuals. Here, we report a new family with three siblings affected by sensorineural hearing loss and peripheral neuropathy. Autozygosity mapping followed by exome sequencing identified a previously reported homozygous missense mutation in PTRH2 (c.254A>C; p.(Gln85Pro)). Sanger sequencing confirmed that the variant segregated with the phenotype. In contrast to the previously reported patient, the affected siblings had normal intelligence, milder microcephaly, delayed puberty, myopia, and moderate insensitivity to pain. Our findings expand the clinical phenotype and further demonstrate the clinical heterogeneity related to PTRH2 variants.
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http://dx.doi.org/10.1002/ajmg.a.38140DOI Listing
April 2017

The role of small in-frame insertions/deletions in inherited eye disorders and how structural modelling can help estimate their pathogenicity.

Orphanet J Rare Dis 2016 09 14;11(1):125. Epub 2016 Sep 14.

School of Biological Science, University of Manchester, Manchester, UK.

Background: Although the majority of small in-frame insertions/deletions (indels) has no/little affect on protein function, a small subset of these changes has been causally associated with genetic disorders. Notably, the molecular mechanisms and frequency by which they give rise to disease phenotypes remain largely unknown. The aim of this study is to provide insights into the role of in-frame indels (≤21 nucleotides) in two genetically heterogeneous eye disorders.

Results: One hundred eighty-one probands with childhood cataracts and 486 probands with retinal dystrophy underwent multigene panel testing in a clinical diagnostic laboratory. In-frame indels were collected and evaluated both clinically and in silico. Variants that could be modeled in the context of protein structure were identified and analysed using integrative structural modeling. Overall, 55 small in-frame indels were detected in 112 of 667 probands (16.8 %); 17 of these changes were novel to this study and 18 variants were reported clinically. A reliable model of the corresponding protein sequence could be generated for 8 variants. Structural modeling indicated a diverse range of molecular mechanisms of disease including disruption of secondary and tertiary protein structure and alteration of protein-DNA binding sites.

Conclusions: In childhood cataract and retinal dystrophy subjects, one small in-frame indel is clinically reported in every ~37 individuals tested. The clinical utility of computational tools evaluating these changes increases when the full complexity of the involved molecular mechanisms is embraced.
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http://dx.doi.org/10.1186/s13023-016-0505-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5024463PMC
September 2016

Mutations in SNORD118 cause the cerebral microangiopathy leukoencephalopathy with calcifications and cysts.

Nat Genet 2016 10 29;48(10):1185-92. Epub 2016 Aug 29.

Faculty of Biology, Medicine and Health, School of Biological Sciences, Division of Evolution and Genomic Sciences, University of Manchester, Manchester, UK.

Although ribosomes are ubiquitous and essential for life, recent data indicate that monogenic causes of ribosomal dysfunction can confer a remarkable degree of specificity in terms of human disease phenotype. Box C/D small nucleolar RNAs (snoRNAs) are evolutionarily conserved non-protein-coding RNAs involved in ribosome biogenesis. Here we show that biallelic mutations in the gene SNORD118, encoding the box C/D snoRNA U8, cause the cerebral microangiopathy leukoencephalopathy with calcifications and cysts (LCC), presenting at any age from early childhood to late adulthood. These mutations affect U8 expression, processing and protein binding and thus implicate U8 as essential in cerebral vascular homeostasis.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5045717PMC
http://dx.doi.org/10.1038/ng.3661DOI Listing
October 2016

Deletion of 19q13 reveals clinical overlap with Dubowitz syndrome.

J Hum Genet 2015 Dec 17;60(12):781-5. Epub 2015 Sep 17.

Manchester Centre for Genomic Medicine, Institute of Human Development, Faculty of Medical and Human Sciences, University of Manchester, Manchester Academic Health Science Centre (MAHSC), Manchester, UK.

