Whole-genome DNA/RNA sequencing identifies truncating mutations in RBCK1 in a novel Mendelian disease with neuromuscular and cardiac involvement

Background: Whole-exome sequencing has identified the causes of several Mendelian diseases by analyzing multiple unrelated cases, but it is more challenging to resolve the cause of extremely rare and suspected Mendelian diseases from individual families. We identified a family quartet with two children, both affected with a previously unreported disease, characterized by progressive muscular weakness and cardiomyopathy, with normal intelligence. During the course of the study, we identified one additional unrelated patient with a comparable phenotype. Methods: We performed whole-genome sequencing (Complete Genomics platform), whole-exome sequencing (Agilent SureSelect exon capture and Illumina Genome Analyzer II platform), SNP genotyping (Illumina HumanHap550 SNP array) and Sanger sequencing on blood samples, as well as RNA-Seq (Illumina HiSeq platform) on transformed lymphoblastoid cell lines. Results: From whole-genome sequence data, we identified RBCK1, a gene encoding an E3 ubiquitin-protein ligase, as the most likely candidate gene, with two protein-truncating mutations in probands in the first family. However, exome data failed to nominate RBCK1 as a candidate gene, due to poor regional coverage. Sanger sequencing identified a private homozygous splice variant in RBCK1 in the proband in the second family, yet SNP genotyping revealed a 1.2Mb copy-neutral region of homozygosity covering RBCK1. RNA-Seq confirmed aberrant splicing of RBCK1 transcripts, resulting in truncated protein products. Conclusions: While the exact mechanism by which these mutations cause disease is unknown, our study represents an example of how the combined use of whole-genome DNA and RNA sequencing can identify a disease-predisposing gene for a novel and extremely rare Mendelian disease

Using synthetic biological parts and microbioreactors to explore the protein expression characteristics of Escherichia coli

Synthetic biology has developed numerous parts for the precise control of protein expression. However, relatively little is known about the burden these place on a host, or their reliability under varying environmental conditions. To address this, we made use of synthetic transcriptional and translational elements to create a combinatorial library of constructs that modulated expression strength of a green fluorescent protein. Combining this library with a microbioreactor platform, we were able to perform a detailed large-scale assessment of transient expression and growth characteristics of two <i>Escherichia coli</i> strains across several temperatures. This revealed significant differences in the robustness of both strains to differing types of protein expression, and a complex response of transcriptional and translational elements to differing temperatures. This study supports the development of reliable synthetic biological systems capable of working across different hosts and environmental contexts. Plasmids developed during this work have been made publicly available to act as a reference set for future research

Geologically recent areas as one key target for identifying active volcanism on Venus

The recently selected NASA VERITAS and DAVINCI missions, the ESA EnVision, the Roscosmos Venera-D will open a new era in the exploration of Venus. One of the key targets of the future orbiting and in situ investigations of Venus is the identification of volcanically active areas on the planet. The study of the areas characterized by recent or ongoing volcano-tectonic activity can inform us on how volcanism and tectonism are currently evolving on Venus. Following this key target, Brossier et al. (2022, https://doi.org/10.1029/2022GL099765) extend the successful approach and methodology used by previous works to Ganis Chasma in Atla Regio. Here we comment on the main results published in Brossier et al. (2022, https://doi.org/10.1029/2022GL099765) and discuss the important implications of their work for the future orbiting and in situ investigation of Venus. Their results add further lines of evidence indicating possibly recent volcanism on Venus

Test of the photon detection system for the LHCb RICH Upgrade in a charged particle beam

The LHCb detector will be upgraded to make more efficient use of the available luminosity at the LHC in Run III and extend its potential for discovery. The Ring Imaging Cherenkov detectors are key components of the LHCb detector for particle identification. In this paper we describe the setup and the results of tests in a charged particle beam, carried out to assess prototypes of the upgraded opto-electronic chain from the Multi-Anode PMT photosensor to the readout and data acquisition system.Comment: 25 pages, 22 figure

Effect of four plant species on soil 15N-access and herbage yield in temporary agricultural grasslands

