Sodium channel slow inactivation interferes with open channel block

Mutations in the voltage-gated sodium channel Nav1.7 are linked to inherited pain syndromes such as erythromelalgia (IEM) and paroxysmal extreme pain disorder (PEPD). PEPD mutations impair Nav1.7 fast inactivation and increase persistent currents. PEPD mutations also increase resurgent currents, which involve the voltage-dependent release of an open channel blocker. In contrast, IEM mutations, whenever tested, leave resurgent currents unchanged. Accordingly, the IEM deletion mutation L955 (ΔL955) fails to produce resurgent currents despite enhanced persistent currents, which have hitherto been considered a prerequisite for resurgent currents. Additionally, ΔL955 exhibits a prominent enhancement of slow inactivation (SI). We introduced mutations into Nav1.7 and Nav1.6 that either enhance or impair SI in order to investigate their effects on resurgent currents. Our results show that enhanced SI is accompanied by impaired resurgent currents, which suggests that SI may interfere with open-channel block

Mismatched single stranded antisense oligonucleotides can induce efficient dystrophin splice switching

<p>Abstract</p> <p>Background</p> <p>Antisense oligomer induced exon skipping aims to reduce the severity of Duchenne muscular dystrophy by redirecting splicing during pre-RNA processing such that the causative mutation is by-passed and a shorter but partially functional Becker muscular dystrophy-like dystrophin isoform is produced. Normal exons are generally targeted to restore the dystrophin reading frame however, an appreciable subset of dystrophin mutations are intra-exonic and therefore have the potential to compromise oligomer efficiency, necessitating personalised oligomer design for some patients. Although antisense oligomers are easily personalised, it remains unclear whether all patient polymorphisms within antisense oligomer target sequences will require the costly process of producing and validating patient specific compounds.</p> <p>Methods</p> <p>Here we report preclinical testing of a panel of splice switching antisense oligomers, designed to excise exon 25 from the dystrophin transcript, in normal and dystrophic patient cells. These patient cells harbour a single base insertion in exon 25 that lies within the target sequence of an oligomer shown to be effective at removing exon 25.</p> <p>Results</p> <p>It was anticipated that such a mutation would compromise oligomer binding and efficiency. However, we show that, despite the mismatch an oligomer, designed and optimised to excise exon 25 from the normal dystrophin mRNA, removes the mutated exon 25 more efficiently than the mutation-specific oligomer.</p> <p>Conclusion</p> <p>This raises the possibility that mismatched AOs could still be therapeutically applicable in some cases, negating the necessity to produce patient-specific compounds.</p

Rapid urbanisation and security: Holistic approach to enhancing security of urban spaces

Rapid urbanisation, particularly driven by rural-urban migration, can pose a wide range of security challenges in the global south and global north. The management of such a transition, in terms of the provision of social goods and quality of life raises significant challenges. Security of contemporary urban environments has become more complex due to a greater range of risk drivers, many of which can be exacerbated by the observed and portended impacts of climate change. This chapter outlines the phenomena underlying the transition to urbanisation - and the security challenges that have been exacerbated by these transitions. In doing so this work a holistic approach to security and highlights a gradual trend in the increased securitisation of issues (such as climate change) that in the past were not considered part of typical ‘security’ dialogues. It also introduces a decision support framework that can aid a broad range of stakeholders in making decisions about the enhancement of security of urban spaces in a context of multiple threats exacerbated by these new security challenges

A Novel System of Cytoskeletal Elements in the Human Pathogen Helicobacter pylori

Pathogenicity of the human pathogen Helicobacter pylori relies upon its capacity to adapt to a hostile environment and to escape from the host response. Therefore, cell shape, motility, and pH homeostasis of these bacteria are specifically adapted to the gastric mucus. We have found that the helical shape of H. pylori depends on coiled coil rich proteins (Ccrp), which form extended filamentous structures in vitro and in vivo, and are differentially required for the maintenance of cell morphology. We have developed an in vivo localization system for this pathogen. Consistent with a cytoskeleton-like structure, Ccrp proteins localized in a regular punctuate and static pattern within H. pylori cells. Ccrp genes show a high degree of sequence variation, which could be the reason for the morphological diversity between H. pylori strains. In contrast to other bacteria, the actin-like MreB protein is dispensable for viability in H. pylori, and does not affect cell shape, but cell length and chromosome segregation. In addition, mreB mutant cells displayed significantly reduced urease activity, and thus compromise a major pathogenicity factor of H. pylori. Our findings reveal that Ccrp proteins, but not MreB, affect cell morphology, while both cytoskeletal components affect the development of pathogenicity factors and/or cell cycle progression

