Loss of function of PGAP1 as a cause of severe encephalopathy identified by Whole Exome Sequencing: Lessons of the bioinformatics pipeline.

We evaluated a multiple consanguineous Turkish family with two children, a boy and a girl, affected by severe encephalopathy, hypotonia, microcephaly and retinal dystrophy by a combination of linkage analysis and Whole Exome Sequencing (WES). We analyzed the sequence data by two different bioinformatics pipelines which did not differ in overall processing strategy but involved differences in software used, minor allele frequency (MAF) thresholds and reference data sets, the usage of in-house control exomes and filter settings to prioritize called variants. Assuming autosomal recessive mode of inheritance, only homozygous variants present in both children were considered. The resulting variant lists differed partially (nine variants identified by both pipelines, ten variants by only one pipeline). Major reasons for this discrepancy were different filters for MAF and different variant prioritizations. Combining the variant lists with the results of linkage analysis and further prioritization by expression data and prediction tools, an intronic homozygous splice variant (c.1090-2A>G; IVS9-2A>G; p.?) in PGAP1 (Post-GPI Attachment To Proteins 1) was identified and validated by cDNA analysis. PGAP1 ensures the first step of maturation of GPI (glycosylphosphatidylinositol)-anchor proteins. Recently, a homozygous loss-of-function mutation in PGAP1 has been reported in one family with two children affected by a similar phenotype. The present report not only illustrates the possible influence of specific filtering settings on the results of WES but also confirms PGAP1 as a cause of severe encephalopathy.

Network-driven discovery yields new insight into Shox2-dependent cardiac rhythm control.

The homeodomain transcription factor SHOX2 is involved in the development and function of the heart's primary pacemaker, the sinoatrial node (SAN), and has been associated with cardiac conduction-related diseases such as atrial fibrillation and sinus node dysfunction. To shed light on Shox2-dependent genetic processes involved in these diseases, we established a murine embryonic stem cell (ESC) cardiac differentiation model to investigate Shox2 pathways in SAN-like cardiomyocytes. Differential RNA-seq-based expression profiling of Shox2+/+ and Shox2-/- ESCs revealed 94 dysregulated transcripts in Shox2-/- ESC-derived SAN-like cells. Of these, 15 putative Shox2 target genes were selected for further validation based on comparative expression analysis with SAN- and right atria-enriched genes. Network-based analyses, integrating data from the Mouse Organogenesis Cell Atlas and the Ingenuity pathways, as well as validation in mouse and zebrafish models confirmed a regulatory role for the novel identified Shox2 target genes including Cav1, Fkbp10, Igfbp5, Mcf2l and Nr2f2. Our results indicate that genetic networks involving SHOX2 may contribute to conduction traits through the regulation of these genes.

Isochromosome 1q as an early genetic event in a child with intracranial ependymoma characterized by molecular cytogenetics.

Data concerning cytogenetic features of childhood ependymoma are rare. In this article, a gain of 1q was identified as the sole alteration in a primary childhood infratentorial ependymoma by comparative genomic hybridization (CGH). A recurrence of this brain tumor was studied using multiplex-fluorescence in situ hybridization (M-FISH) in addition to CGH and G-banding analysis. In accordance with the primary tumor, a gain of 1q corresponding to an isochromosome 1q was observed indicating an early event in the tumor development. Furthermore, M-FISH classified several other rearranged chromosomes including 6q and 17p that have previously been found to be involved in the development and progression of childhood ependymoma.

Comparative genomic hybridization in childhood acute lymphoblastic leukemia: correlation with interphase cytogenetics and loss of heterozygosity analysis.

We used comparative genomic hybridization (CGH) to study DNA copy number changes in 71 children with acute lymphoblastic leukemia (ALL) including 50 B-lineage and 21 T-ALLs. Forty-two patients (59%) showed genomic imbalances whereby gains were more frequently observed than losses (127 vs. 29). Gains most commonly affected the entire chromosomes 21 and 10 (19.7% each), 6, 14, 18, X (15.5% each), 17 (14.1%) and 4 (11.3%). Highly hyperdiploid karyotypes (chromosome number >50) occurred more frequently in B-lineage than in T-lineage ALL (24% vs. 4.8%). In both cell lineages deletions were mainly detected on 9p (14.1%) and 12p (8.4%), and on 6q in T-lineage ALL (4.2%). These findings were compared with loss of heterozygosity (LOH) of 6q, 9p, 11q, and 12p previously performed in 56 of the 71 patients. Among 54 sites of LOH, CGH revealed losses of the respective chromosome arms in 17 LOH-positive regions (31.5%). G-banding analysis and interphase cytogenetics with subregional probes for 14 loci confirmed the presence of genomic imbalances as detected by CGH. We, therefore, conclude that, in the absence of cytogenetic data, CGH represents a suitable method for identifying hyperdiploid karyotypes as well as prognostically relevant deletions in ALL patients.

Nuclear architecture and the induction of chromosomal aberrations.

Progress in fluorescence in situ hybridization, three dimensional microscopy and image analysis has provided the means to study the three-dimensional structure and distribution of chromosome territories within the cell nucleus. In this contribution, we summarize the present state of knowledge of the territorial organization of interphase chromosomes and their topological relationships with other macromolecular domains in the human cell nucleus, and present data from computer simulations of chromosome territory distributions. On this basis, we discuss models of chromosome territory and nuclear architecture and topological consequences for the formation of chromosome exchanges.

Informatics united: exemplary studies combining medical informatics, neuroinformatics and bioinformatics.

OBJECTIVES: Medical informatics, neuroinformatics and bioinformatics provide a wide spectrum of research. Here, we show the great potential of synergies between these research areas on the basis of four exemplary studies where techniques are transferred from one of the disciplines to the other. METHODS: Reviewing and analyzing exemplary and specific projects at the intersection of medical informatics, neuroinformatics, and bioinformatics from our experience in an interdisciplinary research group. RESULTS: Synergy emerges when techniques and solutions from medical informatics, bioinformatics, or neuroinformatics are successfully applied in one of the other disciplines. Synergy was found in 1. the modeling of neurophysiological systems for medical therapy development, 2. the use of image processing techniques from medical computer vision for the analysis of the dynamics of cell nuclei, and 3. the application of neuroinformatics tools for data mining in bioinformatics and as classifiers in clinical oncology. CONCLUSIONS: Each of the three different disciplines have delivered technologies that are readily applicable in the other disciplines. The mutual transfer of knowledge and techniques proved to increase efficiency and accuracy in a manifold of applications. In particular, we expect that clinical decision support systems based on techniques derived from neuro- and bioinformatics have the potential to improve medical diagnostics and will finally lead to a personalized delivery of healthcare.

Multiplex FISH telomere integrity assay identifies an unbalanced cryptic translocation der(5)t(3;5)(q27;p15.3) in a family with three mentally retarded individuals.

Cryptic rearrangements involving the terminal regions of chromosomes are suspected to be the cause of idiopathic mental retardation in a significant number of cases. This finding highlights the necessity of a primary screening test for such chromosome aberrations. Here we present a multiplex fluorescence in situ hybridization telomere integrity assay which allows the detection of submicroscopic aberrations in the telomeric regions of all chromosomes. This novel approach identified an unbalanced cryptic translocation der(5)t(3;5)(q27;p15.3) in a family with three cases of unexplained mental retardation and dysmorphic features. The symptoms of the patients represent neither the classical dup(3q)- nor cri du chat syndrome, although all affected individuals demonstrate several features of both syndromes. The identification of two balanced translocation carriers emphasizes the significance of the telomere integrity assay for genetic counseling and prenatal diagnosis.