Pms2 suppresses large expansions of the (GAA·TTC)n sequence in neuronal tissues

Copyright @ 2012 Bourn et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Expanded trinucleotide repeat sequences are the cause of several inherited neurodegenerative diseases. Disease pathogenesis is correlated with several features of somatic instability of these sequences, including further large expansions in postmitotic tissues. The presence of somatic expansions in postmitotic tissues is consistent with DNA repair being a major determinant of somatic instability. Indeed, proteins in the mismatch repair (MMR) pathway are required for instability of the expanded (CAG·CTG)(n) sequence, likely via recognition of intrastrand hairpins by MutSβ. It is not clear if or how MMR would affect instability of disease-causing expanded trinucleotide repeat sequences that adopt secondary structures other than hairpins, such as the triplex/R-loop forming (GAA·TTC)(n) sequence that causes Friedreich ataxia. We analyzed somatic instability in transgenic mice that carry an expanded (GAA·TTC)(n) sequence in the context of the human FXN locus and lack the individual MMR proteins Msh2, Msh6 or Pms2. The absence of Msh2 or Msh6 resulted in a dramatic reduction in somatic mutations, indicating that mammalian MMR promotes instability of the (GAA·TTC)(n) sequence via MutSα. The absence of Pms2 resulted in increased accumulation of large expansions in the nervous system (cerebellum, cerebrum, and dorsal root ganglia) but not in non-neuronal tissues (heart and kidney), without affecting the prevalence of contractions. Pms2 suppressed large expansions specifically in tissues showing MutSα-dependent somatic instability, suggesting that they may act on the same lesion or structure associated with the expanded (GAA·TTC)(n) sequence. We conclude that Pms2 specifically suppresses large expansions of a pathogenic trinucleotide repeat sequence in neuronal tissues, possibly acting independently of the canonical MMR pathway.IDB was supported by a postdoctoral fellowship from the National Ataxia Foundation. RMP was supported by Ataxia UK. SA was supported by The Wellcome Trust. This research was made possible by grants from the National Institutes of Health (NIH/NINDS) and the Muscular Dystrophy Association to S.I.B

The development and trial of an unmanned aerial system for the measurement of methane flux from landfill and greenhouse gas emission hotspots

This paper describes the development of a new sampling and measurement method to infer methane flux using proxy measurements of CO2 concentration and wind data recorded by Unmanned Aerial Systems (UAS). The flux method described and trialed here is appropriate to the spatial scale of landfill sites and analogous greenhouse gas emission hotspots, making it an important new method for low-cost and rapid case study quantification of fluxes from currently uncertain (but highly important) greenhouse gas sources.We present a case study using these UAS-based measurements to derive instantaneous methane fluxes from a test landfill site in the north of England using a mass balance model tailored for UAS sampling and co-emitted CO2 concentration as a methane-emission proxy. Methane flux (and flux uncertainty) during two trials on 27 November 2014 and 5 March 2015, were found to be 0.140 kg s−1 (±61% at 1σ), and 0.050 kg s−1 (±54% at 1σ), respectively. Uncertainty contributing to the flux was dominated by ambient variability in the background (inflow) concentration (>40%) and wind speed (>10%); with instrumental error contributing only ∼1–2%. The approach described represents an important advance concerning the challenging problem of greenhouse gas hotspot flux calculation, and offers transferability to a wide range of analogous environments. This new measurement solution could add to a toolkit of approaches to better validate source-specific greenhouse emissions inventories – an important new requirement of the UNFCCC COP21 (Paris) climate change agreement

