Human teneurin-1 is a direct target of the homeobox transcription factor EMX2 at a novel alternate promoter

<p>Abstract</p> <p>Background</p> <p>Teneurin-1 is a member of a family of type II transmembrane proteins conserved from <it>C.elegans </it>to vertebrates. Teneurin expression in vertebrates is best studied in mouse and chicken, where the four members teneurin-1 to -4 are predominantly expressed in the developing nervous system in area specific patterns. Based on their distinct, complementary expression a possible function in the establishment of proper connectivity in the brain was postulated. However, the transcription factors contributing to these distinctive expression patterns are largely unknown. Emx2 is a homeobox transcription factor, known to be important for area specification in the developing cortex. A study of Emx2 knock-out mice suggested a role of Emx2 in regulating patterned teneurin expression.</p> <p>Results</p> <p>5'RACE of human teneurin-1 revealed new alternative untranslated exons that are conserved in mouse and chicken. Closer analysis of the conserved region around the newly identified transcription start revealed promoter activity that was induced by EMX2. Mutation of a predicted homeobox binding site decreased the promoter activity in different reporter assays <it>in vitro </it>and <it>in vivo </it>in electroporated chick embryos. We show direct <it>in vivo </it>binding of EMX2 to the newly identified promoter element and finally confirm that the endogenous alternate transcript is specifically upregulated by EMX2.</p> <p>Conclusions</p> <p>We found that human teneurin-1 is directly regulated by EMX2 at a newly identified and conserved promoter region upstream of the published transcription start site, establishing teneurin-1 as the first human EMX2 target gene. We identify and characterize the EMX2 dependent promoter element of human teneurin-1.</p

New insights into the clinical and molecular spectrum of the novel CYFIP2-related neurodevelopmental disorder and impairment of the WRC-mediated actin dynamics

PURPOSE A few de novo missense variants in the cytoplasmic FMRP-interacting protein 2 (CYFIP2) gene have recently been described as a novel cause of severe intellectual disability, seizures, and hypotonia in 18 individuals, with p.Arg87 substitutions in the majority. METHODS We assembled data from 19 newly identified and all 18 previously published individuals with CYFIP2 variants. By structural modeling and investigation of WAVE-regulatory complex (WRC)-mediated actin polymerization in six patient fibroblast lines we assessed the impact of CYFIP2 variants on the WRC. RESULTS Sixteen of 19 individuals harbor two previously described and 11 novel (likely) disease-associated missense variants. We report p.Asp724 as second mutational hotspot (4/19 cases). Genotype-phenotype correlation confirms a consistently severe phenotype in p.Arg87 patients but a more variable phenotype in p.Asp724 and other substitutions. Three individuals with milder phenotypes carry putative loss-of-function variants, which remain of unclear pathogenicity. Structural modeling predicted missense variants to disturb interactions within the WRC or impair CYFIP2 stability. Consistent with its role in WRC-mediated actin polymerization we substantiate aberrant regulation of the actin cytoskeleton in patient fibroblasts. CONCLUSION Our study expands the clinical and molecular spectrum of CYFIP2-related neurodevelopmental disorder and provides evidence for aberrant WRC-mediated actin dynamics as contributing cellular pathomechanism

Constitutive androstane receptor 1 is constitutively bound to chromatin and ‘primed’ for transactivation in hepatocytes

The constitutive androstane receptor (CAR) is a xenobiotic sensor expressed in hepatocytes that activates genes involved in drug metabolism, lipid homeostasis, and cell proliferation. Much progress has been made in understanding the mechanism of activation of human CAR by drugs and xenobiotics. However, many aspects of the activation pathway remain to be elucidated. In this report, we have used viral constructs to express human CAR, its splice variants, and mutant CAR forms in hepatocytes from Car-/- mice in vitro and in vivo. We demonstrate CAR expression rescued the ability of Car-/- hepatocytes to respond to a wide range of CAR activators including phenobarbital. Additionally, two major splice isoforms of human CAR, CAR2 and CAR3, were inactive with almost all the agents tested. In contrast to the current model of CAR activation, ectopic CAR1 is constitutively localised in the nucleus and is loaded onto Cyp2b10 gene in the absence of an inducing agent. In studies to elucidate the role of threonine T38 in CAR regulation, we found that the T38D mutant was inactive even in the presence of CAR activators. However, the T38A mutant was activated by CAR inducers, showing that T38 is not essential for CAR activation. Also, using the inhibitor erlotinib, we could not confirm a role for the epidermal growth factor receptor in CAR regulation. Our data suggest that CAR is constitutively bound to gene regulatory regions and is regulated by exogenous agents through a mechanism which involves protein phosphorylation in the nucleus

