Phenotypic features of CRB1-associated early-onset severe retinal dystrophy and the different molecular approaches to identifying the disease-causing variants.

PURPOSE: The aim of this study was to determine the molecular genetic basis of an early-onset severe retinal dystrophy in three unrelated consecutive patients of Czech origin and to describe their ocular phenotype. METHODS: DNA samples from two probands were analyzed using a genotyping microarray (Asper) followed by either target analysis of 43 genes implicated in retinal disorders by next generation sequencing or whole-exome sequencing, respectively. The third proband underwent conventional Sanger sequencing of CRB1 based on her ocular findings. RESULTS: All three probands harboured a known disease-causing mutation c.2843G>A; p.(Cys948Tyr) in the CRB1 gene. One individual was homozygous for this mutation, while in the other two probands c.2308G>A; p.(Gly770Ser) and c.3121A>G; p.(Met1041Val) were also identified in the heterozygous state, respectively. Both variants were novel and evaluated by in silico analysis as pathogenic. A false-negative result was observed in one of the two samples examined by the genotyping microarray. Disease onset in all patients was before the age of 7 years. Hypermetropic refractive error, bilateral nummular retinal pigmentation, retinal thickening and cystoid spaces in the macula were observed in two probands, aged 6 and 7 years. The third proband, aged 28 years, had bone spicule-like pigmentary changes associated with increased retinal nerve fiber layer. CONCLUSIONS: The first study reporting on the molecular genetic cause of non-syndromic early-onset severe retinal dystrophy in Czech patients identified one homozygous and two compound heterozygote probands with CRB1 mutations. Retina nerve fibre layer measurements should be considered an integral part of the clinical evaluation of retinal dystrophies. Detailed clinical examination and imaging can both direct molecular screening and help to confirm or refute disease causation of identified variants.

A 15 Mb large paracentric chromosome 21 inversion identified in Czech population through a pair of flanking duplications.

BACKGROUND: Inversions are balanced structural chromosome rearrangements, which can influence gene expression and the risk of unbalanced chromosome constitution in offspring. Many examples of inversion polymorphisms exist in human, affecting both heterochromatic regions and euchromatin. RESULTS: We describe a novel, 15 Mb long paracentric inversion, inv(21)(q21.1q22.11), affecting more than a third of human 21q. Despite of its length, the inversion cannot be detected using karyotyping due to similar band patterns on the normal and inverted chromosomes, and is therefore likely to escape attention. Its identification was aided by the repeated observation of the same pair of 150 kb long duplications present in cis on chromosome 21 in three Czech families subjected to microarray analysis. The finding prompted us to hypothesise that this co-occurrence of two remote duplications could be associated with an inversion of the intervening segment, and this speculation turned out to be right. The inversion was confirmed in a series of FISH experiments which also showed that the second copy of each of the duplications was always located at the opposite end of the inversion. The presence of the same pair of duplications in additional individuals reported in public databases indicates that the inversion may also be present in other populations. Three out of the total of about 4000 chromosomes 21 examined in our sample carried the duplications and were inverted, corresponding to carrier frequency of about 1/660. Although the breakpoints affect protein-coding genes, the occurrence of the inversion in normal parents and siblings of our patients and the occurrence of the duplications in unaffected controls in databases indicate that this rare variant is rather non-pathogenic. The inverted segment carried an identical shared haplotype in the three families studied. The haplotypes, however, diverged very rapidly in the flanking regions, possibly pointing to an ancient founder event at the origin of the inversion. CONCLUSIONS: The identification of inv(21)(q21.1q22.11) supports the notion that paracentric inversions are the most common form of chromosomal variation and that some of them may still remain undetected.