Dubowitz syndrome is a presumed autosomal recessive disorder characterized by multiple congenital abnormalities: microcephaly, learning and developmental delay, growth failure, and a predisposition to allergies and eczema. There have been more than 150 individuals reported to have this diagnosis, but no unifying genetic alteration has been identified indicating genetic heterogeneity. We report on a pair of monozygotic twins diagnosed clinically with Dubowitz syndrome by Professor Dubowitz over 30 years ago and identified to have a de novo heterozygous 3.2-Mb deletion at 19q13.11q13.12. Exome sequencing did not identify either a putative pathogenic variant on the trans allele supporting recessive inheritance or any other causative sequence variants. Comparison of the phenotype in our cases shows considerable overlap with the 19q13.11 microdeletion syndrome, suggesting that a subset of individuals diagnosed with Dubowitz syndrome may be due to deletions at 19q13. Our finding further reinforces the genetic and phenotypic heterogeneity of Dubowitz syndrome.
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http://dx.doi.org/10.1038/jhg.2015.111DOI Listing
December 2015

Agnathia-otocephaly complex and asymmetric velopharyngeal insufficiency due to an in-frame duplication in OTX2.

J Hum Genet 2015 Apr 15;60(4):199-202. Epub 2015 Jan 15.

1] Institute of Human Development, University of Manchester, Manchester, UK [2] Manchester Centre for Genomic Medicine, St Mary's Hospital, Central Manchester University Hospitals NHS Foundation Trust, Manchester, UK.

Agnathia-otocephaly complex is a malformation characterized by absent/hypoplastic mandible and abnormally positioned ears. Mutations in two genes, PRRX1 and OTX2, have been described in a small number of families with this disorder. We performed clinical and genetic testing in an additional family. The proband is a healthy female with a complicated pregnancy history that includes two offspring diagnosed with agnathia-otocephaly during prenatal ultrasound scans. Exome sequencing was performed in fetal DNA from one of these two offspring revealing a heterozygous duplication in OTX2: c.271_273dupCAG, p.(Gln91dup). This change leads to the insertion of a glutamine within the OTX2 homeodomain region, and is predicted to alter this signaling molecule's ability to interact with DNA. The same variant was also identified in the proband's clinically unaffected 38-year-old husband and their 9-year-old daughter, who presented with a small mandible, normal ears and velopharyngeal insufficiency due to a short hemi-palate. This unusual presentation of OTX2-related disease suggests that OTX2 might have a role in palatal hypoplasia cases. A previously unreported OTX2 variant associated with extreme intrafamilial variability is described and the utility of exome sequencing as a tool to confirm the diagnosis of agnathia-otocephaly and to inform the reproductive decisions of affected families is highlighted.
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http://dx.doi.org/10.1038/jhg.2014.122DOI Listing
April 2015

Mutations in LZTR1 add to the complex heterogeneity of schwannomatosis.

Neurology 2015 Jan 5;84(2):141-7. Epub 2014 Dec 5.

From the Manchester Centre for Genomic Medicine (M.J.S., W.G.N., D.G.E.) and University of Manchester Biomedical Imaging Institute (S.J.M.), Manchester Academic Health Sciences Centre, and Centre for Imaging Sciences (S.J.M.), University of Manchester, UK; Service de Dermatologie (S.B.) and Service de Genetique Medicale (B.I.), CHU Nantes, France; Institut für Klinische Chemie und Laboratoriumsdiagnostik Universitätsklinikum Jena (C.B.), Germany; Centre for Genomic Medicine (S.G.W., S.S.B., J.O., B.A., S.B.D., J.E.U., W.G.N., D.G.E.), St. Mary's Hospital, Central Manchester University Hospitals NHS Foundation Trust, UK; INSERM U830 (W.R., F.B.), Laboratoire de Genetique et Biologie des Cancers, Paris, France; Department of Clinical Genetics (A.F.), Alder Hey Children's Hospital, Liverpool, UK; Department of Medical Genetics (C.F.R.), Oslo University Hospital, Norway; International Neuroscience Institute (A.S.), Hannover, Germany; Department of Cellular Pathology and Greater Manchester Neurosciences Centre (D.d.P.), Salford Royal Hospitals NHS Foundation Trust; Department of Clinical Genetics (D.H.), Oxford Radcliffe Hospitals NHS Trust, UK; and Institut Curie (F.B.), SIRIC and Departement d'Oncologie Pediatrique d'Adolescents et Jeunes Adultes, Paris, France.

