SHOX triggers the lysosomal pathway of apoptosis via oxidative stress.

The SHOX gene encodes for a transcription factor important for normal bone development. Mutations in the gene are associated with idiopathic short stature and are responsible for the growth failure and skeletal defects found in the majority of patients with Léri-Weill dyschondrosteosis (LWD) and Langer mesomelic dysplasia. SHOX is expressed in growth plate chondrocytes where it is supposed to modulate the proliferation, differentiation and cell death of these cells. Supporting this hypothesis, in vitro studies have shown that SHOX expression induces cell cycle arrest and apoptosis in both transformed and primary cells. In this study, we further characterized the cell death mechanisms triggered by SHOX and compared them with the effects induced by one clinically relevant mutant form of SHOX, detected in LWD patients (SHOX R153L) and a SHOX C-terminally truncated version (L185X). We show that SHOX expression in U2OS osteosarcoma cells leads to oxidative stress that, in turn, induces lysosomal membrane rupture with release of active cathepsin B to the cytosol and subsequent activation of the intrinsic apoptotic pathway characterized by mitochondrial membrane permeabilization and caspase activation. Importantly, cells expressing SHOX R153L or L185X did not display any of these features. Given the fact that many of the events observed in SHOX-expressing cells also characterize the complex cell death process occurring in the growth plate during endochondral ossification, our findings further support the hypothesis that SHOX may play a central role in the regulation of the cell death pathways activated during long bone development.

FGFR3 is a target of the homeobox transcription factor SHOX in limb development.

The short stature homeobox gene SHOX encodes a transcription factor which is important for normal limb development. In humans, SHOX deficiency has been associated with various short stature syndromes including Leri-Weill dyschondrosteosis (LWD), Langer mesomelic dysplasia and Turner syndrome as well as non-syndromic idiopathic short stature. A common feature of these syndromes is disproportionate short stature with a particular shortening of the forearms and lower legs. In our studies employing microarray analyses and cell culture experiments, we revealed a strong positive effect of SHOX on the expression of the fibroblast growth factor receptor gene FGFR3, another well-known factor for limb development. Luciferase reporter gene assays show that SHOX activates the extended FGFR3 promoter, and results from chromatin immunoprecipitation (ChIP)-sequencing, ChIP and electrophoretic mobility shift assay experiments suggest a direct binding of SHOX to multiple upstream sequences of FGFR3. To further investigate these regulations in a cellular system for limb development, the effect of viral overexpression of Shox in limb bud derived chicken micromass cultures was tested. We found that Fgfr3 was negatively regulated by Shox, as demonstrated by quantitative real-time polymerase chain reaction and in situ hybridization. This repressive effect might explain the almost mutually exclusive expression patterns of Fgfr3 and Shox in embryonic chicken limbs. A negative regulation that occurs mainly in the mesomelic segments, a region where SHOX is known to be strongly expressed, offers a possible explanation for the phenotypes seen in patients with FGFR3 (e.g. achondroplasia) and SHOX defects (e.g. LWD). In summary, these data present a link between two frequent short stature phenotypes.

Controlling my genome with my smartphone: first clinical experiences of the PROMISE system.

BACKGROUND: The development of Precision Medicine strategies requires high-dimensional phenotypic and genomic data, both of which are highly privacy-sensitive data types. Conventional data management systems lack the capabilities to sufficiently handle the expected large quantities of such sensitive data in a secure manner. PROMISE is a genetic data management concept that implements a highly secure platform for data exchange while preserving patient interests, privacy, and autonomy. METHODS: The concept of PROMISE to democratize genetic data was developed by an interdisciplinary team. It integrates a sophisticated cryptographic concept that allows only the patient to grant selective access to defined parts of his genetic information with single DNA base-pair resolution cryptography. The PROMISE system was developed for research purposes to evaluate the concept in a pilot study with nineteen cardiomyopathy patients undergoing genotyping, questionnaires, and longitudinal follow-up. RESULTS: The safety of genetic data was very important to 79%, and patients generally regarded the data as highly sensitive. More than half the patients reported that their attitude towards the handling of genetic data has changed after using the PROMISE app for 4 months (median). The patients reported higher confidence in data security and willingness to share their data with commercial third parties, including pharmaceutical companies (increase from 5 to 32%). CONCLUSION: PROMISE democratizes genomic data by a transparent, secure, and patient-centric approach. This clinical pilot study evaluating a genetic data infrastructure is unique and shows that patient's acceptance of data sharing can be increased by patient-centric decision-making.

