Exon Exchange Approach to Repair Duchenne Dystrophin Transcripts

Background: Trans-splicing strategies for mRNA repair involve engineered transcripts designed to anneal target mRNAs in order to interfere with their natural splicing, giving rise to mRNA chimeras where endogenous mutated exons have been replaced by exogenous replacement sequences. A number of trans-splicing molecules have already been proposed for replacing either the 59 or the 39 part of transcripts to be repaired. Here, we show the feasibility of RNA surgery by using a double trans-splicing approach allowing the specific substitution of a given mutated exon. Methodology/Principal Findings: As a target we used a minigene encoding a fragment of the mdx dystrophin gene enclosing the mutated exon (exon 23). This minigene was cotransfected with a variety of exon exchange constructions, differing in their annealing domains. We obtained accurate and efficient replacement of exon 23 in the mRNA target. Adding up a downstream intronic splice enhancer DISE in the exon exchange molecule enhanced drastically its efficiency up to 25–45 % of repair depending on the construction in use. Conclusions/Significance: These results demonstrate the possibility to fix up mutated exons, refurbish deleted exons and introduce protein motifs, while keeping natural untranslated sequences, which are essential for mRNA stability and translation regulation. Conversely to the well-known exon skipping, exon exchange has the advantage to be compatible with almost any type of mutations and more generally to a wide range of genetic conditions. In particular, it allows addressing disorders caused by dominant mutations

Deficiency in origin licensing proteins impairs cilia formation: implications for the aetiology of meier-gorlin syndrome

Mutations in ORC1, ORC4, ORC6, CDT1, and CDC6, which encode proteins required for DNA replication origin licensing, cause Meier-Gorlin syndrome (MGS), a disorder conferring microcephaly, primordial dwarfism, underdeveloped ears, and skeletal abnormalities. Mutations in ATR, which also functions during replication, can cause Seckel syndrome, a clinically related disorder. These findings suggest that impaired DNA replication could underlie the developmental defects characteristic of these disorders. Here, we show that although origin licensing capacity is impaired in all patient cells with mutations in origin licensing component proteins, this does not correlate with the rate of progression through S phase. Thus, the replicative capacity in MGS patient cells does not correlate with clinical manifestation. However, ORC1-deficient cells from MGS patients and siRNA-mediated depletion of origin licensing proteins also have impaired centrosome and centriole copy number. As a novel and unexpected finding, we show that they also display a striking defect in the rate of formation of primary cilia. We demonstrate that this impacts sonic hedgehog signalling in ORC1-deficient primary fibroblasts. Additionally, reduced growth factor-dependent signaling via primary cilia affects the kinetics of cell cycle progression following cell cycle exit and re-entry, highlighting an unexpected mechanism whereby origin licensing components can influence cell cycle progression. Finally, using a cell-based model, we show that defects in cilia function impair chondroinduction. Our findings raise the possibility that a reduced efficiency in forming cilia could contribute to the clinical features of MGS, particularly the bone development abnormalities, and could provide a new dimension for considering developmental impacts of licensing deficiency

A massive, quiescent galaxy at redshift of z=3.717

In the early Universe finding massive galaxies that have stopped forming stars present an observational challenge as their rest-frame ultraviolet emission is negligible and they can only be reliably identified by extremely deep near-infrared surveys. These have revealed the presence of massive, quiescent early-type galaxies appearing in the universe as early as z$\sim$2, an epoch 3 Gyr after the Big Bang. Their age and formation processes have now been explained by an improved generation of galaxy formation models where they form rapidly at z$\sim$3-4, consistent with the typical masses and ages derived from their observations. Deeper surveys have now reported evidence for populations of massive, quiescent galaxies at even higher redshifts and earlier times, however the evidence for their existence, and redshift, has relied entirely on coarsely sampled photometry. These early massive, quiescent galaxies are not predicted by the latest generation of theoretical models. Here, we report the spectroscopic confirmation of one of these galaxies at redshift z=3.717 with a stellar mass of 1.7$\times$10$^{11}$ M$_\odot$ whose absorption line spectrum shows no current star-formation and which has a derived age of nearly half the age of the Universe at this redshift. The observations demonstrates that the galaxy must have quickly formed the majority of its stars within the first billion years of cosmic history in an extreme and short starburst. This ancestral event is similar to those starting to be found by sub-mm wavelength surveys pointing to a possible connection between these two populations. Early formation of such massive systems is likely to require significant revisions to our picture of early galaxy assembly.Comment: 6 pages, 7 figures. This is the final preprint corresponding closely to the published version. Uploaded 6 months after publication in accordance with Nature polic