Objectives: We aimed to determine the proportion of individuals in our schwannomatosis cohort whose disease is associated with an LZTR1 mutation.

Methods: We used exome sequencing, Sanger sequencing, and copy number analysis to screen 65 unrelated individuals with schwannomatosis who were negative for a germline NF2 or SMARCB1 mutation. We also screened samples from 39 patients with a unilateral vestibular schwannoma (UVS), plus at least one other schwannoma, but who did not have an identifiable germline or mosaic NF2 mutation.

Results: We identified germline LZTR1 mutations in 6 of 16 patients (37.5%) with schwannomatosis who had at least one affected relative, 11 of 49 (22%) sporadic patients, and 2 of 39 patients with UVS in our cohort. Three germline mutation-positive patients in total had developed a UVS. Mosaicism was excluded in 3 patients without germline mutation in NF2, SMARCB1, or LZTR1 by mutation screening in 2 tumors from each.

Conclusions: Our data confirm the relationship between mutations in LZTR1 and schwannomatosis. They indicate that germline mutations in LZTR1 confer an increased risk of vestibular schwannoma, providing further overlap with NF2, and that further causative genes for schwannomatosis remain to be identified.
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http://dx.doi.org/10.1212/WNL.0000000000001129DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4336087PMC
January 2015

Compound heterozygosity of low-frequency promoter deletions and rare loss-of-function mutations in TXNL4A causes Burn-McKeown syndrome.

Am J Hum Genet 2014 Dec 26;95(6):698-707. Epub 2014 Nov 26.

Institute of Human Genetics, Helmholtz Zentrum München, 85764 Neuherberg, Germany; Institut für Praenatale Medizin & Humangenetik, 42103 Wuppertal, Germany.

Mutations in components of the major spliceosome have been described in disorders with craniofacial anomalies, e.g., Nager syndrome and mandibulofacial dysostosis type Guion-Almeida. The U5 spliceosomal complex of eight highly conserved proteins is critical for pre-mRNA splicing. We identified biallelic mutations in TXNL4A, a member of this complex, in individuals with Burn-McKeown syndrome (BMKS). This rare condition is characterized by bilateral choanal atresia, hearing loss, cleft lip and/or palate, and other craniofacial dysmorphisms. Mutations were found in 9 of 11 affected families. In 8 families, affected individuals carried a rare loss-of-function mutation (nonsense, frameshift, or microdeletion) on one allele and a low-frequency 34 bp deletion (allele frequency 0.76%) in the core promoter region on the other allele. In a single highly consanguineous family, formerly diagnosed as oculo-oto-facial dysplasia, the four affected individuals were homozygous for a 34 bp promoter deletion, which differed from the promoter deletion in the other families. Reporter gene and in vivo assays showed that the promoter deletions led to reduced expression of TXNL4A. Depletion of TXNL4A (Dib1) in yeast demonstrated reduced assembly of the tri-snRNP complex. Our results indicate that BMKS is an autosomal-recessive condition, which is frequently caused by compound heterozygosity of low-frequency promoter deletions in combination with very rare loss-of-function mutations.
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http://dx.doi.org/10.1016/j.ajhg.2014.10.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4259969PMC
December 2014

Germline mutations in SUFU cause Gorlin syndrome-associated childhood medulloblastoma and redefine the risk associated with PTCH1 mutations.

