The driver landscape of sporadic chordoma

Chordoma is a malignant, often incurable bone tumour showing notochordal differentiation. Here, we defined the somatic driver landscape of 104 cases of sporadic chordoma. We reveal somatic duplications of the notochordal transcription factor brachyury (T) in up to 27% of cases. These variants recapitulate the rearrangement architecture of the pathogenic germline duplications of T that underlie familial chordoma. In addition, we find potentially clinically actionable PI3K signalling mutations in 16% of cases. Intriguingly, one of the most frequently altered genes, mutated exclusively by inactivating mutation, was LYST (10%), which may represent a novel cancer gene in chordoma

Deep Sequencing of Target Linkage Assay-Identified Regions in Familial Breast Cancer: Methods, Analysis Pipeline and Troubleshooting

Background: The classical candidate-gene approach has failed to identify novel breast cancer susceptibility genes. Nowadays, massive parallel sequencing technology allows the development of studies unaffordable a few years ago. However, analysis protocols are not yet sufficiently developed to extract all information from the huge amount of data obtained. Methodology/Principal Findings: In this study, we performed high throughput sequencing in two regions located on chromosomes 3 and 6, recently identified by linkage studies by our group as candidate regions for harbouring breast cancer susceptibility genes. In order to enrich for the coding regions of all described genes located in both candidate regions, a hybrid-selection method on tiling microarrays was performed. Conclusions/Significance: We developed an analysis pipeline based on SOAP aligner to identify candidate variants with a high real positive confirmation rate (0.89), with which we identified eight variants considered candidates for functiona

Erratum to: Mutation screening in 86 known X-linked mental retardation genes by droplet-based multiplex PCR and massive parallel sequencing

Item does not contain fulltext[This corrects the article DOI: 10.1007/s11568-010-9137-y.].1 december 200

Molecular changes in the expression of human colonic nutrient transporters during the transition from normality to malignancy

Healthy colonocytes derive 60–70% of their energy supply from short-chain fatty acids, particularly butyrate. Butyrate has profound effects on differentiation, proliferation and apoptosis of colonic epithelial cells by regulating expression of various genes associated with these processes. We have previously shown that butyrate is transported across the luminal membrane of the colonic epithelium via a monocarboxylate transporter, MCT1. In this paper, using immunohistochemistry and in situ hybridisation histochemistry, we have determined the profile of MCT1 protein and mRNA expression along the crypt to surface axis of healthy human colonic tissue. There is a gradient of MCT1 protein expression in the apical membrane of the cells along the crypt-surface axis rising to a peak in the surface epithelial cells. MCT1 mRNA is expressed along the crypt-surface axis and is most abundant in cells lining the crypt. Analysis of healthy colonic tissues and carcinomas using immunohistochemistry and Western blotting revealed a significant decline in the expression of MCT1 protein during transition from normality to malignancy. This was reflected in a corresponding reduction in MCT1 mRNA expression, as measured by Northern analysis. Carcinoma samples displaying reduced levels of MCT1 were found to express the high affinity glucose transporter, GLUT1, suggesting that there is a switch from butyrate to glucose as an energy source in colonic epithelia during transition to malignancy. The expression levels of MCT1 in association with GLUT1 could potentially be used as determinants of the malignant state of colonic tissue

Massively parallel sequencing of patients with intellectual disability, congenital anomalies and/or autism spectrum disorders with a targeted gene panel

10.1371/journal.pone.0093409PLoS ONE94-POLN

Efficient Utilization of Rare Variants for Detection of Disease-Related Genomic Regions

When testing association between rare variants and diseases, an efficient analytical approach involves considering a set of variants in a genomic region as the unit of analysis. One factor complicating this approach is that the vast majority of rare variants in practical applications are believed to represent background neutral variation. As a result, analyzing a single set with all variants may not represent a powerful approach. Here, we propose two alternative strategies. In the first, we analyze the subsets of rare variants exhaustively. In the second, we categorize variants selectively into two subsets: one in which variants are overrepresented in cases, and the other in which variants are overrepresented in controls. When the proportion of neutral variants is moderate to large we show, by simulations, that the both proposed strategies improve the statistical power over methods analyzing a single set with total variants. When applied to a real sequencing association study, the proposed methods consistently produce smaller p-values than their competitors. When applied to another real sequencing dataset to study the difference of rare allele distributions between ethnic populations, the proposed methods detect the overrepresentation of variants between the CHB (Chinese Han in Beijing) and YRI (Yoruba people of Ibadan) populations with small p-values. Additional analyses suggest that there is no difference between the CHB and CHD (Chinese Han in Denver) datasets, as expected. Finally, when applied to the CHB and JPT (Japanese people in Tokyo) populations, existing methods fail to detect any difference, while it is detected by the proposed methods in several regions