Wavelet Cycle Spinning Denoising of NDE Ultrasonic Signals Using a Random Selection of Shifts

Wavelets are a powerful tool for signal and image denoising. Most of the denoising applications in different fields were based on the thresholding of the discrete wavelet transform (DWT) coefficients. Nevertheless, DWT transform is not a time or shift invariant transform and results depend on the selected shift. Improvements on the denoising performance can be obtained using the stationary wavelet transform (SWT) (also called shift-invariant or undecimated wavelet transform). Denoising using SWT has previously shown a robust and usually better performance than denoising using DWT but with a higher computational cost. In this paper, wavelet shrinkage schemes are applied for reducing noise in synthetic and experimental non-destructive evaluation ultrasonic A-scans, using DWT and a cycle-spinning implementation of SWT. A new denoising procedure, which we call random partial cycle spinning (RPCS), is presented. It is based on a cycle-spinning over a limited number of shifts that are selected in a random way. Wavelet denoising based on DWT, SWT and RPCS have been applied to the same sets of ultrasonic A-scans and their performances in terms of SNR are compared. In all cases three well known threshold selection rules (Universal, Minimax and Sure), with decomposition level dependent selection, have been used. It is shown that the new procedure provides a good robust denoising performance, without the DWT fluctuating performance, and close to SWT but with a much lower computational cost.This work was partially supported by Spanish MCI Project DPI2011-22438San Emeterio Prieto, JL.; Rodríguez-Hernández, MA. (2015). Wavelet Cycle Spinning Denoising of NDE Ultrasonic Signals Using a Random Selection of Shifts. Journal of Nondestructive Evaluation. 34(1):1-8. https://doi.org/10.1007/s10921-014-0270-8S18341Galloway, R.L., McDermott, B.A., Thurstone, F.L.: A frequency diversity process for speckle reduction in real-time ultrasonic images. IEEE Trans. Ultrason. Ferroelectr. Freq. Control 35, 45–49 (1988)Newhouse, V.L., Bilgutay, N.M., Saniie, J., Furgason, E.S.: Flaw-to-grain echo enhancement by split spectrum processing. Ultrasonics 20, 59–68 (1982)Karpur, P., Canelones, O.J.: Split spectrum processing: a new filtering approach for improved signal-to-noise ratio enhancement of ultrasonic signals. Ultrasonics 30, 351–357 (1992)Donoho, D.L., Johnstone, I.M.: Ideal spatial adaptation by wavelet shrinkage. Biometrika 81, 425–455 (1994)Donoho, D.L., Johnstone, I.M., Kerkyacharian, G., Picard, D.: Wavelet shrinkage: asymptotia? J. R Stat. Soc. Ser. B 57, 301–369 (1995)Donoho, D.L., Johnstone, I.M.: Adapting to unknown smoothness via wavelet shrinkage. J. Am. Stat. Assoc. 90, 1200–1224 (1995)Johnstone, I.M., Silverman, B.W.: Wavelet threshold estimators for data with correlated noise. J. R Stat. Soc. 59, 319–351 (1997)Jansen, M.: Noise Reduction by Wavelet Thresholding. Lecture Notes in Statistics 161. Springer, New York (2001). doi: 10.1007/978-1-4613-0145-5Nason, G.P., Silverman, B.W.:The stationary wavelet transform and some statistical applications. In: Antoniadis, A., Oppenheim, G. (eds.) Wavelets and Statistics. Lecture Notes in Statistics, Vol. 103, pp 281–299. Springer, New York (1995)Lang, M., Guo, H., Odegard, J.E., Burrus, C.S.: Noise reduction using an undecimated discrete wavelet transform. IEEE Signal Proc. Lett. 3, 10–12 (1996)Coifman, R.R., Donoho, D.L.: Translation-invariant de-noising. In: Antoniadis, A., Oppenheim, G. (eds.) Wavelets and Statistics. Lecture Notes in Statistics, vol. 103, pp 125–150, Springer, New York (1995) .Abbate, A., Koay, J., Frankel, J., Schroeder, S.C., Das, P.: Signal detection and noise suppression using a wavelet