J Clin Oncol 2014 Dec 17;32(36):4155-61. Epub 2014 Nov 17.

Miriam J. Smith, Simon G. Williams, Sanjeev S. Bhaskar, James O'Sullivan, Beverley Anderson, Sarah B. Daly, Jill E. Urquhart, Zaynab Bholah, William G. Newman, and D. Gareth R. Evans, Manchester Centre for Genomic Medicine, University of Manchester, Manchester Academic Health Sciences Centre, and Central Manchester University Hospitals National Health Service (NHS) Foundation Trust; Deemesh Oudit, Christie NHS Foundation Trust; Edmund Cheesman and Anna Kelsey, Central Manchester University Hospital NHS Foundation Trust, Royal Manchester Children's Hospital; Martin G. McCabe, Institute of Cancer Sciences, University of Manchester, Manchester Academic Health Science Centre, Manchester, United Kingdom; and Christian Beetz, Institut für Klinische Chemie und Laboratoriumsdiagnostik Universitätsklinikum Jena, Jena, Germany.

Purpose: Heterozygous germline PTCH1 mutations are causative of Gorlin syndrome (naevoid basal cell carcinoma), but detection rates > 70% have rarely been reported. We aimed to define the causative mutations in individuals with Gorlin syndrome without PTCH1 mutations.

Methods: We undertook exome sequencing on lymphocyte DNA from four unrelated individuals from families with Gorlin syndrome with no PTCH1 mutations found by Sanger sequencing, multiplex ligation-dependent probe amplification (MLPA), or RNA analysis.

Results: A germline heterozygous nonsense mutation in SUFU was identified in one of four exomes. Sanger sequencing of SUFU in 23 additional PTCH1-negative Gorlin syndrome families identified a SUFU mutation in a second family. Copy-number analysis of SUFU by MLPA revealed a large heterozygous deletion in a third family. All three SUFU-positive families fulfilled diagnostic criteria for Gorlin syndrome, although none had odontogenic jaw keratocysts. Each SUFU-positive family included a single case of medulloblastoma, whereas only two (1.7%) of 115 individuals with Gorlin syndrome and a PTCH1 mutation developed medulloblastoma.

Conclusion: We demonstrate convincing evidence that SUFU mutations can cause classical Gorlin syndrome. Our study redefines the risk of medulloblastoma in Gorlin syndrome, dependent on the underlying causative gene. Previous reports have found a 5% risk of medulloblastoma in Gorlin syndrome. We found a < 2% risk in PTCH1 mutation-positive individuals, with a risk up to 20× higher in SUFU mutation-positive individuals. Our data suggest childhood brain magnetic resonance imaging surveillance is justified in SUFU-related, but not PTCH1-related, Gorlin syndrome.
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http://dx.doi.org/10.1200/JCO.2014.58.2569DOI Listing
December 2014

The genetic basis of DOORS syndrome: an exome-sequencing study.

Lancet Neurol 2014 Jan 29;13(1):44-58. Epub 2013 Nov 29.

Department of Clinical and Experimental Epilepsy, UCL Institute of Neurology, London, UK; Epilepsy Society, Buckinghamshire, UK. Electronic address:

Background: Deafness, onychodystrophy, osteodystrophy, mental retardation, and seizures (DOORS) syndrome is a rare autosomal recessive disorder of unknown cause. We aimed to identify the genetic basis of this syndrome by sequencing most coding exons in affected individuals.

Methods: Through a search of available case studies and communication with collaborators, we identified families that included at least one individual with at least three of the five main features of the DOORS syndrome: deafness, onychodystrophy, osteodystrophy, intellectual disability, and seizures. Participants were recruited from 26 centres in 17 countries. Families described in this study were enrolled between Dec 1, 2010, and March 1, 2013. Collaborating physicians enrolling participants obtained clinical information and DNA samples from the affected child and both parents if possible. We did whole-exome sequencing in affected individuals as they were enrolled, until we identified a candidate gene, and Sanger sequencing to confirm mutations. We did expression studies in human fibroblasts from one individual by real-time PCR and western blot analysis, and in mouse tissues by immunohistochemistry and real-time PCR.