MutLα heterodimers modify the molecular phenotype of Friedreich ataxia

This article has been made available through the Brunel Open Access Publishing Fund.Background: Friedreich ataxia (FRDA), the most common autosomal recessive ataxia disorder, is caused by a dynamic GAA repeat expansion mutation within intron 1 of FXN gene, resulting in down-regulation of frataxin expression. Studies of cell and mouse models have revealed a role for the mismatch repair (MMR) MutS-heterodimer complexes and the PMS2 component of the MutLα complex in the dynamics of intergenerational and somatic GAA repeat expansions: MSH2, MSH3 and MSH6 promote GAA repeat expansions, while PMS2 inhibits GAA repeat expansions. Methodology/Principal Findings: To determine the potential role of the other component of the MutLα complex, MLH1, in GAA repeat instability in FRDA, we have analyzed intergenerational and somatic GAA repeat expansions from FXN transgenic mice that have been crossed with Mlh1 deficient mice. We find that loss of Mlh1 activity reduces both intergenerational and somatic GAA repeat expansions. However, we also find that loss of either Mlh1 or Pms2 reduces FXN transcription, suggesting different mechanisms of action for Mlh1 and Pms2 on GAA repeat expansion dynamics and regulation of FXN transcription. Conclusions/Significance: Both MutLα components, PMS2 and MLH1, have now been shown to modify the molecular phenotype of FRDA. We propose that upregulation of MLH1 or PMS2 could be potential FRDA therapeutic approaches to increase FXN transcription. © 2014 Ezzatizadeh et al.This article has been made available through the Brunel Open Access Publishing Fund

Generation and characterisation of Friedreich ataxia YG8R mouse fibroblast and neural stem cell models

This article has been made available through the Brunel Open Access Publishing Fund.Background: Friedreich ataxia (FRDA) is an autosomal recessive neurodegenerative disease caused by GAA repeat expansion in the first intron of the FXN gene, which encodes frataxin, an essential mitochondrial protein. To further characterise the molecular abnormalities associated with FRDA pathogenesis and to hasten drug screening, the development and use of animal and cellular models is considered essential. Studies of lower organisms have already contributed to understanding FRDA disease pathology, but mammalian cells are more related to FRDA patient cells in physiological terms. Methodology/Principal Findings: We have generated fibroblast cells and neural stem cells (NSCs) from control Y47R mice (9 GAA repeats) and GAA repeat expansion YG8R mice (190+120 GAA repeats). We then differentiated the NSCs in to neurons, oligodendrocytes and astrocytes as confirmed by immunocytochemical analysis of cell specific markers. The three YG8R mouse cell types (fibroblasts, NSCs and differentiated NSCs) exhibit GAA repeat stability, together with reduced expression of frataxin and reduced aconitase activity compared to control Y47R cells. Furthermore, YG8R cells also show increased sensitivity to oxidative stress and downregulation of Pgc-1α and antioxidant gene expression levels, especially Sod2. We also analysed various DNA mismatch repair (MMR) gene expression levels and found that YG8R cells displayed significant reduction in expression of several MMR genes, which may contribute to the GAA repeat stability. Conclusions/Significance: We describe the first fibroblast and NSC models from YG8R FRDA mice and we confirm that the NSCs can be differentiated into neurons and glia. These novel FRDA mouse cell models, which exhibit a FRDA-like cellular and molecular phenotype, will be valuable resources to further study FRDA molecular pathogenesis. They will also provide very useful tools for preclinical testing of frataxin-increasing compounds for FRDA drug therapy, for gene therapy, and as a source of cells for cell therapy testing in FRDA mice. © 2014 Sandi et al

A near-field Gaussian plume inversion flux quantification method, applied to unmanned aerial vehicle sampling

The accurate quantification of methane emissions from point sources is required to better quantify emissions for sector-specific reporting and inventory validation. An unmanned aerial vehicle (UAV) serves as a platform to sample plumes near to source. This paper describes a near-field Gaussian plume inversion (NGI) flux technique, adapted for downwind sampling of turbulent plumes, by fitting a plume model to measured flux density in three spatial dimensions. The method was refined and tested using sample data acquired from eight UAV flights, which measured a controlled release of methane gas. Sampling was conducted to a maximum height of 31 m (i.e. above the maximum height of the emission plumes). The method applies a flux inversion to plumes sampled near point sources. To test the method, a series of random walk sampling simulations were used to derive an NGI upper uncertainty bound by quantifying systematic flux bias due to a limited spatial sampling extent typical for short-duration small UAV flights (less than 30 min). The development of the NGI method enables its future use to quantify methane emissions for point sources, facilitating future assessments of emissions from specific source-types and source areas. This allows for atmospheric measurement-based fluxes to be derived using downwind UAV sampling for relatively rapid flux analysis, without the need for access to difficult-to-reach areas