Positive plant diversity-productivity relationships have been reported for experimental semi-natural grasslands (Cardinale et al. 2006; Hector et al. 1999; Tilman et al. 1996) as well as temporary agricultural grasslands (Frankow-Lindberg et al. 2009; Kirwan et al. 2007; Nyfeler et al. 2009; Picasso et al. 2008). Generally, these relationships are explained, on the one hand, by niche differentiation and facilitation (Hector et al. 2002; Tilman et al. 2002) and, on the other hand, by greater probability of including a highly productive plant species in high diversity plots (Huston 1997). Both explanations accept that diversity is significant because species differ in characteristics, such as root architecture, nutrient acquisition and water use efficiency, to name a few, resulting in composition and diversity being important for improved productivity and resource use (Naeem et al. 1994; Tilman et al. 2002). Plant diversity is generally low in temporary agricultural grasslands grown for ruminant fodder production. Grass in pure stands is common, but requires high nitrogen (N) inputs. In terms of N input, two-species grass-legume mixtures are more sustainable than grass in pure stands and consequently dominate low N input grasslands (Crews and Peoples 2004; Nyfeler et al. 2009; Nyfeler et al. 2011). In temperate grasslands, N is often the limiting factor for productivity (Whitehead 1995). Plant available soil N is generally concentrated in the upper soil layers, but may leach to deeper layers, especially in grasslands that include legumes (Scherer-Lorenzen et al. 2003) and under conditions with surplus precipitation (Thorup-Kristensen 2006). To improve soil N use efficiency in temporary grasslands, we propose the addition of deep-rooting plant species to a mixture of perennial ryegrass and white clover, which are the most widespread forage plant species in temporary grasslands in a temperate climate (Moore 2003). Perennial ryegrass and white clover possess relatively shallow root systems (Kutschera and Lichtenegger 1982; Kutschera and Lichtenegger 1992) with effective rooting depths of <0.7 m on a silt loamy site (Pollock and Mead 2008). Grassland species, such as lucerne and chicory, grow their tap-roots into deep soil layers and exploit soil nutrients and water in soil layers that the commonly grown shallow-rooting grassland species cannot reach (Braun et al. 2010; Skinner 2008). Chicory grown as a catch crop after barley reduced the inorganic soil N down to 2.5 m depth during the growing season, while perennial ryegrass affected the inorganic soil N only down to 1 m depth (Thorup-Kristensen 2006). Further, on a Wakanui silt loam in New Zealand chicory extracted water down to 1.9 m and lucerne down to 2.3 m soil depth, which resulted in greater herbage yields compared with a perennial ryegrass-white clover mixture, especially for dryland plots (Brown et al. 2005). There is little information on both the ability of deep- and shallow-rooting grassland species to access soil N from different vertical soil layers and the relation of soil N-access and herbage yield in temporary agricultural grasslands. Therefore, the objective of the present work was to test the hypotheses 1) that a mixture comprising both shallow- and deep-rooting plant species has greater herbage yields than a shallow-rooting binary mixture and pure stands, 2) that deep-rooting plant species (chicory and lucerne) are superior in accessing soil N from 1.2 m soil depth compared with shallow-rooting plant species, 3) that shallow-rooting plant species (perennial ryegrass and white clover) are superior in accessing soil N from 0.4 m soil depth compared with deep-rooting plant species, 4) that a mixture of deep- and shallow-rooting plant species has greater access to soil N from three soil layers compared with a shallow-rooting two-species mixture and that 5) the leguminous grassland plants, lucerne and white clover, have a strong impact on grassland N acquisition, because of their ability to derive N from the soil and the atmosphere

Tertiary-Treated Municipal Wastewater is a Significant Point Source of Antibiotic Resistance Genes Into Duluth-Superior Harbor

In this study, the impact of tertiary-treated municipal wastewater on the quantity of several antibiotic resistance determinants in Duluth-Superior Harbor was investigated by collecting surface water and sediment samples from 13 locations in Duluth-Superior Harbor, the St. Louis River, and Lake Superior. Quantitative PCR (qPCR) was used to target three different genes encoding resistance to tetracycline (tet(A), tet(X), and tet(W)), the gene encoding the integrase of class 1 integrons (intI1), and total bacterial abundance (16S rRNA genes) as well as total and human fecal contamination levels (16S rRNA genes specific to the genus Bacteroides). The quantities of tet(A), tet(X), tet(W), intI1, total Bacteroides, and human-specific Bacteroides were typically 20-fold higher in the tertiary-treated wastewater than in nearby surface water samples. In contrast, the quantities of these genes in the St. Louis River and Lake Superior were typically below detection. Analysis of sequences of tet(W) gene fragments from four different samples collected throughout the study site supported the conclusion that tertiary-treated municipal wastewater is a point source of resistance genes into Duluth-Superior Harbor. This study demonstrates that the discharge of exceptionally treated municipal wastewater can have a statistically significant effect on the quantities of antibiotic resistance genes in otherwise pristine surface waters