Findings: 26 families were included in the study. We did exome sequencing in the first 17 enrolled families; we screened for TBC1D24 by Sanger sequencing in subsequent families. We identified TBC1D24 mutations in 11 individuals from nine families (by exome sequencing in seven families, and Sanger sequencing in two families). 18 families had individuals with all five main features of DOORS syndrome, and TBC1D24 mutations were identified in half of these families. The seizure types in individuals with TBC1D24 mutations included generalised tonic-clonic, complex partial, focal clonic, and infantile spasms. Of the 18 individuals with DOORS syndrome from 17 families without TBC1D24 mutations, eight did not have seizures and three did not have deafness. In expression studies, some mutations abrogated TBC1D24 mRNA stability. We also detected Tbc1d24 expression in mouse phalangeal chondrocytes and calvaria, which suggests a role of TBC1D24 in skeletogenesis.

Interpretation: Our findings suggest that mutations in TBC1D24 seem to be an important cause of DOORS syndrome and can cause diverse phenotypes. Thus, individuals with DOORS syndrome without deafness and seizures but with the other features should still be screened for TBC1D24 mutations. More information is needed to understand the cellular roles of TBC1D24 and identify the genes responsible for DOORS phenotypes in individuals who do not have a mutation in TBC1D24.

Funding: US National Institutes of Health, the CIHR (Canada), the NIHR (UK), the Wellcome Trust, the Henry Smith Charity, and Action Medical Research.
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http://dx.doi.org/10.1016/S1474-4422(13)70265-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3895324PMC
January 2014

Protein kinase cδ deficiency causes mendelian systemic lupus erythematosus with B cell-defective apoptosis and hyperproliferation.

Arthritis Rheum 2013 Aug;65(8):2161-71

Centre de Référence des Maladies Rénales Rares, Hospices Civils de Lyon, INSERM U1111, UMS3444/US8, Université Claude Bernard Lyon 1, and Université de Lyon, Lyon, France.

Objective: Systemic lupus erythematosus (SLE) is a prototype autoimmune disease that is assumed to occur via a complex interplay of environmental and genetic factors. Rare causes of monogenic SLE have been described, providing unique insights into fundamental mechanisms of immune tolerance. The aim of this study was to identify the cause of an autosomal-recessive form of SLE.

Methods: We studied 3 siblings with juvenile-onset SLE from 1 consanguineous kindred and used next-generation sequencing to identify mutations in the disease-associated gene. We performed extensive biochemical, immunologic, and functional assays to assess the impact of the identified mutations on B cell biology.

Results: We identified a homozygous missense mutation in PRKCD, encoding protein kinase δ (PKCδ), in all 3 affected siblings. Mutation of PRKCD resulted in reduced expression and activity of the encoded protein PKCδ (involved in the deletion of autoreactive B cells), leading to resistance to B cell receptor- and calcium-dependent apoptosis and increased B cell proliferation. Thus, as for mice deficient in PKCδ, which exhibit an SLE phenotype and B cell expansion, we observed an increased number of immature B cells in the affected family members and a developmental shift toward naive B cells with an immature phenotype.

Conclusion: Our findings indicate that PKCδ is crucial in regulating B cell tolerance and preventing self-reactivity in humans, and that PKCδ deficiency represents a novel genetic defect of apoptosis leading to SLE.
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http://dx.doi.org/10.1002/art.38008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4066615PMC
August 2013

Loss-of-function mutations in SMARCE1 cause an inherited disorder of multiple spinal meningiomas.

Nat Genet 2013 Mar 3;45(3):295-8. Epub 2013 Feb 3.