HUWE1 mutations in Juberg-Marsidi and Brooks syndromes: the results of an X-chromosome exome sequencing study

Background: X linked intellectual disability (XLID) syndromes account for a substantial number of males with ID. Much progress has been made in identifying the genetic cause in many of the syndromes described 20-40 years ago. Next generation sequencing (NGS) has contributed to the rapid discovery of XLID genes and identifying novel mutations in known XLID genes for many of these syndromes. Methods: 2 NGS approaches were employed to identify mutations in X linked genes in families with XLID disorders. 1 involved exome sequencing of genes on the X chromosome using the Agilent SureSelect Human X Chromosome Kit. The second approach was to conduct targeted NGS sequencing of 90 known XLID genes. Results: We identified the same mutation, a c.12928 G>C transversion in the HUWE1 gene, which gives rise to a p.G4310R missense mutation in 2 XLID disorders: Juberg-Marsidi syndrome (JMS) and Brooks syndrome. Although the original families with these disorders were considered separate entities, they indeed overlap clinically. A third family was also found to have a novel HUWE1 mutation. Conclusions: As we identified a HUWE1 mutation in an affected male from the original family reported by Juberg and Marsidi, it is evident the syndrome does not result from a mutation in ATRX as reported in the literature. Additionally, our data indicate that JMS and Brooks syndromes are allelic having the same HUWE1 mutation.Michael J Friez, Susan Sklower Brooks, Roger E Stevenson, Michael Field, Monica J Basehore, Lesley C Adès, Courtney Sebold, Stephen McGee, Samantha Saxon, Cindy Skinner, Maria E Craig, Lucy Murray, Richard J Simensen, Ying Yzu Yap, Marie A Shaw, Alison Gardner, Mark Corbett, Raman Kumar, Matthias Bosshard, Barbara van Loon, Patrick S Tarpey, Fatima Abidi, Jozef Gecz, Charles E Schwart

Lack of Cul4b, an E3 Ubiquitin Ligase Component, Leads to Embryonic Lethality and Abnormal Placental Development

Cullin-RING ligases (CRLs) complexes participate in the regulation of diverse cellular processes, including cell cycle progression, transcription, signal transduction and development. Serving as the scaffold protein, cullins are crucial for the assembly of ligase complexes, which recognize and target various substrates for proteosomal degradation. Mutations in human CUL4B, one of the eight members in cullin family, are one of the major causes of X-linked mental retardation. We here report the generation and characterization of Cul4b knockout mice, in which exons 3 to 5 were deleted. In contrast to the survival to adulthood of human hemizygous males with CUL4B null mutation, Cul4b null mouse embryos show severe developmental arrest and usually die before embryonic day 9.5 (E9.5). Accumulation of cyclin E, a CRL (CUL4B) substrate, was observed in Cul4b null embryos. Cul4b heterozygotes were recovered at a reduced ratio and exhibited a severe developmental delay. The placentas in Cul4b heterozygotes were disorganized and were impaired in vascularization, which may contribute to the developmental delay. As in human CUL4B heterozygotes, Cul4b null cells were selected against in Cul4b heterozygotes, leading to various degrees of skewed X-inactivation in different tissues. Together, our results showed that CUL4B is indispensable for embryonic development in the mouse

Improved Detection of Rare Genetic Variants for Diseases

Technology advances have promoted gene-based sequencing studies with the aim of identifying rare mutations responsible for complex diseases. A complication in these types of association studies is that the vast majority of non-synonymous mutations are believed to be neutral to phenotypes. It is thus critical to distinguish potential causative variants from neutral variation before performing association tests. In this study, we used existing predicting algorithms to predict functional amino acid substitutions, and incorporated that information into association tests. Using simulations, we comprehensively studied the effects of several influential factors, including the sensitivity and specificity of functional variant predictions, number of variants, and proportion of causative variants, on the performance of association tests. Our results showed that incorporating information regarding functional variants obtained from existing prediction algorithms improves statistical power under certain conditions, particularly when the proportion of causative variants is moderate. The application of the proposed tests to a real sequencing study confirms our conclusions. Our work may help investigators who are planning to pursue gene-based sequencing studies