Natural Exon Skipping Sets the Stage for Exon Skipping as Therapy for Dystrophic Epidermolysis Bullosa

Contains fulltext : 214010.pdf (publisher's version ) (Open Access

Recurrent FXYD2 p.Gly41Arg mutation in patients with isolated dominant hypomagnesaemia

Background Magnesium (Mg2+) is an essential ion for cell growth, neuroplasticity and muscle contraction. Blood Mg2+ levels <0.7 mmol/L may cause a heterogeneous clinical phenotype, including muscle cramps and epilepsy and disturbances in K+ and Ca2+ homeostasis. Over the last decade, the genetic origin of several familial forms of hypomagnesaemia has been found. In 2000, mutations in FXYD2, encoding the γ-subunit of the Na+-K+-ATPase, were identified to cause isolated dominant hypomagnesaemia (IDH) in a large Dutch family suffering from hypomagnesaemia, hypocalciuria and chondrocalcinosis. However, no additional patients have been identified since then. Methods Here, two families with hypomagnesaemia and hypocalciuria were screened for mutations in the FXYD2 gene. Moreover, the patients were clinically and genetically characterized. Results We report a p.Gly41Arg FXYD2 mutation in two families with hypomagnesaemia and hypocalciuria. Interestingly, this is the same mutation as was described in the original study. As in the initial family, several patients suffered from muscle cramps, chondrocalcinosis and epilepsy. Haplotype analysis revealed an overlapping haplotype in all families, suggesting a founder effect. Conclusions The recurrent p.Gly41Arg FXYD2 mutation in two new families with IDH confirms that FXYD2 mutation causes hypomagnesaemia. Until now, no other FXYD2 mutations have been reported which could indicate that other FXYD2 mutations will not cause hypomagnesaemia or are embryonically letha

Mechanisms of Natural Gene Therapy in Dystrophic Epidermolysis Bullosa

Item does not contain fulltextRevertant mosaicism has been reported in several inherited diseases, including the genetic skin fragility disorder epidermolysis bullosa (EB). Here, we describe the largest cohort of seven patients with revertant mosaicism and dystrophic EB (DEB), associated with mutations in the COL7A1 gene, and determine the underlying molecular mechanisms. We show that revertant mosaicism occurs both in autosomal dominantly and recessively inherited DEB. We found that null mutations resulting in complete loss of collagen VII and severe disease, as well as missense or splice-site mutations associated with some preserved collagen VII function and a milder phenotype, were corrected by revertant mosaicism. The mutation, subtype, and severity of the disease are thus not decisive for the presence of revertant mosaicism. Although collagen VII is synthesized and secreted by both keratinocytes and fibroblasts, evidence for reversion was only found in keratinocytes. The reversion mechanisms included back mutations/mitotic recombinations in 70% of the cases and second-site mutations affecting splicing in 30%. We conclude that revertant mosaicism is more common than previously assumed in patients with DEB, and our findings will have implications for future therapeutic strategies using the patient's naturally corrected cells as a source for cell-based therapies

ADULT syndrome : phenotype in a Brazilian family with the R298Q mutation

Transcription factor p63 is important for embryonic regulation of ectodermal, orofacial, and limb development.1 Mutations in the p63 gene are associated with six overlapping phenotypes:2 Ankyloblepharon‐ectodermal defects‐cleft lip/palate (AEC) syndrome (including Rapp‐Hodgkin syndrome), Ectrodactyly‐Ectodermal dysplasia‐cleft lip/palate (EEC) syndrome, Limb‐mammary syndrome (LMS), Split‐hand/foot malformation (SHFM), isolated cleft lip/palate and Acro‐dermo‐ungual‐lacrimal‐tooth (ADULT) syndrome.3-

KIF1A variants are a frequent cause of autosomal dominant hereditary spastic paraplegia

Contains fulltext : 214064.pdf (publisher's version ) (Open Access

Design and Validation of a Conformation Sensitive Capillary Electrophoresis-Based Mutation Scanning System and Automated Data Analysis of the More than 15 kbp-Spanning Coding Sequence of the SACS Gene

