Silent polymorphisms in the RYR1 gene do not\ud modify the phenotype of the p.4898 I>T\ud pathogenic mutation in central core disease:\ud a case report

Background: Central core disease is a congenital myopathy, characterized by presence of central core-like areas in\ud muscle fibers. Patients have mild or moderate weakness, hypotonia and motor developmental delay. The disease is\ud caused by mutations in the human ryanodine receptor gene (RYR1), which encodes a calcium-release channel.\ud Since the RYR1 gene is huge, containing 106 exons, mutation screening has been limited to three ‘hot spots’, with\ud particular attention to the C-terminal region. Recent next- generation sequencing methods are now identifying\ud multiple numbers of variants in patients, in which interpretation and phenotype prevision is difficult.\ud Case presentation: In a Brazilian Caucasian family, clinical, histopathological and molecular analysis identified a\ud new case of central core disease in a 48-year female. Sanger sequencing of the C-terminal region of the RYR1\ud gene identified two different missense mutations: c.14256 A > C polymorphism in exon 98 and c.14693 T > C in\ud exon 102, which have already been described as pathogenic. Trans-position of the 2 mutations was confirmed\ud because patient’s daughter, mother and sister carried only the exon 98’s mutation, a synonymous variant that was\ud subsequently found in the frequency of 013–0,05 of alleles. Further next generation sequencing study of the whole\ud RYR1 gene in the patient revealed the presence of additional 5 common silent polymorphisms in homozygosis and\ud 8 polymorphisms in heterozygosis.\ud Conclusions: Considering that patient’s relatives showed no pathologic phenotype, and the phenotype presented\ud by the patient is within the range observed in other central core disease patients with the same mutation, it was\ud concluded that the c.14256 A > C polymorphism alone is not responsible for disease, and the associated additional\ud silent polymorphisms are not acting as modifiers of the primary pathogenic mutation in the affected patient. The\ud case described above illustrates the present reality where new methods for wide genome screening are becoming\ud more accessible and able to identify a great variety of mutations and polymorphisms of unknown function in\ud patients and their families.Fundação de Amparo a Pesquisa do Estado de São Paulo - Centro de Pesquisa, Inovação e Difusão (FAPESP-CEPID)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq-INCT)Associação Brasileira de Distrofia Muscular (ABDIM)CAPES-COFECU

Integrative Variation Analysis Reveals that a Complex Genotype May Specify Phenotype in Siblings with Syndromic Autism Spectrum Disorder

<div><p>It has been proposed that copy number variations (CNVs) are associated with increased risk of autism spectrum disorder (ASD) and, in conjunction with other genetic changes, contribute to the heterogeneity of ASD phenotypes. Array comparative genomic hybridization (aCGH) and exome sequencing, together with systems genetics and network analyses, are being used as tools for the study of complex disorders of unknown etiology, especially those characterized by significant genetic and phenotypic heterogeneity. Therefore, to characterize the complex genotype-phenotype relationship, we performed aCGH and sequenced the exomes of two affected siblings with ASD symptoms, dysmorphic features, and intellectual disability, searching for <i>de novo</i> CNVs, as well as for <i>de novo</i> and rare inherited point variations—single nucleotide variants (SNVs) or small insertions and deletions (indels)—with probable functional impacts. With aCGH, we identified, in both siblings, a duplication in the 4p16.3 region and a deletion at 8p23.3, inherited by a paternal balanced translocation, t(4, 8) (p16; p23). Exome variant analysis found a total of 316 variants, of which 102 were shared by both siblings, 128 were in the male sibling exome data, and 86 were in the female exome data. Our integrative network analysis showed that the siblings’ shared translocation could explain their similar syndromic phenotype, including overgrowth, macrocephaly, and intellectual disability. However, exome data aggregate genes to those already connected from their translocation, which are important to the robustness of the network and contribute to the understanding of the broader spectrum of psychiatric symptoms. This study shows the importance of using an integrative approach to explore genotype-phenotype variability.</p></div

Summary of the characteristics observed in our patients, compared with those of other cases described in the literature.

<p>Summary of the characteristics observed in our patients, compared with those of other cases described in the literature.</p

Details of the hemizygous variations found in the exome of the male sibling.

<p>Details of the hemizygous variations found in the exome of the male sibling.</p

Summary diagram of the variations found in the two siblings.

<p>Summary diagram of the variations found in the two siblings.</p

Female sibling STRING network analysis.

<p>Biggest component of the connecting brain-expressed genes present in the unbalanced translocation, altered by the shared and exclusive rare inherited SNV and indels found in the female sibling.</p

Comparison of broker and bridge genes in the biggest connected components of the networks of the two siblings.

<p>Comparison of broker and bridge genes in the biggest connected components of the networks of the two siblings.</p

FISH images.

<p>(A) image showing that, in the female sibling, there were three chr4p copies, marked in green with a GS-118B13 probe (green arrow), and one chr8p copy, marked in red with a RP11-338B22 probe (red arrow); (B) image showing that, in the male sibling, there were also three chr4p copies, marked in red with a GS-118B13 probe (red arrow), and one chr8p copy, marked in green with a GS-77L23 probe (green arrow); (C) image showing that, in the father, there was a balanced translocation, t(4;8); and (D) image showing that, in the mother, the chromosomes were normal.</p

CNVs at the chromosomal regions 15q13.3, 16p11.2 and 22q13 in the Brazilian individuals with ASD.

<p><b>Neurol. Dis.</b> - Neurological Disorder; “<b>–</b>” – not reported in the literature; “<b>+</b>” reported in the literature; del – deletion; dup – duplication; n.a. – not available; pat - paternal; mat - maternal;</p>¥<p>- ASD-affected individual 10 was described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0107705#pone.0107705-Jehee1" target="_blank">[37]</a>; * CNVs at 15q13.3 and 15q11.2 are present in both parents;</p><p>CNVs at the chromosomal regions 15q13.3, 16p11.2 and 22q13 in the Brazilian individuals with ASD.</p