Potentiation of sensory responses in ventrobasal thalamus in vivo via selective modulation of mGlu1 receptors with a positive allosteric modulator.

Metabotropic glutamate subtype 1 (mGlu1) receptor is thought to play a role in synaptic responses in thalamic relay nuclei. The aim of this study was to evaluate the positive allosteric modulator (PAM) Ro67-4853 as a tool to modulate thalamic mGlu1 receptors on single thalamic neurones in vivo. Ro67-4853, applied by iontophoresis onto ventrobasal thalamus neurones of urethane-anaesthetised rats, selectively enhanced responses to the agonist (S)-3,5-dihydroxy-phenylglycine (DHPG), an effect consistent with mGlu1 potentiation. The PAM was also able to enhance maintained responses to 10 Hz trains of sensory stimulation of the vibrissae, but had little effect on responses to single sensory stimuli. Thus Ro67-4853 appears to be a highly selective tool that can be useful in investigating how mGlu1 receptor potentiation can alter neural processing in vivo. Our results show the importance of mGlu1 in sensory processing and attention mechanisms at the thalamic level and suggest that positive modulation of mGlu1 receptors might be a useful mechanism for enhancing cognitive and attentional processes

Interactions of the Bacillus subtilis DnaE polymerase with replisomal proteins modulate its activity and fidelity

During Bacillus subtilis replication two replicative polymerases function at the replisome to collectively carry out genome replication. In a reconstituted in vitro replication assay, PolC is the main polymerase while the lagging strand DnaE polymerase briefly extends RNA primers synthesized by the primase DnaG prior to handing-off DNA synthesis to PolC. Here, we show in vivo that (i) the polymerase activity of DnaE is essential for both the initiation and elongation stages of DNA replication, (ii) its error rate varies inversely with PolC concentration, and (iii) its misincorporations are corrected by the mismatch repair system post-replication. We also found that the error rates in cells encoding mutator forms of both PolC and DnaE are significantly higher (up to 15-fold) than in PolC mutants. In vitro, we showed that (i) the polymerase activity of DnaE is considerably stimulated by DnaN, SSB and PolC, (ii) its error-prone activity is strongly inhibited by DnaN, and (iii) its errors are proofread by the 30 . 50 exonuclease activity of PolC in a stable template-DnaE –PolC complex. Collectively our data show that protein –protein interactions within the replisome modulate the activity and fidelity of DnaE, and confirm the prominent role of DnaE during B. subtilis replication

Visualizing the Spatial Relationship of the Genome with the Nuclear Envelope Using Fluorescence In Situ Hybridization

The genome has a special relationship with the nuclear envelope in cells. Much of the genome is anchored at the nuclear periphery, tethered by chromatin binding proteins such nuclear lamins and other integral membrane proteins. Even though there are global assays such as DAM-ID or ChIP to assess what parts of the genome are associated with the nuclear envelope, it is also essential to be able to visualize regions of the genome in order to reveal their individual relationships with nuclear structures in single cells. This is executed by fluorescence in situ hybridization (FISH) in 2-dimensional flattened nuclei (2D-FISH) or 3-dimensionally preserved cells (3D-FISH) in combination with indirect immunofluorescence to reveal structural proteins. This chapter explains the protocols for 2D- and 3D-FISH in combination with indirect immunofluorescence and discusses options for image capture and analysis. Due to the nuclear envelope proteins being part of the non-extractable nucleoskeleton, we also describe how to prepare DNA halos through salt extraction and how they can be used to study genome behavior and association when combined with 2D-FISH.SPARKs children’s charity for funding CSC; Brunel University London Progeria Research Fund; The Gordon Memorial Trust; EURO-laminopathies consortium FP6

Chromosomal Rearrangements and Altered Nuclear Organization: Recent Mechanistic Models in Cancer

