John Compton

Common loss-of-function variants of the epidermal barrier protein filaggrin are a major predisposing factor for atopic dermatitis

Atopic disease, including atopic dermatitis (eczema), allergy and asthma, has increased in frequency in recent decades and now affects ∼20% of the population in the developed world. Twin and family studies have shown that predisposition to atopic disease is highly heritable. Although most genetic studies have focused on immunological mechanisms, a primary epithelial barrier defect has been anticipated. Filaggrin is a key protein that facilitates terminal differentiation of the epidermis and formation of the skin barrier. Here we show that two independent loss-of-function genetic variants (R510X and 2282del4) in the gene encoding filaggrin (FLG) are very strong predisposing factors for atopic dermatitis. These variants are carried by ∼9% of people of European origin. These variants also show highly significant association with asthma occurring in the context of atopic dermatitis. This work establishes a key role for impaired skin barrier function in the development of atopic disease.

Loss-of-function mutations in the gene encoding filaggrin cause ichthyosis vulgaris.

Ichthyosis vulgaris (OMIM 146700) is the most common inherited disorder of keratinization and one of the most frequent single-gene disorders in humans. The most widely cited incidence figure is 1 in 250 based on a survey of 6,051 healthy English schoolchildren. We have identified homozygous or compound heterozygous mutations R501X and 2282del4 in the gene encoding filaggrin (FLG) as the cause of moderate or severe ichthyosis vulgaris in 15 kindreds. In addition, these mutations are semidominant; heterozygotes show a very mild phenotype with incomplete penetrance. The mutations show a combined allele frequency of ∼4% in populations of European ancestry, explaining the high incidence of ichthyosis vulgaris. Profilaggrin is the major protein of keratohyalin granules in the epidermis. During terminal differentiation, it is cleaved into multiple filaggrin peptides that aggregate keratin filaments. The resultant matrix is cross-linked to form a major component of the cornified cell envelope. We find that loss or reduction of this major structural protein leads to varying degrees of impaired keratinization.

Assuring the quality of next-generation sequencing in clinical laboratory practice

Amy S Gargis, Centers for Disease Control and Prevention Lisa Kalman, Centers for Disease Control and Prevention Meredith W Berry, SeqWright Inc David P Bick, Medical College of Wisconsin David P Dimmock, Medical College of Wisconsin Tina Hambuch, Illumina Clinical Services Fei Lu, SeqWright Inc Elaine Lyon, University of Utah Karl V Voelkerding, University of Utah Barbara Zehnbauer, Emory University

An evidence-based approach to establish the functional and clinical significance of copy number variants in intellectual and developmental disabilities

Purpose: Copy number variants have emerged as a major cause of human disease such as autism and intellectual disabilities. Because copy number variants are common in normal individuals, determining the functional and clinical significance of rare copy number variants in patients remains challenging. The adoption of whole-genome chromosomal microarray analysis as a first-tier diagnostic test for individuals with unexplained developmental disabilities provides a unique opportunity to obtain large copy number variant datasets generated through routine patient care.Methods: A consortium of diagnostic laboratories was established (the International Standards for Cytogenomic Arrays consortium) to share copy number variant and phenotypic data in a central, public database. We present the largest copy number variant case-control study to date comprising 15,749 International Standards for Cytogenomic Arrays cases and 10,118 published controls, focusing our initial analysis on recurrent deletions and duplications involving 14 copy number variant regions.Results: Compared with controls, 14 deletions and seven duplications were significantly overrepresented in cases, providing a clinical diagnosis as pathogenic.Conclusion: Given the rapid expansion of clinical chromosomal microarray analysis testing, very large datasets will be available to determine the functional significance of increasingly rare copy number variants. This data will provide an evidence-based guide to clinicians across many disciplines involved in the diagnosis, management, and care of these patients and their families.

Prevalence and nonrandom distribution of exonic mutations in interferon regulatory factor 6 in 307 families with Van der Woude syndrome and 37 families with popliteal pterygium syndrome.