Insights From Analysis of Human Antigen-Specific Memory B Cell Repertoires

Memory B cells that are generated during an infection or following vaccination act as sentinels to guard against future infections. Upon repeat antigen exposure memory B cells differentiate into new antibody-secreting plasma cells to provide rapid and sustained protection. Some pathogens evade or suppress the humoral immune system, or induce memory B cells with a diminished ability to differentiate into new plasma cells. This leaves the host vulnerable to chronic or recurrent infections. Single cell approaches coupled with next generation antibody gene sequencing facilitate a detailed analysis of the pathogen-specific memory B cell repertoire. Monoclonal antibodies that are generated from antibody gene sequences allow a functional analysis of the repertoire. This review discusses what has been learned thus far from analysis of diverse pathogen-specific memory B cell compartments and describes major differences in their repertoires. Such information may illuminate ways to advance the goal of improving vaccine and therapeutic antibody design

Prevalence and architecture of de novo mutations in developmental disorders.

The genomes of individuals with severe, undiagnosed developmental disorders are enriched in damaging de novo mutations (DNMs) in developmentally important genes. Here we have sequenced the exomes of 4,293 families containing individuals with developmental disorders, and meta-analysed these data with data from another 3,287 individuals with similar disorders. We show that the most important factors influencing the diagnostic yield of DNMs are the sex of the affected individual, the relatedness of their parents, whether close relatives are affected and the parental ages. We identified 94 genes enriched in damaging DNMs, including 14 that previously lacked compelling evidence of involvement in developmental disorders. We have also characterized the phenotypic diversity among these disorders. We estimate that 42% of our cohort carry pathogenic DNMs in coding sequences; approximately half of these DNMs disrupt gene function and the remainder result in altered protein function. We estimate that developmental disorders caused by DNMs have an average prevalence of 1 in 213 to 1 in 448 births, depending on parental age. Given current global demographics, this equates to almost 400,000 children born per year

Heterozygous Variants in KMT2E Cause a Spectrum of Neurodevelopmental Disorders and Epilepsy.

We delineate a KMT2E-related neurodevelopmental disorder on the basis of 38 individuals in 36 families. This study includes 31 distinct heterozygous variants in KMT2E (28 ascertained from Matchmaker Exchange and three previously reported), and four individuals with chromosome 7q22.2-22.23 microdeletions encompassing KMT2E (one previously reported). Almost all variants occurred de novo, and most were truncating. Most affected individuals with protein-truncating variants presented with mild intellectual disability. One-quarter of individuals met criteria for autism. Additional common features include macrocephaly, hypotonia, functional gastrointestinal abnormalities, and a subtle facial gestalt. Epilepsy was present in about one-fifth of individuals with truncating variants and was responsive to treatment with anti-epileptic medications in almost all. More than 70% of the individuals were male, and expressivity was variable by sex; epilepsy was more common in females and autism more common in males. The four individuals with microdeletions encompassing KMT2E generally presented similarly to those with truncating variants, but the degree of developmental delay was greater. The group of four individuals with missense variants in KMT2E presented with the most severe developmental delays. Epilepsy was present in all individuals with missense variants, often manifesting as treatment-resistant infantile epileptic encephalopathy. Microcephaly was also common in this group. Haploinsufficiency versus gain-of-function or dominant-negative effects specific to these missense variants in KMT2E might explain this divergence in phenotype, but requires independent validation. Disruptive variants in KMT2E are an under-recognized cause of neurodevelopmental abnormalities

Bi-allelic Loss-of-Function CACNA1B Mutations in Progressive Epilepsy-Dyskinesia.