C3-halogenation of cytisine generates potent and efficacious nicotinic receptor agonists.

Neuronal nicotinic acetylcholine receptors subserve predominantly modulatory roles in the brain, making them attractive therapeutic targets. Natural products provide key leads in the quest for nicotinic receptor subtype-selective compounds. Cytisine, found in Leguminosae spp., binds with high affinity to alpha4beta2* nicotinic receptors. We have compared the effect of C3 and C5 halogenation of cytisine and methylcytisine (MCy) on their interaction with native rat nicotinic receptors. 3-Bromocytisine (3-BrCy) and 3-iodocytisine (3-ICy) exhibited increased binding affinity (especially at alpha7 nicotinic receptors; Ki approximately 0.1 microM) and functional potency, whereas C5-halogenation was detrimental. 3-BrCy and 3-ICy were more potent than cytisine at evoking [3H]dopamine release from striatal slices (EC50 approximately 11 nM), [3H]noradrenaline release from hippocampal slices (EC50 approximately 250 nM), increases in intracellular Ca2+ in PC12 cells and inward currents in Xenopus oocytes expressing human alpha3beta4 nicotinic receptor (EC50 approximately 2 microM). These compounds were also more efficacious than cytisine. C3-halogenation of cytisine is proposed to stabilize the open conformation of the nicotinic receptor but does not enhance subtype selectivity.

Percepción de riesgo ante las inundaciones en personas que habitan en zonas vulnerables de Lima, Perú / Risk perception of people living in vulnerable zones of Lima (Peru) during floods

Introducción: En el último año, el Perú experimentó un fenómeno climático denominado el Niño costero. Este generó lluvias, huaicos e inundaciones, afectando a miles de personas y ocasionando la declaración en emergencia sanitaria del país. A pesar de la ocurrencia de fenómenos similares en las zonas afectadas, estas suelen ser ocupadas como lugares de residencia. Objetivo: Identificar las características de la percepción de riesgo ante las inundaciones en personas que habitan en zonas vulnerables. Método: Investigación empírica cualitativa desarrollada en el año 2017. Se aplicó la técnica de la entrevista semiestructurada a nueve personas residentes en cuatro distritos de Lima (Perú) propensos a inundaciones. La información recolectada se analizó con el apoyo del software para el análisis cualitativo de datos Atlas.ti v.7.0, utilizando elementos de la Teoría Fundamentada para desarrollar los procesos de codificación y categorización. Resultados: Se encontró que los participantes no poseen percepción de riesgo ante la amenaza de las inundaciones. Conciben a las inundaciones como una amenaza cuando experimentan sus consecuencias. Conclusiones: Se requieren desarrollar investigaciones de carácter interdisciplinario que permitan generar evidencia para el desarrollo de intervenciones orientadas a la construcción de una percepción de riesgo coherente con las consecuencias e impacto de las inundaciones(AU)

Introduction: In the last year, Peru experienced a climatic phenomenon called Niño Costero. This generated rains, alluviums and floods affecting thousands of people and causing the declaration of health emergency in the country. In spite of the occurrence of similar phenomena in the affected areas, these are usually occupied as places to live. Objective: To identify the characteristics of risk's perception during floods in people who live in vulnerable zones. Method: A qualitative empirical research was developed in the year 2017. The semi-structured interview technique was conducted in nine people living in four districts of Lima (Peru) whom were susceptible to flooding. The information collected was analyzed with the support of Atlas.ti v. 7.0., software for the qualitative data analysis, using elements of the Grounded Theory to develop the coding and categorization processes. Results: It was found that the participants do not elaborate a perception of risk before the threat of floods. They envisage the floods as a threat when experience their consequences. Conclusions: It is necessary to implement interdisciplinary researches that allow to generate evidence for the development of interventions aimed to the creation of a risk's perception that will be coherent with the consequences and the impact of floods(AU)

Height matters-from monogenic disorders to normal variation.