Current and novel therapeutic opportunities for systemic therapy in biliary cancer

none24Biliary tract cancers (BTCs) are a group of rare and aggressive malignancies that arise in the biliary tree within and outside the liver. Beyond surgical resection, which is beneficial for only a small proportion of patients, current strategies for treating patients with BTCs include chemotherapy, as a single agent or combination regimens, in the adjuvant and palliative setting. Increased characterisation of the molecular landscape of these tumours has facilitated the identification of molecular vulnerabilities, such as IDH mutations and FGFR fusions, that can be exploited for the treatment of BTC patients. Beyond targeted therapies, active research avenues explore the development of novel therapeutics that target the crosstalk between cancer and stroma, the cellular pathways involved in the regulation of cell death, the chemoresistance phenotype and the dysregulation of RNA. In this review, we discuss the therapeutic opportunities currently available in the management of BTC patients, and explore the strategies that can support the implementation of precision oncology in BTCs, including novel molecular targets, liquid biopsies and patient-derived predictive tools.openMarin J.J.G.; Prete M.G.; Lamarca A.; Tavolari S.; Landa-Magdalena A.; Brandi G.; Segatto O.; Vogel A.; Macias R.I.R.; Rodrigues P.M.; Casta A.L.; Mertens J.; Rodrigues C.M.P.; Fernandez-Barrena M.G.; Da Silva Ruivo A.; Marzioni M.; Mentrasti G.; Acedo P.; Munoz-Garrido P.; Cardinale V.; Banales J.M.; Valle J.W.; Bridgewater J.; Braconi C.Marin, J. J. G.; Prete, M. G.; Lamarca, A.; Tavolari, S.; Landa-Magdalena, A.; Brandi, G.; Segatto, O.; Vogel, A.; Macias, R. I. R.; Rodrigues, P. M.; Casta, A. L.; Mertens, J.; Rodrigues, C. M. P.; Fernandez-Barrena, M. G.; Da Silva Ruivo, A.; Marzioni, M.; Mentrasti, G.; Acedo, P.; Munoz-Garrido, P.; Cardinale, V.; Banales, J. M.; Valle, J. W.; Bridgewater, J.; Braconi, C

IL-17 induces an expanded range of downstream genes in reconstituted human epidermis model

Background: IL-17 is the defining cytokine of the Th17, Tc17, and γδ T cell populations that plays a critical role in mediating inflammation and autoimmunity. Psoriasis vulgaris is an inflammatory skin disease mediated by Th1 and Th17 cytokines with relevant contributions of IFN-γ, TNF-α, and IL-17. Despite the pivotal role IL-17 plays in psoriasis, and in contrast to the other key mediators involved in the psoriasis cytokine cascade that are capable of inducing broad effects on keratinocytes, IL-17 was demonstrated to regulate the expression of a limited number of genes in monolayer keratinocytes cultured in vitro. Methodology/Principal Findings: Given the clinical efficacy of anti-IL-17 agents is associated with an impressive reduction in a large set of inflammatory genes, we sought a full-thickness skin model that more closely resemble in vivo epidermal architecture. Using a reconstructed human epidermis (RHE), IL-17 was able to upregulate 419 gene probes and downregulate 216 gene probes. As possible explanation for the increased gene induction in the RHE model is that C/CAATenhancer- binding proteins (C/EBP) -β, the transcription factor regulating IL-17-responsive genes, is expressed preferentially in differentiated keratinocytes. Conclusions/Significance: The genes identified in IL-17-treated RHE are likely relevant to the IL-17 effects in psoriasis, since ixekizumab (anti-IL-17A agent) strongly suppressed the «RHE» genes in psoriasis patients treated in vivo with this IL-17 antagonist. © 2014 Chiricozzi et al

Hydration and cooling in elite athletes: relationship with performance, body mass loss and body temperatures during the Doha 2019 IAAF World Athletics Championships.