transform signal processor: application to ultrasonic flaw detection. IEEE Trans. Ultrason. Ferroelectr. Freq. Control 44, 14–26 (1997)Lázaro, J.C., San Emeterio, J.L., Ramos, A., Fernandez, J.L.: Influence of thresholding procedures in ultrasonic grain noise reduction using wavelets. Ultrasonics 40, 263–267 (2002)Matz, V., Smid, R., Starman, S., Kreidl, M.: Signal-to-noise ratio enhancement based on wavelet filtering in ultrasonic testing. Ultrasonics 49, 752–759 (2009)Kubinyi, M., Kreibich, O., Neuzil, J., Smid, R.: EMAT noise suppression using information fusion in stationary wavelet packets. IEEE Trans. Ultrason. Ferroelectr. Freq. Control 58, 1027–1036 (2011)Shi, G.M., Chen, X.Y., Song, X.X., Qui, F., Ding, A.L.: Signal matching wavelet for ultrasonic flaw detection in high background noise. IEEE Trans. Ultrason. Ferroelectr. Freq. Control 58, 776–787 (2011)Song, S.P., Que, P.W.: Wavelet based noise suppression technique and its application to ultrasonic flaw detection. Ultrasonics 44, 188–193 (2006)Rodriguez, M.A., San Emeterio, J.L., Lázaro, J.C., Ramos, A.: Ultrasonic flaw detection in NDE of highly scattering materials using wavelet and Wigner-Ville transform processing. Ultrasonics 42, 847–851 (2004)Zhang, G.M., Zhang, S.Y., Wang, Y.W.: Application of adaptive time-frequency decomposition in ultrasonic NDE of highly-scattering materials. Ultrasonics 38, 961–964 (2000)Drai, R., Khelil, M., Benchaala, A.: Time frequency and wavelet transform applied to selected problems in ultrasonics NDE. NDT & E Int. 35, 567–572 (2002)Pardo, E., San Emeterio, J.L.: Noise reduction in ultrasonic NDT using undecimated wavelet transforms. Ultrasonics 44, e1063–e1067 (2006)Kechida, A., Drai, R., Guessoum, A.: Texture analysis for flaw detection in ultrasonic images. J. Nondestruct. Eval. 31, 108–116 (2012). doi: 10.1007/s10921-011-0126-4Rucka, M., Wilde, K.: Experimental study on ultrasonic monitoring of splitting failure in reinforced concrete. J. Nondestruct. Eval. 32, 372–383 (2013). doi: 10.1007/s10921-013-0191-yHosseini, S.M.H., Duczek, S., Gabbert, U.: Damage localization in plates using mode conversion characteristics of ultrasonic guided waves. J. Nondestruct. Eval. 33, 152–165 (2014). doi: 10.1007/s10921-013-0211-yMohammed, M.S., Ki-Seong, K.: Shift-invariant wavelet packet for signal de-noising in ultrasonic testing. Insight 54, 366–370 (2012)San Emeterio, J.L., Rodriguez-Hernandez, M.A.: Wavelet denoising of ultrasonic A-scans by random partial cycle spinning. 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Acute kidney disease and renal recovery : consensus report of the Acute Disease Quality Initiative (ADQI) 16 Workgroup

Consensus definitions have been reached for both acute kidney injury (AKI) and chronic kidney disease (CKD) and these definitions are now routinely used in research and clinical practice. The KDIGO guideline defines AKI as an abrupt decrease in kidney function occurring over 7 days or less, whereas CKD is defined by the persistence of kidney disease for a period of > 90 days. AKI and CKD are increasingly recognized as related entities and in some instances probably represent a continuum of the disease process. For patients in whom pathophysiologic processes are ongoing, the term acute kidney disease (AKD) has been proposed to define the course of disease after AKI; however, definitions of AKD and strategies for the management of patients with AKD are not currently available. In this consensus statement, the Acute Disease Quality Initiative (ADQI) proposes definitions, staging criteria for AKD, and strategies for the management of affected patients. We also make recommendations for areas of future research, which aim to improve understanding of the underlying processes and improve outcomes for patients with AKD

