Compound heterozygous mutations in RIPPLY2 associated with vertebral segmentation defects

Segmentation defects of the vertebrae (SDV) are caused by aberrant somite formation during embryogenesis and result in irregular formation of the vertebrae and ribs. The Notch signal transduction pathway plays a critical role in somite formation and patterning in model vertebrates. In humans, mutations in several genes involved in the Notch pathway are associated with SDV, with both autosomal recessive (MESP2, DLL3, LFNG, HES7) and autosomal dominant (TBX6) inheritance. However, many individuals with SDV do not carry mutations in these genes. Using whole-exome capture and massive parallel sequencing, we identified compound heterozygous mutations in RIPPLY2 in two brothers with multiple regional SDV, with appropriate familial segregation. One novel mutation (c.A238T:p.Arg80*) introduces a premature stop codon. In transiently transfected C2C12 mouse myoblasts, the RIPPLY2 mutant protein demonstrated impaired transcriptional repression activity compared with wild-type RIPPLY2 despite similar levels of expression. The other mutation (c.240-4T>G), with minor allele frequency <0.002, lies in the highly conserved splice site consensus sequence 5' to the terminal exon. Ripply2 has a well-established role in somitogenesis and vertebral column formation, interacting at both gene and protein levels with SDV-associated Mesp2 and Tbx6. We conclude that compound heterozygous mutations in RIPPLY2 are associated with SDV, a new gene for this condition. © The Author 2014.

Next generation sequencing identifies novel CACNA1A gene mutations in Episodic Ataxia type 2

Episodic Ataxia type 2 (EA2) is a rare autosomal dominantly inherited neurological disorder characterized by recurrent disabling imbalance, vertigo and episodes of ataxia lasting minutes to hours. EA2 is caused most often by loss of function mutations of the calcium channel gene CACNA1A. In addition to EA2, mutations in CACNA1A are responsible for two other allelic disorders: familial hemiplegic migraine type1 (FHM1) and spinocerebellar ataxia type 6 (SCA6). Herein, we have utilised Next Generation Sequencing (NGS) to screen the coding sequence, exon-intron boundaries and UTRs of five genes where mutation is known to produce symptoms related to EA2, including CACNA1A. We performed this screening in a group of 31 unrelated patients with EA2 symptoms. Both novel and known mutations were detected through NGS technology, and confirmed through Sanger sequencing. Genetic testing showed in total 15 mutation bearing patients (48%), of which 9 were novel mutations (6 missense and 3 small frameshift deletion mutations) and six known mutations (4 missense and 2 nonsense).These results demonstrate the efficiency of our NGS-panel for detecting known and novel mutations for EA2 in the CACNA1A gene, also identifying a novel missense mutation in ATP1A2 which is not a normal target for EA2 screening.

Genome-wide DNA methylation profiling of CD8+ T cells shows a distinct epigenetic signature to CD4+ T cells in multiple sclerosis patients

Background Multiple sclerosis (MS) is thought to be a T cell-mediated autoimmune disorder. MS pathogenesis is likely due to a genetic predisposition triggered by a variety of environmental factors. Epigenetics, particularly DNA methylation, provide a logical interface for environmental factors to influence the genome. In this study we aim to identify DNA methylation changes associated with MS in CD8+ T cells in 30 relapsing remitting MS patients and 28 healthy blood donors using Illumina 450K methylation arrays. Findings Seventy-nine differentially methylated CpGs were associated with MS. The methylation profile of CD8+ T cells was distinctive from our previously published data on CD4+ T cells in the same cohort. Most notably, there was no major CpG effect at the MS risk gene HLA-DRB1 locus in the CD8+ T cells. Conclusion CD8+ T cells and CD4+ T cells have distinct DNA methylation profiles. This case–control study highlights the importance of distinctive cell subtypes when investigating epigenetic changes in MS and other complex diseases.

Epigenetics Underpinning DNA Damage Repair

Epigenetics is the study of heritable changes in gene expression that are not the result of genetic alterations. These changes include DNA methylation, histone modifications, or indeed microRNA expression. Chromatin is a tightly compacted DNA–protein complex that allows approximately two meters of DNA to be packaged inside a cell, only a few micrometers across. Although the resulting DNA structure is very stable, it is not very amiable to DNA-dependent processes, so mechanisms have to exist to allow processes such as transcription, replication, and DNA repair to occur. This chapter will look at how a cell responds to and deals with genomic instability at the epigenetic level and highlight how critical chromatin remodeling is for correct DNA repair and cell survival following DNA damage. This chapter will initially look at the DNA repair pathways that function in human cells and then at how the repair of DNA damage is controlled by epigenetics.