Copyright: © 2021 by the authors. The last decade has seen significant progress in understanding how the genome is organized spatially within interphase nuclei. Recent analyses have confirmed earlier molecular cytogenetic studies on chromosome positioning within interphase nuclei and provided new information about the topologically associated domains (TADs). Examining the nuances of how genomes are organized within interphase nuclei will provide information fundamental to understanding gene regulation and expression in health and disease. Indeed, the radial spatial positioning of individual gene loci within nuclei has been associated with up- and down-regulation of specific genes, and disruption of normal genome organization within nuclei will result in compromised cellular health. In cancer cells, where reorganization of the nuclear architecture may occur in the presence of chromosomal rearrangements such as translocations, inversions, or deletions, gene repositioning can change their expression. To date, very few studies have focused on radial gene positioning and the correlation to gene expression in cancers. Further investigations would improve our understanding of the biological mechanisms at the basis of cancer and, in particular, in leukemia initiation and progression, especially in those cases where the molecular consequences of chromosomal rearrangements are still unclear. In this review, we summarize the main milestones in the field of genome organization in the nucleus and the alterations to this organization that can lead to cancer diseases.Research Plan PIACERI L.2 and L.3 Starting Grant from the Department of Biological, Geological, and Environmental Sciences, the University of Catania; University of Catania, Catania, Italy; Brunel University London

Individual and Contextual Factors Associated with Low Childhood Immunisation Coverage in Sub-Saharan Africa: A Multilevel Analysis

Background: In 2010, more than six million children in sub-Saharan Africa did not receive the full series of three doses of the diphtheria-tetanus-pertussis vaccine by one year of age. An evidence-based approach to addressing this burden of un-immunised children requires accurate knowledge of the underlying factors. We therefore developed and tested a model of childhood immunisation that includes individual, community and country-level characteristics. Method and Findings: We conducted multilevel logistic regression analysis of Demographic and Health Survey data for 27,094 children aged 12–23 months, nested within 8,546 communities from 24 countries in sub-Saharan Africa. According to the intra-country and intra-community correlation coefficient implied by the estimated intercept component variance, 21% and 32% of the variance in unimmunised children were attributable to country- and community-level factors respectively. Children born to mothers (OR 1.35, 95%CI 1.18 to 1.53) and fathers (OR 1.13, 95%CI 1.12 to 1.40) with no formal education were more likely to be unimmunised than those born to parents with secondary or higher education. Children from the poorest households were 36% more likely to be unimmunised than counterparts from the richest households. Maternal access to media significantly reduced the odds of children being unimmunised (OR 0.94, 95%CI 0.94 to 0.99). Mothers with health seeking behaviours were less likely to have unimmunised children (OR 0.56, 95%CI 0.54 to 0.58). However, children from urban areas (OR 1.12, 95% CI 1.01 to 1.23), communities with high illiteracy rates (OR 1.13, 95% CI 1.05 to 1.23), and countries with high fertility rates (OR 4.43, 95% CI 1.04 to 18.92) were more likely to be unimmunised. Conclusion: We found that individual and contextual factors were associated with childhood immunisation, suggesting that public health programmes designed to improve coverage of childhood immunisation should address people, and the communities and societies in which they live

Comparison of infant malaria incidence in districts of Maputo province, Mozambique

<p>Abstract</p> <p>Background</p> <p>Malaria is one of the principal health problems in Mozambique, representing 48% of total external consultations and 63% of paediatric hospital admissions in rural and general hospitals with 26.7% of total mortality. <it>Plasmodium falciparum </it>is responsible for 90% of all infections being also the species associated with most severe cases. The aim of this study was to identify zones of high malaria risk, showing their spatially and temporal pattern.</p> <p>Methods</p> <p>Space and time Poison model for the analysis of malaria data is proposed. This model allows for the inclusion of environmental factors: rainfall, temperature and humidity as predictor variables. Modelling and inference use the fully Bayesian approach via Markov Chain Monte Carlo (MCMC) simulation techniques. The methodology is applied to analyse paediatric data arising from districts of Maputo province, Mozambique, between 2007 and 2008.</p> <p>Results</p> <p>Malaria incidence risk is greater for children in districts of Manhiça, Matola and Magude. Rainfall and humidity are significant predictors of malaria incidence. The risk increased with rainfall (relative risk - RR: .006761, 95% interval: .001874, .01304), and humidity (RR: .049, 95% interval: .03048, .06531). Malaria incidence was found to be independent of temperature.</p> <p>Conclusions</p> <p>The model revealed a spatial and temporal pattern of malaria incidence. These patterns were found to exhibit a stable malaria transmission in most non-coastal districts. The findings may be useful for malaria control, planning and management.</p

Cellular Radiosensitivity: How much better do we understand it?