Proteomic Analyses of Host and Pathogen Responses during Bovine Mastitis

The pursuit of biomarkers for use as clinical screening tools, measures for early detection, disease monitoring, and as a means for assessing therapeutic responses has steadily evolved in human and veterinary medicine over the past two decades. Concurrently, advances in mass spectrometry have markedly expanded proteomic capabilities for biomarker discovery. While initial mass spectrometric biomarker discovery endeavors focused primarily on the detection of modulated proteins in human tissues and fluids, recent efforts have shifted to include proteomic analyses of biological samples from food animal species. Mastitis continues to garner attention in veterinary research due mainly to affiliated financial losses and food safety concerns over antimicrobial use, but also because there are only a limited number of efficacious mastitis treatment options. Accordingly, comparative proteomic analyses of bovine milk have emerged in recent years. Efforts to prevent agricultural-related food-borne illness have likewise fueled an interest in the proteomic evaluation of several prominent strains of bacteria, including common mastitis pathogens. The interest in establishing biomarkers of the host and pathogen responses during bovine mastitis stems largely from the need to better characterize mechanisms of the disease, to identify reliable biomarkers for use as measures of early detection and drug efficacy, and to uncover potentially novel targets for the development of alternative therapeutics. The following review focuses primarily on comparative proteomic analyses conducted on healthy versus mastitic bovine milk. However, a comparison of the host defense proteome of human and bovine milk and the proteomic analysis of common veterinary pathogens are likewise introduced

Dynamics of Co-Transcriptional Pre-mRNA Folding Influences the Induction of Dystrophin Exon Skipping by Antisense Oligonucleotides

Antisense oligonucleotides (AONs) mediated exon skipping offers potential therapy for Duchenne muscular dystrophy. However, the identification of effective AON target sites remains unsatisfactory for lack of a precise method to predict their binding accessibility. This study demonstrates the importance of co-transcriptional pre-mRNA folding in determining the accessibility of AON target sites for AON induction of selective exon skipping in DMD. Because transcription and splicing occur in tandem, AONs must bind to their target sites before splicing factors. Furthermore, co-transcriptional pre-mRNA folding forms transient secondary structures, which redistributes accessible binding sites. In our analysis, to approximate transcription elongation, a “window of analysis” that included the entire targeted exon was shifted one nucleotide at a time along the pre-mRNA. Possible co-transcriptional secondary structures were predicted using the sequence in each step of transcriptional analysis. A nucleotide was considered “engaged” if it formed a complementary base pairing in all predicted secondary structures of a particular step. Correlation of frequency and localisation of engaged nucleotides in AON target sites accounted for the performance (efficacy and efficiency) of 94% of 176 previously reported AONs. Four novel insights are inferred: (1) the lowest frequencies of engaged nucleotides are associated with the most efficient AONs; (2) engaged nucleotides at 3′ or 5′ ends of the target site attenuate AON performance more than at other sites; (3) the performance of longer AONs is less attenuated by engaged nucleotides at 3′ or 5′ ends of the target site compared to shorter AONs; (4) engaged nucleotides at 3′ end of a short target site attenuates AON efficiency more than at 5′ end

Meta-analysis of the effects of predation on animal prey abundance: evidence from UK vertebrates

Background: Controlling vertebrate predators is one of the most widespread forms of wildlife management and it continues to cause conflict between stakeholders worldwide. It is important for managers and policy-makers to make decisions on this issue that are based on the best available scientific evidence. Therefore, it is first important to understand if there is indeed an impact of vertebrate predators on prey, and then to quantify this impact. Methodology/Principal Findings: Using the UK as a case study, we use a meta-analytical approach to review the available evidence to assess the effect of vertebrate predation on animal prey abundance. We find a significant effect of predators on prey abundance across our studies. On average, there is a 1.6 fold increase in prey abundance in the absence of predation. However, we show significant heterogeneity in effect sizes, and discuss how the method of predator control, whether the predator is native or non-native, and aspects of study design, may be potential causes. Conclusions/Significance: Our results allow some cautious policy recommendations to be made regarding the management of predator and prey populations. Meta-analysis is an important tool for understanding general patterns in the effect of predators on prey abundance across studies. Such an approach is especially valuable where management decisions need to be made in the absence of site-specific information

A new model for the study of high-K(+)-induced preconditioning in cultured neurones: role of N-methyl-d-aspartate and alpha7-nicotinic acetylcholine receptors.

Spreading depression (SD), whether elicited by local application of high K(+) medium to the cortical surface or by other stimuli, can increase the brain's tolerance to a subsequent, severe ischaemic insult in vivo, a phenomenon termed preconditioning. Herein, we have developed and validated a robust in vitro protocol for high-K(+)-preconditioning of cultured neurones. This new model is especially appropriate to unravel the molecular mechanisms underlying neuronal preconditioning and subsequent ischaemic tolerance. With this new, optimised preparation, preconditioning was found to be dependent upon culture day in vitro, cell density, K(+) concentration and duration of treatment. Finally, preconditioning was shown to be dependent upon N-methyl-d-aspartate (NMDA), CAM-kinase II signalling and alpha7-nicotinic (alpha7 nACh) receptor function, which is analogous to in vivo preconditioning induced by various stimuli