Purpose: Interferon regulatory factor 6 encodes a member of the IRF family of transcription factors. Mutations in interferon regulatory factor 6 cause Van der Woude and popliteal pterygium syndrome, two related orofacial clefting disorders. Here, we compared and contrasted the frequency and distribution of exonic mutations in interferon regulatory factor 6 between two large geographically distinct collections of families with Van der Woude and between one collection of families with popliteal pterygium syndrome.Methods: We performed direct sequence analysis of interferon regulatory factor 6 exons on samples from three collections, two with Van der Woude and one with popliteal pterygium syndrome.Results: We identified mutations in interferon regulatory factor 6 exons in 68% of families in both Van der Woude collections and in 97% of families with popliteal pterygium syndrome. In sum, 106 novel disease-causing variants were found. The distribution of mutations in the interferon regulatory factor 6 exons in each collection was not random; exons 3, 4, 7, and 9 accounted for 80%. In the Van der Woude collections, the mutations were evenly divided between protein truncation and missense, whereas most mutations identified in the popliteal pterygium syndrome collection were missense. Further, the missense mutations associated with popliteal pterygium syndrome were localized significantly to exon 4, at residues that are predicted to bind directly to DNA.Conclusion: The nonrandom distribution of mutations in the interferon regulatory factor 6 exons suggests a two-tier approach for efficient mutation screens for interferon regulatory factor 6. The type and distribution of mutations are consistent with the hypothesis that Van der Woude is caused by haploinsufficiency of interferon regulatory factor 6. On the other hand, the distribution of popliteal pterygium syndrome-associated mutations suggests a different, though not mutually exclusive, effect on interferon regulatory factor 6 function.

Comparative analysis of IRF6 variants in families with Van der Woude syndrome and popliteal pterygium syndrome using public whole-exome databases

Purpose:Mutations in the transcription factor IRF6 cause allelic autosomal dominant clefting syndromes, Van der Woude syndrome, and popliteal pterygium syndrome. We compared the distribution of IRF6 coding and splice-site mutations from 549 families with Van der Woude syndrome or popliteal pterygium syndrome with that of variants from the 1000 Genomes and National Heart, Lung, and Blood Institute Exome Sequencing Projects.Methods:We compiled all published pathogenic IRF6 mutations and performed direct sequencing of IRF6 in families with Van der Woude syndrome or popliteal pterygium syndrome.Results:Although mutations causing Van der Woude syndrome or popliteal pterygium syndrome were nonrandomly distributed with significantly increased frequencies in the DNA-binding domain (P = 0.0001), variants found in controls were rare and evenly distributed in IRF6. Of 194 different missense or nonsense variants described as potentially pathogenic, we identified only two in more than 6,000 controls. PolyPhen and SIFT (sorting intolerant from tolerant) reported 5.9% of missense mutations in patients as benign, suggesting that use of current in silico prediction models to determine function can have significant false negatives.Conclusion:Mutation of IRF6 occurs infrequently in controls, suggesting that for IRF6 there is a high probability that disruption of the coding sequence, particularly the DNA-binding domain, will result in syndromic features. Prior associations of coding sequence variants in IRF6 with clefting syndromes have had few false positives.Genet Med 2013:15(5):338–344

Oligonucleotide array CGH studies in myeloproliferative neoplasms: Comparison with JAK2V617F mutational status and conventional chromosome analysis

Comparative genomic hybridization (CGH), using oligo arrays with either 44,000 or 105,000 oligonucleotides, was performed on granulocyte-derived DNA from 71 patients with BCR-ABL-negative classic myeloproliferative neoplasms (MPNs): 32 primary myelofibrosis (PMF), 26 polycythemia vera (PV) and 13 essential thrombocythemia (ET). Copy number changes (CNCs) were detected in 44%, 35%, and 15% of the cases with PMF, PV and ET, respectively. In ET and PMF, CNCs were more frequently detected in the presence of JAK2V617F (50% vs. 19%; p=0.05). Conventional chromosome analysis was obtained in 57 patients either at diagnosis or within 1 year of the array CGH study; all 21 patients with PV and 11 with ET displayed normal cytogenetic findings despite the presence of CNCs in 29% and 18%, respectively. In PMF, the respective rates of CNCs and abnormal karyotype were 48% and 36%; karyotypic abnormalities, including unbalanced translocations, were often detected by array CGH as chromosomal gains or losses. This preliminary report suggests a potential value for array CGH in terms of both clinical diagnostics and genomic research in MPNs.