In this study, we developed and analytically validated a fully automated, robust confirmation sensitive capillary electrophoresis (CSCE) method to perform mutation scanning of the large SACS gene. This method facilitates a rapid and cost-effective molecular diagnosis of autosomal recessive spastic ataxia of Charlevoix-Saguenay. Critical issues addressed during the development of the CSCE system included the position of a DNA variant relative to the primers and the CG-content of the amplicons. The validation was performed in two phases; a retrospective analysis of 32 samples containing 41 different known DNA variants and a prospective analysis of 20 samples of patients clinically suspected of having autosomal recessive spastic ataxia of Charlevoix-Saguenay. These 20 samples appeared to contain 73 DNA variants. In total, in 32 out of the 45 amplicons, a DNA variant was present, which allowed verification of the detection capacity during the validation process. After optimization of the original design, the overall analytical sensitivity of CSCE for the SACS gene was 100%, and the analytical specificity of CSCE was 99.8%. In conclusion, CSCE is a robust technique with a high analytical sensitivity and specificity, and it can readily be used for mutation scanning of the large SACS gene. Furthermore this technique is less time-consuming and less expensive, as compared with standard automated sequencing

A novel translation re-initiation mechanism for the p63 gene revealed by amino-terminal truncating mutations in Rapp-Hodgkin/Hay-Wells-like syndromes

Missense mutations in the 3′ end of the p63 gene are associated with either RHS (Rapp-Hodgkin syndrome) or AEC (Ankyloblepharon Ectodermal defects Cleft lip/palate) syndrome. These mutations give rise to mutant p63α protein isoforms with dominant effects towards their wild-type counterparts. Here we report four RHS/AEC-like patients with mutations (p.Gln9fsX23, p.Gln11X, p.Gln16X), that introduce premature termination codons in the N-terminal part of the p63 protein. These mutations appear to be incompatible with the current paradigms of dominant-negative/gain-of-function outcomes for other p63 mutations. Moreover it is difficult to envisage how the remaining small N-terminal polypeptide contributes to a dominant disease mechanism. Primary keratinocytes from a patient containing the p.Gln11X mutation revealed a normal and aberrant p63-related protein that was just slightly smaller than the wild-type p63. We show that the smaller p63 protein is produced by translation re-initiation at the next downstream methionine, causing truncation of a non-canonical transactivation domain in the ΔN-specific isoforms. Interestingly, this new ΔΔNp63 isoform is also present in the wild-type keratinocytes albeit in small amounts compared with the p.Gln11X patient. These data establish that the p.Gln11X-mutation does not represent a null-allele leading to haploinsufficiency, but instead gives rise to a truncated ΔNp63 protein with dominant effects. Given the nature of other RHS/AEC-like syndrome mutations, we conclude that these mutations affect only the ΔNp63α isoform and that this disruption is fundamental to explaining the clinical characteristics of these particular ectodermal dysplasia syndromes

The inversa type of recessive dystrophic epidermolysis bullosa is caused by specific arginine and glycine substitutions in type VII collagen

International audienceBackground The inversa type of recessive dystrophic epidermolysis bullosa (RDEB-I) is a rare variant of dystrophic epidermolysis bullosa, characterized by blistering in the body flexures, trunk, and mucosae. The cause of this specific distribution is unknown. So far, 20 COL7A1 genotypes have been described in RDEB-I and genotype-phenotype correlations have not been studied extensively. The aim of the study was to gain more insight into the pathophysiology of this intriguing RDEB-I phenotype. Methods We identified 20 Dutch and British RDEB-I patients and full genotypes in 18 of them. We reviewed the literature on RDEB-I genotypes and conducted an extensive genotype-phenotype correlation study for RDEB-I. Results All 20 patients had generalized blistering at birth and during early infancy. In most patients, the age of transition from generalized to inversa distribution was before the age of 4 years. We noted a spectrum of disease severity ranging from the mildest 'mucosal only' phenotype to the severest phenotype with limited acral involvement. The 29 genotypes of our RDEB-I patients and those reported in the literature revealed that RDEB-I is associated with specific recessive arginine and glycine substitutions in the triple-helix domain of type VII collagen. Discussion and conclusion Why these specific arginine and glycine substitutions cause the inversa distribution remains unknown. We could not identify clear differences in location and nature of substituting amino acids between these mutations and missense mutations causing other RDEB phenotypes. We hypothesize that the higher skin temperature in the affected areas plays an important role in the pathophysiology of RDEB-I