Genetic Medicine, Manchester Academic Health Sciences Centre (MAHSC), St. Mary's Hospital, University of Manchester, Manchester, UK.

One-third of all primary central nervous system tumors in adults are meningiomas. Rarely, meningiomas occur at multiple sites, usually occurring in individuals with type 2 neurofibromatosis (NF2). We sequenced the exomes of three unrelated individuals with familial multiple spinal meningiomas without NF2 mutations. We identified two individuals with heterozygous loss-of-function mutations in the SWI/SNF chromatin-remodeling complex subunit gene SMARCE1. Sequencing of SMARCE1 in six further individuals with spinal meningiomas identified two additional heterozygous loss-of-function mutations. Tumors from individuals with SMARCE1 mutations were of clear-cell histological subtype, and all had loss of SMARCE1 protein, consistent with a tumor suppressor mechanism. Our findings identify multiple-spinal-meningioma disease as a new discrete entity and establish a key role for the SWI/SNF complex in the pathogenesis of both meningiomas and tumors with clear-cell histology.
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http://dx.doi.org/10.1038/ng.2552DOI Listing
March 2013

Mutations in ADAR1 cause Aicardi-Goutières syndrome associated with a type I interferon signature.

Nat Genet 2012 Nov 23;44(11):1243-8. Epub 2012 Sep 23.

Manchester Academic Health Science Centre, University of Manchester, Genetic Medicine, UK.

Adenosine deaminases acting on RNA (ADARs) catalyze the hydrolytic deamination of adenosine to inosine in double-stranded RNA (dsRNA) and thereby potentially alter the information content and structure of cellular RNAs. Notably, although the overwhelming majority of such editing events occur in transcripts derived from Alu repeat elements, the biological function of non-coding RNA editing remains uncertain. Here, we show that mutations in ADAR1 (also known as ADAR) cause the autoimmune disorder Aicardi-Goutières syndrome (AGS). As in Adar1-null mice, the human disease state is associated with upregulation of interferon-stimulated genes, indicating a possible role for ADAR1 as a suppressor of type I interferon signaling. Considering recent insights derived from the study of other AGS-related proteins, we speculate that ADAR1 may limit the cytoplasmic accumulation of the dsRNA generated from genomic repetitive elements.
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http://dx.doi.org/10.1038/ng.2414DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4154508PMC
November 2012

A paradigm shift in the delivery of services for diagnosis of inherited retinal disease.

J Med Genet 2012 May;49(5):322-6

Genetic Medicine, Manchester Academic Health Sciences Centre, Central Manchester Foundation Trust, St Mary’s Hospital, Manchester, UK.

Objectives: Current technologies for delivering gene testing are labour-intensive and expensive. Over the last 3 years, new high-throughput DNA sequencing techniques (next generation sequencing; NGS), with the capability to analyse multiple genes or entire genomes, have been rapidly adopted into research. This study examines the possibility of incorporating NGS into a clinical UK service context.

Methods: The study applied NGS of 105 genes to 50 patients known to be affected by inherited forms of blindness in the setting of a UK National Health Service-accredited diagnostic molecular genetics laboratory. The study assessed the ability of an NGS protocol to identify likely disease-causing genetic variants when compared with current methodologies available through UK diagnostic laboratories.

Results: Conventional testing is only applicable to the minority of patients with inherited retinal disease and identifies mutations in fewer than one in four of those patients tested. By contrast, the NGS assay is directed at all patients with such disorders and identifies disease-causing mutations in 50--55%, which is a dramatic increase. This includes patients with apparently 'sporadic' disease, and those for whom clinical management and prognosis are altered as a consequence of defining their disease at a molecular level.