MetaPath: identifying differentially abundant metabolic pathways in metagenomic datasets

Enabled by rapid advances in sequencing technology, metagenomic studies aim to characterize entire communities of microbes bypassing the need for culturing individual bacterial members. One major goal of metagenomic studies is to identify specific functional adaptations of microbial communities to their habitats. The functional profile and the abundances for a sample can be estimated by mapping metagenomic sequences to the global metabolic network consisting of thousands of molecular reactions. Here we describe a powerful analytical method (MetaPath) that can identify differentially abundant pathways in metagenomic datasets, relying on a combination of metagenomic sequence data and prior metabolic pathway knowledge. First, we introduce a scoring function for an arbitrary subnetwork and find the max-weight subnetwork in the global network by a greedy search algorithm. Then we compute two p values (p abund and p struct ) using nonparametric approaches to answer two different statistical questions: (1) is this subnetwork differentically abundant? (2) What is the probability of finding such good subnetworks by chance given the data and network structure? Finally, significant metabolic subnetworks are discovered based on these two p values. In order to validate our methods, we have designed a simulated metabolic pathways dataset and show that MetaPath outperforms other commonly used approaches. We also demonstrate the power of our methods in analyzing two publicly available metagenomic datasets, and show that the subnetworks identified by MetaPath provide valuable insights into the biological activities of the microbiome. We have introduced a statistical method for finding significant metabolic subnetworks from metagenomic datasets. Compared with previous methods, results from MetaPath are more robust against noise in the data, and have significantly higher sensitivity and specificity (when tested on simulated datasets). When applied to two publicly available metagenomic datasets, the output of MetaPath is consistent with previous observations and also provides several new insights into the metabolic activity of the gut microbiome. The software is freely available at http://metapath.cbcb.umd.edu .https://doi.org/10.1186/1753-6561-5-S2-S

Chromosomes of Theridiidae spiders (Entelegynae): Interspecific karyotype diversity in Argyrodes and diploid number intraspecific variability in Nesticodes rufipes

Theridiidae is a derived family within the Araneoidea clade. In contrast to closely related groups, the 2n(male) = 20+X1 X 2 with acro/telocentric chromosomes is the most widespread karyotype among the theridiid spiders. In this work, the cytogenetic analysis of Argyrodes elevatus revealed original chromosome features different from those previously registered for Theridiidae, including the presence of 2n(male) = 20+X with meta/submetacentric chromosomes. Most individuals of Nesticodes rufipes showed family conserved karyotype characteristics. However, one individual had a 2n(male) = 24 due to the presence of an extra chromosome pair, which exhibited regular behavior and reductional segregation during meiosis. After silver staining, mitotic cells exhibited NORs localized on the terminal regions of the short arms of pairs 2, 3, and 4 of A. elevatus and on the terminal regions of long arms of pair 4 of N. rufipes. The comparative analysis with data from phylogenetically related species allowed the clarification of the origin of the interspecific and intraspecific chromosome variability observed in Argyrodes and in N. rufipes, respectively

Global, regional, and national comparative risk assessment of 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks, 1990-2015: a systematic analysis for the Global Burden of Disease Study 2015