Clinical and genetic characterization of individuals with predicted deleterious PHIP variants

Heterozygous deleterious variants in PHIP have been associated with behavioral problems, intellectual disability/developmental delay, obesity/overweight, and dysmorphic features (BIDOD syndrome). We report an additional 10 individuals with pleckstrin homology domain-interacting protein (PHIP)-predicted deleterious variants (four frameshift, three missense, two nonsense, and one splice site; six of which are confirmed de novo). The mutation spectrum is diverse, and there is no clustering of mutations across the protein. The clinical phenotype of these individuals is consistent with previous reports and includes behavioral problems, intellectual disability, developmental delay, hypotonia, and dysmorphic features. The additional individuals we report have a lower frequency of obesity than previous reports and a higher frequency of gastrointestinal problems, social deficits, and behavioral challenges. Characterizing additional individuals with diverse mutations longitudinally will provide better natural history data to assist with medical management and educational and behavioral support

ARTICLE Bi-allelic loss-of-function variants in PPFIBP1 cause a neurodevelopmental disorder with microcephaly, epilepsy, and periventricular calcifications

PPFIBP1 encodes for the liprin-β1 protein, which has been shown to play a role in neuronal outgrowth and synapse formation in Drosophila melanogaster. By exome and genome sequencing, we detected nine ultra-rare homozygous loss-of-function variants in 16 individuals from 12 unrelated families. The individuals presented with moderate to profound developmental delay, often refractory early-onset epilepsy, and progressive microcephaly. Further common clinical findings included muscular hyper- and hypotonia, spasticity, failure to thrive and short stature, feeding difficulties, impaired vision, and congenital heart defects. Neuroimaging revealed abnormalities of brain morphology with leukoencephalopathy, ventriculomegaly, cortical abnormalities, and intracranial periventricular calcifications as major features. In a fetus with intracranial calcifications, we identified a rare homozygous missense variant that by structural analysis was predicted to disturb the topology of the SAM domain region that is essential for protein-protein interaction. For further insight into the effects of PPFIBP1 loss of function, we performed automated behavioral phenotyping of a Caenorhabditis elegans PPFIBP1/hlb-1 knockout model, which revealed defects in spontaneous and light-induced behavior and confirmed resistance to the acetylcholinesterase inhibitor aldicarb, suggesting a defect in the neuronal presynaptic zone. In conclusion, we establish bi-allelic loss-of-function variants in PPFIBP1 as a cause of an autosomal recessive severe neurodevelopmental disorder with early-onset epilepsy, microcephaly, and periventricular calcifications

New Insights Into the Clinical and Molecular Spectrum of the Novel CYFIP2-Related Neurodevelopmental Disorder and Impairment of the WRC-Mediated Actin Dynamics

Purpose: A few de novo missense variants in the cytoplasmic FMRP-interacting protein 2 (CYFIP2) gene have recently been described as a novel cause of severe intellectual disability, seizures, and hypotonia in 18 individuals, with p.Arg87 substitutions in the majority. Methods: We assembled data from 19 newly identified and all 18 previously published individuals with CYFIP2 variants. By structural modeling and investigation of WAVE-regulatory complex (WRC)-mediated actin polymerization in six patient fibroblast lines we assessed the impact of CYFIP2 variants on the WRC. Results: Sixteen of 19 individuals harbor two previously described and 11 novel (likely) disease-associated missense variants. We report p.Asp724 as second mutational hotspot (4/19 cases). Genotype–phenotype correlation confirms a consistently severe phenotype in p.Arg87 patients but a more variable phenotype in p.Asp724 and other substitutions. Three individuals with milder phenotypes carry putative loss-of-function variants, which remain of unclear pathogenicity. Structural modeling predicted missense variants to disturb interactions within the WRC or impair CYFIP2 stability. Consistent with its role in WRC-mediated actin polymerization we substantiate aberrant regulation of the actin cytoskeleton in patient fibroblasts. Conclusion: Our study expands the clinical and molecular spectrum of CYFIP2-related neurodevelopmental disorder and provides evidence for aberrant WRC-mediated actin dynamics as contributing cellular pathomechanism