Conclusions: The new NGS approach delivers a step change in the diagnosis of inherited eye disease, provides precise diagnostic information and extends the possibility of targeted treatments including gene therapy. The approach represents an exemplar that illustrates the opportunity that NGS provides for broadening the availability of genetic testing. The technology will be applied to many conditions that are associated with high levels of genetic heterogeneity.
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http://dx.doi.org/10.1136/jmedgenet-2012-100847DOI Listing
May 2012

Mutations in CTC1, encoding conserved telomere maintenance component 1, cause Coats plus.

Nat Genet 2012 Jan 22;44(3):338-42. Epub 2012 Jan 22.

Manchester Academic Health Science Centre, University of Manchester, Genetic Medicine, UK.

Coats plus is a highly pleiotropic disorder particularly affecting the eye, brain, bone and gastrointestinal tract. Here, we show that Coats plus results from mutations in CTC1, encoding conserved telomere maintenance component 1, a member of the mammalian homolog of the yeast heterotrimeric CST telomeric capping complex. Consistent with the observation of shortened telomeres in an Arabidopsis CTC1 mutant and the phenotypic overlap of Coats plus with the telomeric maintenance disorders comprising dyskeratosis congenita, we observed shortened telomeres in three individuals with Coats plus and an increase in spontaneous γH2AX-positive cells in cell lines derived from two affected individuals. CTC1 is also a subunit of the α-accessory factor (AAF) complex, stimulating the activity of DNA polymerase-α primase, the only enzyme known to initiate DNA replication in eukaryotic cells. Thus, CTC1 may have a function in DNA metabolism that is necessary for but not specific to telomeric integrity.
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http://dx.doi.org/10.1038/ng.1084DOI Listing
January 2012

Exome sequence identifies RIPK4 as the Bartsocas-Papas syndrome locus.

Am J Hum Genet 2012 Jan 22;90(1):69-75. Epub 2011 Dec 22.

Faculty of Medical and Human Sciences, Manchester Academic Health Sciences Centre, University of Manchester, UK.

Pterygium syndromes are complex congenital disorders that encompass several distinct clinical conditions characterized by multiple skin webs affecting the flexural surfaces often accompanied by craniofacial anomalies. In severe forms, such as in the autosomal-recessive Bartsocas-Papas syndrome, early lethality is common, complicating the identification of causative mutations. Using exome sequencing in a consanguineous family, we identified the homozygous mutation c.1127C>A in exon 7 of RIPK4 that resulted in the introduction of the nonsense mutation p.Ser376X into the encoded ankyrin repeat-containing kinase, a protein that is essential for keratinocyte differentiation. Subsequently, we identified a second mutation in exon 2 of RIPK4 (c.242T>A) that resulted in the missense variant p.Ile81Asn in the kinase domain of the protein. We have further demonstrated that RIPK4 is a direct transcriptional target of the protein p63, a master regulator of stratified epithelial development, which acts as a nodal point in the cascade of molecular events that prevent pterygium syndromes.
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http://dx.doi.org/10.1016/j.ajhg.2011.11.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3257958PMC
January 2012

Exome sequencing identifies CCDC8 mutations in 3-M syndrome, suggesting that CCDC8 contributes in a pathway with CUL7 and OBSL1 to control human growth.

Am J Hum Genet 2011 Jul 7;89(1):148-53. Epub 2011 Jul 7.

Department of Endocrinology, Manchester Academic Health Sciences Centre, School of Biomedicine, University of Manchester, UK.

3-M syndrome, a primordial growth disorder, is associated with mutations in CUL7 and OBSL1. Exome sequencing now identifies mutations in CCDC8 as a cause of 3-M syndrome. CCDC8 is a widely expressed gene that is transcriptionally associated to CUL7 and OBSL1, and coimmunoprecipitation indicates a physical interaction between CCDC8 and OBSL1 but not CUL7. We propose that CUL7, OBSL1, and CCDC8 are members of a pathway controlling mammalian growth.
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http://dx.doi.org/10.1016/j.ajhg.2011.05.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3135816PMC
July 2011

Clustered coding variants in the glutamate receptor complexes of individuals with schizophrenia and bipolar disorder.