The occurrence of non-epileptic hyperkinetic movements in the context of developmental epileptic encephalopathies is an increasingly recognized phenomenon. Identification of causative mutations provides an important insight into common pathogenic mechanisms that cause both seizures and abnormal motor control. We report bi-allelic loss-of-function CACNA1B variants in six children from three unrelated families whose affected members present with a complex and progressive neurological syndrome. All affected individuals presented with epileptic encephalopathy, severe neurodevelopmental delay (often with regression), and a hyperkinetic movement disorder. Additional neurological features included postnatal microcephaly and hypotonia. Five children died in childhood or adolescence (mean age of death: 9 years), mainly as a result of secondary respiratory complications. CACNA1B encodes the pore-forming subunit of the pre-synaptic neuronal voltage-gated calcium channel Cav2.2/N-type, crucial for SNARE-mediated neurotransmission, particularly in the early postnatal period. Bi-allelic loss-of-function variants in CACNA1B are predicted to cause disruption of Ca2+ influx, leading to impaired synaptic neurotransmission. The resultant effect on neuronal function is likely to be important in the development of involuntary movements and epilepsy. Overall, our findings provide further evidence for the key role of Cav2.2 in normal human neurodevelopment.MAK is funded by an NIHR Research Professorship and receives funding from the Wellcome Trust, Great Ormond Street Children's Hospital Charity, and Rosetrees Trust. E.M. received funding from the Rosetrees Trust (CD-A53) and Great Ormond Street Hospital Children's Charity. K.G. received funding from Temple Street Foundation. A.M. is funded by Great Ormond Street Hospital, the National Institute for Health Research (NIHR), and Biomedical Research Centre. F.L.R. and D.G. are funded by Cambridge Biomedical Research Centre. K.C. and A.S.J. are funded by NIHR Bioresource for Rare Diseases. The DDD Study presents independent research commissioned by the Health Innovation Challenge Fund (grant number HICF-1009-003), a parallel funding partnership between the Wellcome Trust and the Department of Health, and the Wellcome Trust Sanger Institute (grant number WT098051). We acknowledge support from the UK Department of Health via the NIHR comprehensive Biomedical Research Centre award to Guy's and St. Thomas' National Health Service (NHS) Foundation Trust in partnership with King's College London. This research was also supported by the NIHR Great Ormond Street Hospital Biomedical Research Centre. J.H.C. is in receipt of an NIHR Senior Investigator Award. The research team acknowledges the support of the NIHR through the Comprehensive Clinical Research Network. The views expressed are those of the author(s) and not necessarily those of the NHS, the NIHR, Department of Health, or Wellcome Trust. E.R.M. acknowledges support from NIHR Cambridge Biomedical Research Centre, an NIHR Senior Investigator Award, and the University of Cambridge has received salary support in respect of E.R.M. from the NHS in the East of England through the Clinical Academic Reserve. I.E.S. is supported by the National Health and Medical Research Council of Australia (Program Grant and Practitioner Fellowship)

Carnivore detection at the domestic/wildlife interface within Mpumalanga Province, South Africa

South African protected areas account for 8% of the total landmass according to World Bank indicators. Effective conservation of biodiversity in protected areas requires the development of specific reserve management objectives addressing species and disease management. The primary objective of the current study was to identify predictors of carnivore detection in an effort to inform carnivore species management plans on Andover and Manyeleti nature reserves in South Africa. A limited number of camera traps were placed randomly using a grid system. Species detection data were analysed using mixed-effects logistic regression and Spearman’s correlation coefficients. Deterministic inverse distance weighted distribution maps were used to describe the spatial distribution of carnivore species. Camera traps identified similar species as traditional call-up surveys during the study and would be useful as an adjunct census method. Carnivore detection was associated with several variables, including the presence of specific prey species. The measured intra-and interspecies interactions suggested the risk of disease transmission among species, and vaccination for prevalent diseases should be considered to manage this risk.SUPPLEMENTARY MATERIAL: Table S1: Summary of camera traps and census, Table S2: The significant univariate associations between any carnivore group (lion, leopard, serval, cheetah), canine (African wild dog, jackal), other (spotted hyena, civet, and genet) group detection and continuous predictor variables based on camera trap data collected within two protected areas, Mpumalanga Province, South Africa for the years 2015–2017., Table S3: The important univariate associations between lion, leopard, cheetah, serval, African wild dog, jackal, spotted hyena, civet, and genet detection and continuous predictor variables on camera trap data collected within two protected areas, Mpumalanga Province, South Africa for the years 2015–2017, Table S4: The important univariate associations between lion, leopard, serval, and cheetah detection and continuous predictor variables on camera trap data collected within two protected areas, Mpumalanga Province, South Africa, for the years 2015–2017, Table S5: The important univariate associations between spotted hyena, genet, and civet detection and continuous predictor variables on camera trap data collected within two protected areas, Mpumalanga Province, South Africa, for the years 2015–2017, Table S6: The important univariate associations between spotted hyena, detection, and continuous predictor variables on camera trap data collected within two protected areas, Mpumalanga Province, South Africa, for the years 2015–2017.The National Research Foundation (NRF), South Africahttps://www.mdpi.com/journal/animalsam2022Centre for Veterinary Wildlife StudiesProduction Animal Studie