SummaryBackground The Global Burden of Diseases, Injuries, and Risk Factors Study 2015 provides an up-to-date synthesis of the evidence for risk factor exposure and the attributable burden of disease. By providing national and subnational assessments spanning the past 25 years, this study can inform debates on the importance of addressing risks in context. Methods We used the comparative risk assessment framework developed for previous iterations of the Global Burden of Disease Study to estimate attributable deaths, disability-adjusted life-years (DALYs), and trends in exposure by age group, sex, year, and geography for 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks from 1990 to 2015. This study included 388 risk-outcome pairs that met World Cancer Research Fund-defined criteria for convincing or probable evidence. We extracted relative risk and exposure estimates from randomised controlled trials, cohorts, pooled cohorts, household surveys, census data, satellite data, and other sources. We used statistical models to pool data, adjust for bias, and incorporate covariates. We developed a metric that allows comparisons of exposure across risk factors—the summary exposure value. Using the counterfactual scenario of theoretical minimum risk level, we estimated the portion of deaths and DALYs that could be attributed to a given risk. We decomposed trends in attributable burden into contributions from population growth, population age structure, risk exposure, and risk-deleted cause-specific DALY rates. We characterised risk exposure in relation to a Socio-demographic Index (SDI). Findings Between 1990 and 2015, global exposure to unsafe sanitation, household air pollution, childhood underweight, childhood stunting, and smoking each decreased by more than 25%. Global exposure for several occupational risks, high body-mass index (BMI), and drug use increased by more than 25% over the same period. All risks jointly evaluated in 2015 accounted for 57·8% (95% CI 56·6–58·8) of global deaths and 41·2% (39·8–42·8) of DALYs. In 2015, the ten largest contributors to global DALYs among Level 3 risks were high systolic blood pressure (211·8 million [192·7 million to 231·1 million] global DALYs), smoking (148·6 million [134·2 million to 163·1 million]), high fasting plasma glucose (143·1 million [125·1 million to 163·5 million]), high BMI (120·1 million [83·8 million to 158·4 million]), childhood undernutrition (113·3 million [103·9 million to 123·4 million]), ambient particulate matter (103·1 million [90·8 million to 115·1 million]), high total cholesterol (88·7 million [74·6 million to 105·7 million]), household air pollution (85·6 million [66·7 million to 106·1 million]), alcohol use (85·0 million [77·2 million to 93·0 million]), and diets high in sodium (83·0 million [49·3 million to 127·5 million]). From 1990 to 2015, attributable DALYs declined for micronutrient deficiencies, childhood undernutrition, unsafe sanitation and water, and household air pollution; reductions in risk-deleted DALY rates rather than reductions in exposure drove these declines. Rising exposure contributed to notable increases in attributable DALYs from high BMI, high fasting plasma glucose, occupational carcinogens, and drug use. Environmental risks and childhood undernutrition declined steadily with SDI; low physical activity, high BMI, and high fasting plasma glucose increased with SDI. In 119 countries, metabolic risks, such as high BMI and fasting plasma glucose, contributed the most attributable DALYs in 2015. Regionally, smoking still ranked among the leading five risk factors for attributable DALYs in 109 countries; childhood underweight and unsafe sex remained primary drivers of early death and disability in much of sub-Saharan Africa. Interpretation Declines in some key environmental risks have contributed to declines in critical infectious diseases. Some risks appear to be invariant to SDI. Increasing risks, including high BMI, high fasting plasma glucose, drug use, and some occupational exposures, contribute to rising burden from some conditions, but also provide opportunities for intervention. Some highly preventable risks, such as smoking, remain major causes of attributable DALYs, even as exposure is declining. Public policy makers need to pay attention to the risks that are increasingly major contributors to global burden. Funding Bill & Melinda Gates Foundation

A change in the optical polarization associated with a gamma-ray flare in the blazar 3C 279

It is widely accepted that strong and variable radiation detected over all accessible energy bands in a number of active galaxies arises from a relativistic, Doppler-boosted jet pointing close to our line of sight. The size of the emitting zone and the location of this region relative to the central supermassive black hole are, however, poorly known, with estimates ranging from light-hours to a light-year or more. Here we report the coincidence of a gamma-ray flare with a dramatic change of optical polarization angle. This provides evidence for co-spatiality of optical and gamma-ray emission regions and indicates a highly ordered jet magnetic field. The results also require a non-axisymmetric structure of the emission zone, implying a curved trajectory for the emitting material within the jet, with the dissipation region located at a considerable distance from the black hole, at about 10^5 gravitational radii.Comment: Published in Nature issued on 18 February 2010. Corresponding authors: Masaaki Hayashida and Greg Madejsk