PURPOSE: To characterise hydration, cooling, body mass loss, and core (Tcore) and skin (Tsk) temperatures during World Athletics Championships in hot-humid conditions. METHODS: Marathon and race-walk (20 km and 50 km) athletes (n=83, 36 women) completed a pre-race questionnaire. Pre-race and post-race body weight (n=74), Tcore (n=56) and Tsk (n=49; thermography) were measured. RESULTS: Most athletes (93%) had a pre-planned drinking strategy (electrolytes (83%), carbohydrates (81%)) while ice slurry was less common (11%; p<0.001). More men than women relied on electrolytes and carbohydrates (91%-93% vs 67%-72%, p≤0.029). Drinking strategies were based on personal experience (91%) rather than external sources (p<0.001). Most athletes (80%) planned pre-cooling (ice vests (53%), cold towels (45%), neck collars (21%) and ice slurry (21%)) and/or mid-cooling (93%; head/face dousing (65%) and cold water ingestion (52%)). Menthol usage was negligible (1%-2%). Pre-race Tcore was lower in athletes using ice vests (37.5°C±0.4°C vs 37.8°C±0.3°C, p=0.024). Tcore (pre-race 37.7°C±0.3°C, post-race 39.6°C±0.6°C) was independent of event, ranking or performance (p≥0.225). Pre-race Tsk was correlated with faster race completion (r=0.32, p=0.046) and was higher in non-finishers (did not finish (DNF); 33.8°C±0.9°C vs 32.6°C±1.4°C, p=0.017). Body mass loss was higher in men than women (-2.8±1.5% vs -1.3±1.6%, p<0.001), although not associated with performance. CONCLUSION: Most athletes' hydration strategies were pre-planned based on personal experience. Ice vests were the most adopted pre-cooling strategy and the only one minimising Tcore, suggesting that event organisers should be cognisant of logistics (ie, freezers). Dehydration was moderate and unrelated to performance. Pre-race Tsk was related to performance and DNF, suggesting that Tsk modulation should be incorporated into pre-race strategies

Hydration and cooling in elite athletes: relationship with performance, body mass loss and body temperatures during the Doha 2019 IAAF World Athletics Championships

Purpose: To characterise hydration, cooling, body mass loss, and core (Tcore) and skin (Tsk) temperatures during World Athletics Championships in hot-humid conditions. Methods: Marathon and race-walk (20 km and 50 km) athletes (n=83, 36 women) completed a pre-race questionnaire. Pre-race and post-race body weight (n=74), Tcore (n=56) and Tsk (n=49; thermography) were measured. Results: Most athletes (93%) had a pre-planned drinking strategy (electrolytes (83%), carbohydrates (81%)) while ice slurry was less common (11%; p<0.001). More men than women relied on electrolytes and carbohydrates (91%–93% vs 67%–72%, p≤0.029). Drinking strategies were based on personal experience (91%) rather than external sources (p<0.001). Most athletes (80%) planned pre-cooling (ice vests (53%), cold towels (45%), neck collars (21%) and ice slurry (21%)) and/or midcooling (93%; head/face dousing (65%) and cold water ingestion (52%)). Menthol usage was negligible (1%–2%). Pre-race Tcore was lower in athletes using ice vests (37.5°C±0.4°C vs 37.8°C±0.3°C, p=0.024). Tcore (pre-race 37.7°C±0.3°C, post-race 39.6°C±0.6°C) was independent of event, ranking or performance (p≥0.225). Pre-race Tsk was correlated with faster race completion (r=0.32, p=0.046) and was higher in non-finishers (did not finish (DNF); 33.8°C±0.9°C vs 32.6°C±1.4°C, p=0.017). Body mass loss was higher in men than women (−2.8±1.5% vs −1.3±1.6%, p<0.001), although not associated with performance. Conclusion: Most athletes’ hydration strategies were pre-planned based on personal experience. Ice vests were the most adopted pre-cooling strategy and the only one minimising Tcore, suggesting that event organisers should be cognisant of logistics (ie, freezers). Dehydration was moderate and unrelated to performance. Pre-race Tsk was related to performance and DNF, suggesting that Tsk modulation should be incorporated into pre-race strategies