Novel Missense Mutation A789V in IQSEC2 underlies X-Linked intellectual disability in the MRX78 family

Disease gene discovery in neurodevelopmental disorders, including X-linked intellectual disability (XLID) has recently been accelerated by next-generation DNA sequencing approaches. To date, more than 100 human X chromosome genes involved in neuronal signaling pathways and networks implicated in cognitive function have been identified. Despite these advances, the mutations underlying disease in a large number of XLID families remained unresolved. We report the resolution of MRX78, a large family with six affected males and seven affected females, showing X-linked inheritance. Although a previous linkage study had mapped the locus to the short arm of chromosome X (Xp11.4-p11.23), this region contained too many candidate genes to be analyzed using conventional approaches. However, our X-chromosome exome resequencing, bioinformatics analysis and inheritance testing revealed a missense mutation (c.C2366T, p.A789V) in IQSEC2, encoding a neuronal GDP-GTP exchange factor for Arf family GTPases (ArfGEF) previously implicated in XLID. Molecular modeling of IQSEC2 revealed that the A789V substitution results in the insertion of a larger side-chain into a hydrophobic pocket in the catalytic Sec7 domain of IQSEC2. The A789V change is predicted to result in numerous clashes with adjacent amino acids and disruption of local folding of the Sec7 domain. Consistent with this finding, functional assays revealed that recombinant IQSEC2A789V was not able to catalyze GDP-GTP exchange on Arf6 as efficiently as wild-type IQSEC2. Taken together, these results strongly suggest that the A789V mutation in IQSEC2 is the underlying cause of XLID in the MRX78 family

DNA damage in subpopulations of human lymphocytes irradiated with doses in the range of 0-1 Gy of X-radiation

We compared three methods usually applied in biological dosimetry for estimation of radiation-induced DNA damage in human T and B lymphocytes: alkaline comet assay, micronucleus (MN) test and formation of histone gamma-H2AX foci. Human peripheral blood lymphocytes were fractionated using T cells and B cells isolation kits. Cells were irradiated with doses in the range of 0-1 Gy of X-rays. Induction of DNA damage was assessed by the standard alkaline comet assay, MN test and histone gammaH2AX foci immunofluorescence assay. Notwithstanding different end-points measured by the applied methods, all tests revealed a similar induction of DNA damage in B lymphocytes as compared with T lymphocytes. The results indicated that all three tests detect DNA damage with similar sensitivity, the lowest dose being approximately 0.3 Gy. The difference between irradiated and control cells was expressed as the ratio of the value obtained for irradiated cells (1 Gy) to that for control cells. The highest ratio was obtained for formation of gammaH2AX foci and was 6.2 for T and 13.8 for B lymphocytes, whereas those for comet assay and micronucleus test were 3.5; 3.6 and 5.6; 4.8, respectively

Analysis of clinical symptoms and selected hematological indices in hospitalized children with Ascaris lumbricoides infection from the northeastern region of Poland

Ascariasis is the most common soil-transmitted helminth infection in the world. The objective of this study was to analyze the clinical symptoms and selected hematological indices of ascariasis in hospitalized children from the northeastern region of Poland. Patients in the Pediatric Ward hospitalized in the Regional Hospital in Dąbrowa Białostocka in the period of 2005–2007 were included in this retrospective study. The intestinal stage of ascariasis was diagnosed on the basis of positive coprological survey performed using the decantation technique. A total of 938 patients were included in the study, 1801 stool samples were evaluated, and A. lumbricoides-positive tests were obtained from 252 children. Ascaris-positive young children (3 yrs) accounted for 3.0% of all hospitalized children, Ascarispositive preschool-aged children (4–7 yrs) accounted for 8.1% and school-aged children (8–18 yrs) for 15.8%. Seasonal patterns were observed in the prevalence of A. lumbricoides (maximum in August–December). There was no relationship between BMI z-score, hemoglobin levels and prevalence of infection with Ascaris lumbricoides. Significant predictors of intestinal stage ascariasis in a multivariate logistic regression model were: abdominal pain as a reason for hospital admission (OR–2.19; 95%CI 1.62–2.95; p<0.001) and age from 4 to 7 years (OR–2.0; 95%CI 1.41–2.80; p<0.001). The prevalence rate of ascariasis was not higher in the group of patients with atopic diseases (bronchial asthma, allergic rhinitis, atopic dermatitis) and co-existing ascariasis did not affect the eosinophil counts in the peripheral blood. Ascariasis is still a current pediatric clinical problem characterized by non-specific clinical manifestations, which should be taken into consideration in the differential diagnosis of children’s diseases

Minigene-Based Splice Assays Reveal the Effect of Non-Canonical Splice Site Variants in USH2A

Non-canonical splice site variants are increasingly recognized as a relevant cause of the USH2A-associated diseases, non-syndromic autosomal recessive retinitis pigmentosa and Usher syndrome type 2. Many non-canonical splice site variants have been reported in public databases, but an effect on pre-mRNA splicing has only been functionally verified for a subset of these variants. In this study, we aimed to extend the knowledge regarding splicing events by assessing a selected set of USH2A non-canonical splice site variants and to study their potential pathogenicity. Eleven non-canonical splice site variants were selected based on four splice prediction tools. Ten different USH2A constructs were generated and minigene splice assays were performed in HEK293T cells. An effect on pre-mRNA splicing was observed for all 11 variants. Various events, such as exon skipping, dual exon skipping and partial exon skipping were observed and eight of the tested variants had a full effect on splicing as no conventionally spliced mRNA was detected. We demonstrated that non-canonical splice site variants in USH2A are an important contributor to the genetic etiology of the associated disorders. This type of variant generally should not be neglected in genetic screening, both in USH2A-associated disease as well as other hereditary disorders. In addition, cases with these specific variants may now receive a conclusive genetic diagnosis