Purpose: Ionizing radiation exposure gives rise to a variety of lesions in DNA that result in genetic instability and potentially tumorigenesis or cell death. Radiation extends its effects on DNA by direct interaction or by radiolysis of H2O that generates free radicals or aqueous electrons capable of interacting with and causing indirect damage to DNA. While the various lesions arising in DNA after radiation exposure can contribute to the mutagenising effects of this agent, the potentially most damaging lesion is the DNA double strand break (DSB) that contributes to genome instability and/or cell death. Thus in many cases failure to recognise and/or repair this lesion determines the radiosensitivity status of the cell. DNA repair mechanisms including homologous recombination (HR) and non-homologous end-joining (NHEJ) have evolved to protect cells against DNA DSB. Mutations in proteins that constitute these repair pathways are characterised by radiosensitivity and genome instability. Defects in a number of these proteins also give rise to genetic disorders that feature not only genetic instability but also immunodeficiency, cancer predisposition, neurodegeneration and other pathologies. Conclusions: In the past fifty years our understanding of the cellular response to radiation damage has advanced enormously with insight being gained from a wide range of approaches extending from more basic early studies to the sophisticated approaches used today. In this review we discuss our current understanding of the impact of radiation on the cell and the organism gained from the array of past and present studies and attempt to provide an explanation for what it is that determines the response to radiation

Natural Host Genetic Resistance to Lentiviral CNS Disease: A Neuroprotective MHC Class I Allele in SIV-Infected Macaques

Human immunodeficiency virus (HIV) infection frequently causes neurologic disease even with anti-retroviral treatment. Although associations between MHC class I alleles and acquired immunodeficiency syndrome (AIDS) have been reported, the role MHC class I alleles play in restricting development of HIV-induced organ-specific diseases, including neurologic disease, has not been characterized. This study examined the relationship between expression of the MHC class I allele Mane-A*10 and development of lentiviral-induced central nervous system (CNS) disease using a well-characterized simian immunodeficiency (SIV)/pigtailed macaque model. The risk of developing CNS disease (SIV encephalitis) was 2.5 times higher for animals that did not express the MHC class I allele Mane-A*10 (P = 0.002; RR = 2.5). Animals expressing the Mane-A*10 allele had significantly lower amounts of activated macrophages, SIV RNA, and neuronal dysfunction in the CNS than Mane-A*10 negative animals (P<0.001). Mane-A*10 positive animals with the highest CNS viral burdens contained SIV gag escape mutants at the Mane-A*10-restricted KP9 epitope in the CNS whereas wild type KP9 sequences dominated in the brain of Mane-A*10 negative animals with comparable CNS viral burdens. These concordant findings demonstrate that particular MHC class I alleles play major neuroprotective roles in lentiviral-induced CNS disease

Influenza H5 Hemagglutinin DNA Primes the Antibody Response Elicited by the Live Attenuated Influenza A/Vietnam/1203/2004 Vaccine in Ferrets

Priming immunization plays a key role in protecting individuals or populations to influenza viruses that are novel to humans. To identify the most promising vaccine priming strategy, we have evaluated different prime-boost regimens using inactivated, DNA and live attenuated vaccines in ferrets. Live attenuated influenza A/Vietnam/1203/2004 (H5N1) candidate vaccine (LAIV, VN04 ca) primed ferrets efficiently while inactivated H5N1 vaccine could not prime the immune response in seronegative ferrets unless an adjuvant was used. However, the H5 HA DNA vaccine alone was as successful as an adjuvanted inactivated VN04 vaccine in priming the immune response to VN04 ca virus. The serum antibody titers of ferrets primed with H5 HA DNA followed by intranasal vaccination of VN04 ca virus were comparable to that induced by two doses of VN04 ca virus. Both LAIV-LAIV and DNA-LAIV vaccine regimens could induce antibody responses that cross-neutralized antigenically distinct H5N1 virus isolates including A/HongKong/213/2003 (HK03) and prevented nasal infection of HK03 vaccine virus. Thus, H5 HA DNA vaccination may offer an alternative option for pandemic preparedness