Height is a classic polygenic quantitative trait with a high level of heritability. As it is a simple and stable parameter to measure, height is a model for both common, complex disorders and monogenic, Mendelian disease. In this Review, we examine height from the perspective of monogenic and complex genetics and discuss the lessons learned so far. We explore several examples of rare sequence variants with large effects on height and compare these variants to the common variants identified in genome-wide association studies that have small effects on height. We discuss how copy number changes or genetic interactions might contribute to the unidentified aspects of the heritability of height. We also ask whether information derived from genome-wide association studies on specific loci in the vicinity of genes can be used for further research in clinical paediatric endocrinology. Furthermore, we address key challenges that remain for gene discovery and for the transition of moving from genomic localization to mechanistic insights, with an emphasis on using next-generation sequencing to identify causative variants of people at the extremes of height distribution.

The homeobox transcription factor HOXA9 is a regulator of SHOX in U2OS cells and chicken micromass cultures.

The homeobox gene SHOX encodes for a transcription factor that plays an important role during limb development. Mutations or deletions of SHOX in humans cause short stature in Turner, Langer and Leri-Weill syndrome as well as idiopathic short stature. During embryonic development, SHOX is expressed in a complex spatio-temporal pattern that requires the presence of specific regulatory mechanisms. Up to now, it was known that SHOX is regulated by two upstream promoters and several enhancers on either side of the gene, but no regulators have been identified that can activate or repress the transcription of SHOX by binding to these regulatory elements. We have now identified the homeodomain protein HOXA9 as a positive regulator of SHOX expression in U2OS cells. Using luciferase assays, chromatin immunoprecipitation and electrophoretic mobility shift assays, we could narrow down the HOXA9 binding site to two AT-rich sequences of 31 bp within the SHOX promoter 2. Virus-induced Hoxa9 overexpression in a chicken micromass model validated the regulation of Shox by Hoxa9 (negative regulation). As Hoxa9 and Shox are both expressed in overlapping regions of the developing limb buds, a regulatory relationship of Hoxa9 and Shox during the process of limb development can be assumed.

Alternative splicing and nonsense-mediated RNA decay contribute to the regulation of SHOX expression.

The human SHOX gene is composed of seven exons and encodes a paired-related homeodomain transcription factor. SHOX mutations or deletions have been associated with different short stature syndromes implying a role in growth and bone formation. During development, SHOX is expressed in a highly specific spatiotemporal expression pattern, the underlying regulatory mechanisms of which remain largely unknown. We have analysed SHOX expression in diverse embryonic, fetal and adult human tissues and detected expression in many tissues that were not known to express SHOX before, e.g. distinct brain regions. By using RT-PCR and comparing the results with RNA-Seq data, we have identified four novel exons (exon 2a, 7-1, 7-2 and 7-3) contributing to different SHOX isoforms, and also established an expression profile for the emerging new SHOX isoforms. Interestingly, we found the exon 7 variants to be exclusively expressed in fetal neural tissues, which could argue for a specific role of these variants during brain development. A bioinformatical analysis of the three novel 3'UTR exons yielded insights into the putative role of the different 3'UTRs as targets for miRNA binding. Functional analysis revealed that inclusion of exon 2a leads to nonsense-mediated RNA decay altering SHOX expression in a tissue and time specific manner. In conclusion, SHOX expression is regulated by different mechanisms and alternative splicing coupled with nonsense-mediated RNA decay constitutes a further component that can be used to fine tune the SHOX expression level.

Enhancer elements upstream of the SHOX gene are active in the developing limb.

Léri-Weill Dyschondrosteosis (LWD) is a dominant skeletal disorder characterized by short stature and distinct bone anomalies. SHOX gene mutations and deletions of regulatory elements downstream of SHOX resulting in haploinsufficiency have been found in patients with LWD. SHOX encodes a homeodomain transcription factor and is known to be expressed in the developing limb. We have now analyzed the regulatory significance of the region upstream of the SHOX gene. By comparative genomic analyses, we identified several conserved non-coding elements, which subsequently were tested in an in ovo enhancer assay in both chicken limb bud and cornea, where SHOX is also expressed. In this assay, we found three enhancers to be active in the developing chicken limb, but none were functional in the developing cornea. A screening of 60 LWD patients with an intact SHOX coding and downstream region did not yield any deletion of the upstream enhancer region. Thus, we speculate that SHOX upstream deletions occur at a lower frequency because of the structural organization of this genomic region and/or that SHOX upstream deletions may cause a phenotype that differs from the one observed in LWD.