Xenobiotic CAR activators induce Dlk1-Dio3 locus non-coding RNA expression in mouse liver

Predicting the impact of human exposure to chemicals such as pharmaceuticals and agrochemicals requires the development of reliable and predictive biomarkers suitable for the detection of early events potentially leading to adverse outcomes. In particular, drug-induced non-genotoxic carcinogenesis (NGC) during preclinical development of novel therapeutics intended for chronic administration in humans is a major challenge for drug safety. We previously demonstrated Constitutive Androstane Receptor (CAR) and WNT signaling-dependent up-regulation of the pluripotency associated Dlk1-Dio3 imprinted gene cluster non-coding RNAs (ncRNAs) in the liver of mice treated with tumorpromoting doses of phenobarbital (PB). Here, to explore the sensitivity and the specificity of this candidate liver tumor promotion ncRNAs signature we compared phenotypic, transcriptional and proteomic data from wild-type, CAR/PXR double knock-out and CAR/PXR double humanized animals treated with tumor-promoting doses of PB or chlordane, both well-established CAR activators. We further investigated selected transcriptional profiles from mouse liver samples exposed to seven NGC compounds working through different mode of actions, overall suggesting CAR-activation specificity of the Dlk1-Dio3 long ncRNAs activation. We propose that Dlk1-Dio3 long ncRNAs up-regulation is an early CAR-activation dependent transcriptional signature during xenobiotic-induced mouse liver tumor promotion. This signature may further contribute mode of action-based ‘weight of evidence’ cancer risk assessment for xenobiotic-induced rodent liver tumors

Erratum: Evaluation of DNA Methylation Episignatures for Diagnosis and Phenotype Correlations in 42 Mendelian Neurodevelopmental Disorders (The American Journal of Human Genetics (2020) 106(3) (356–370), (S0002929720300197), (10.1016/j.ajhg.2020.01.019))

(The American Journal of Human Genetics 106, 356–370; March 5, 2020) In the version of this paper originally published, the underlying cause for Hunter McAlpine syndrome was incorrectly described in Table 1. The relevant description has been changed to read “Chr5q35-qter duplication involving NSD1” in the updated Table 1 reflected here. The authors apologize for this error

Solve-RD: systematic pan-European data sharing and collaborative analysis to solve rare diseases.

For the first time in Europe hundreds of rare disease (RD) experts team up to actively share and jointly analyse existing patient\u27s data. Solve-RD is a Horizon 2020-supported EU flagship project bringing together \u3e300 clinicians, scientists, and patient representatives of 51 sites from 15 countries. Solve-RD is built upon a core group of four European Reference Networks (ERNs; ERN-ITHACA, ERN-RND, ERN-Euro NMD, ERN-GENTURIS) which annually see more than 270,000 RD patients with respective pathologies. The main ambition is to solve unsolved rare diseases for which a molecular cause is not yet known. This is achieved through an innovative clinical research environment that introduces novel ways to organise expertise and data. Two major approaches are being pursued (i) massive data re-analysis of \u3e19,000 unsolved rare disease patients and (ii) novel combined -omics approaches. The minimum requirement to be eligible for the analysis activities is an inconclusive exome that can be shared with controlled access. The first preliminary data re-analysis has already diagnosed 255 cases form 8393 exomes/genome datasets. This unprecedented degree of collaboration focused on sharing of data and expertise shall identify many new disease genes and enable diagnosis of many so far undiagnosed patients from all over Europe