PLoS One 2011 Apr 29;6(4):e19011. Epub 2011 Apr 29.

Wellcome Trust Sanger Institute, Genome Campus, Hinxton, Cambridgeshire, United Kingdom.

Current models of schizophrenia and bipolar disorder implicate multiple genes, however their biological relationships remain elusive. To test the genetic role of glutamate receptors and their interacting scaffold proteins, the exons of ten glutamatergic 'hub' genes in 1304 individuals were re-sequenced in case and control samples. No significant difference in the overall number of non-synonymous single nucleotide polymorphisms (nsSNPs) was observed between cases and controls. However, cluster analysis of nsSNPs identified two exons encoding the cysteine-rich domain and first transmembrane helix of GRM1 as a risk locus with five mutations highly enriched within these domains. A new splice variant lacking the transmembrane GPCR domain of GRM1 was discovered in the human brain and the GRM1 mutation cluster could perturb the regulation of this variant. The predicted effect on individuals harbouring multiple mutations distributed in their ten hub genes was also examined. Diseased individuals possessed an increased load of deleteriousness from multiple concurrent rare and common coding variants. Together, these data suggest a disease model in which the interplay of compound genetic coding variants, distributed among glutamate receptors and their interacting proteins, contribute to the pathogenesis of schizophrenia and bipolar disorders.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0019011PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3084736PMC
April 2011

Whole-Exome sequencing identifies FAM20A mutations as a cause of amelogenesis imperfecta and gingival hyperplasia syndrome.

Am J Hum Genet 2011 May 5;88(5):616-20. Epub 2011 May 5.

Faculty of Medical and Human Sciences, Manchester Academic Health Sciences Centre, University of Manchester, Oxford Road, Manchester, UK.

Amelogenesis imperfecta (AI) describes a clinically and genetically heterogeneous group of disorders of biomineralization resulting from failure of normal enamel formation. AI is found as an isolated entity or as part of a syndrome, and an autosomal-recessive syndrome associating AI and gingival hyperplasia was recently reported. Using whole-exome sequencing, we identified a homozygous nonsense mutation in exon 2 of FAM20A that was not present in the Single Nucleotide Polymorphism database (dbSNP), the 1000 Genomes database, or the Centre d'Etude du Polymorphisme Humain (CEPH) Diversity Panel. Expression analyses indicated that Fam20a is expressed in ameloblasts and gingivae, providing biological plausibility for mutations in FAM20A underlying the pathogenesis of this syndrome.
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http://dx.doi.org/10.1016/j.ajhg.2011.04.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3146735PMC
May 2011

Expansion of GAA trinucleotide repeats in mammals.

Genomics 2006 Jan 28;87(1):57-67. Epub 2005 Nov 28.

Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.

We have previously shown that GAA trinucleotide repeats have undergone significant expansion in the human genome. Here we present the analysis of the length distribution of all 10 nonredundant trinucleotide repeat motifs in 20 complete eukaryotic genomes (6 mammalian, 2 nonmammalian vertebrates, 4 arthropods, 4 fungi, and 1 each of nematode, amoebozoa, alveolate, and plant), which showed that the abundance of large expansions of GAA trinucleotide repeats is specific to mammals. Analysis of human-chimpanzee-gorilla orthologs revealed that loci with large expansions are species-specific and have occurred after divergence from the common ancestor. PCR analysis of human controls revealed large expansions at multiple human (GAA)(30+) loci; nine loci showed expanded alleles containing >65 triplets, analogous to disease-causing expansions in Friedreich ataxia, including two that are in introns of genes of unknown function. The abundance of long GAA trinucleotide repeat tracts in mammalian genomes represents a significant mutation potential and source of interindividual variability.
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http://dx.doi.org/10.1016/j.ygeno.2005.09.006DOI Listing
January 2006