A Serratia marcescens PigP Homolog Controls Prodigiosin Biosynthesis, Swarming Motility and Hemolysis and Is Regulated by cAMP-CRP and HexS

Swarming motility and hemolysis are virulence-associated determinants for a wide array of pathogenic bacteria. The broad host-range opportunistic pathogen Serratia marcescens produces serratamolide, a small cyclic amino-lipid, that promotes swarming motility and hemolysis. Serratamolide is negatively regulated by the transcription factors HexS and CRP. Positive regulators of serratamolide production are unknown. Similar to serratamolide, the antibiotic pigment, prodigiosin, is regulated by temperature, growth phase, HexS, and CRP. Because of this co-regulation, we tested the hypothesis that a homolog of the PigP transcription factor of the atypical Serratia species ATCC 39006, which positively regulates prodigiosin biosynthesis, is also a positive regulator of serratamolide production in S. marcescens. Mutation of pigP in clinical, environmental, and laboratory strains of S. marcescens conferred pleiotropic phenotypes including the loss of swarming motility, hemolysis, and severely reduced prodigiosin and serratamolide synthesis. Transcriptional analysis and electrophoretic mobility shift assays place PigP in a regulatory pathway with upstream regulators CRP and HexS. The data from this study identifies a positive regulator of serratamolide production, describes novel roles for the PigP transcription factor, shows for the first time that PigP directly regulates the pigment biosynthetic operon, and identifies upstream regulators of pigP. This study suggests that PigP is important for the ability of S. marcescens to compete in the environment. © 2013 Shanks et al

Whole-transcriptome, high-throughput RNA sequence analysis of the bovine macrophage response to Mycobacterium bovis infection in vitro

BACKGROUND: Mycobacterium bovis, the causative agent of bovine tuberculosis, is an intracellular pathogen that can persist inside host macrophages during infection via a diverse range of mechanisms that subvert the host immune response. In the current study, we have analysed and compared the transcriptomes of M. bovis-infected monocyte-derived macrophages (MDM) purified from six Holstein-Friesian females with the transcriptomes of non-infected control MDM from the same animals over a 24 h period using strand-specific RNA sequencing (RNA-seq). In addition, we compare gene expression profiles generated using RNA-seq with those previously generated by us using the high-density Affymetrix® GeneChip® Bovine Genome Array platform from the same MDM-extracted RNA. RESULTS: A mean of 7.2 million reads from each MDM sample mapped uniquely and unambiguously to single Bos taurus reference genome locations. Analysis of these mapped reads showed 2,584 genes (1,392 upregulated; 1,192 downregulated) and 757 putative natural antisense transcripts (558 upregulated; 119 downregulated) that were differentially expressed based on sense and antisense strand data, respectively (adjusted P-value ≤ 0.05). Of the differentially expressed genes, 694 were common to both the sense and antisense data sets, with the direction of expression (i.e. up- or downregulation) positively correlated for 693 genes and negatively correlated for the remaining gene. Gene ontology analysis of the differentially expressed genes revealed an enrichment of immune, apoptotic and cell signalling genes. Notably, the number of differentially expressed genes identified from RNA-seq sense strand analysis was greater than the number of differentially expressed genes detected from microarray analysis (2,584 genes versus 2,015 genes). Furthermore, our data reveal a greater dynamic range in the detection and quantification of gene transcripts for RNA-seq compared to microarray technology. CONCLUSIONS: This study highlights the value of RNA-seq in identifying novel immunomodulatory mechanisms that underlie host-mycobacterial pathogen interactions during infection, including possible complex post-transcriptional regulation of host gene expression involving antisense RNA

Human exercise-induced circulating progenitor cell mobilization is nitric oxide-dependent and is blunted in South Asian men

This article is available open access through the publisher’s website. Copyright @ 2010 American Heart Foundation.Objective— Circulating progenitor cells (CPC) have emerged as potential mediators of vascular repair. In experimental models, CPC mobilization is critically dependent on nitric oxide (NO). South Asian ethnicity is associated with reduced CPC. We assessed CPC mobilization in response to exercise in Asian men and examined the role of NO in CPC mobilization per se. Methods and Results— In 15 healthy, white European men and 15 matched South Asian men, CPC mobilization was assessed during moderate-intensity exercise. Brachial artery flow-mediated vasodilatation was used to assess NO bioavailability. To determine the role of NO in CPC mobilization, identical exercise studies were performed during intravenous separate infusions of saline, the NO synthase inhibitor l-NMMA, and norepinephrine.  Flow-mediated vasodilatation (5.8%±0.4% vs 7.9%±0.5%; P=0.002) and CPC mobilization (CD34+/KDR+ 53.2% vs 85.4%; P=0.001; CD133+/CD34+/KDR+ 48.4% vs 73.9%; P=0.05; and CD34+/CD45− 49.3% vs 78.4; P=0.006) was blunted in the South Asian group. CPC mobilization correlated with flow-mediated vasodilatation and l-NMMA significantly reduced exercise-induced CPC mobilization (CD34+/KDR+ −3.3% vs 68.4%; CD133+/CD34+/KDR+ 0.7% vs 71.4%; and CD34+/CD45− −30.5% vs 77.8%; all P<0.001). Conclusion— In humans, NO is critical for CPC mobilization in response to exercise. Reduced NO bioavailability may contribute to imbalance between vascular damage and repair mechanisms in South Asian men.British Heart Foundatio

A novel cell culture model for studying differentiation and apoptosis in the mouse mammary gland.

BACKGROUND: This paper describes the derivation and characterization of a novel, conditionally immortal mammary epithelial cell line named KIM-2. These cells were derived from mid-pregnant mammary glands of a mouse harbouring one to two copies of a transgene comprised of the ovine beta-lactoglobulin milk protein gene promoter, driving expression of a temperature-sensitive variant of simian virus-40 (SV40) large T antigen (T-Ag). RESULTS: KIM-2 cells have a characteristic luminal epithelial cell morphology and a stable, nontransformed phenotype at the semipermissive temperature of 37 degrees C. In contrast, at the permissive temperature of 33 degrees C the cells have an elongated spindle-like morphology and become transformed after prolonged culture. Differentiation of KIM-2 cells at 37 degrees C, in response to lactogenic hormones, results in the formation of polarized dome-like structures with tight junctions. This is accompanied by expression of the milk protein genes that encode beta-casein and whey acidic protein (WAP), and activation of the prolactin signalling molecule, signal transducer and activator of transcription (STAT)5. Fully differentiated KIM-2 cultures at 37 degrees C become dependent on lactogenic hormones for survival and undergo extensive apoptosis upon hormone withdrawal, as indicated by nuclear morphology and flow cytometric analysis. KIM-2 cells can be genetically modified by stable transfection and clonal lines isolated that retain the characteristics of untransfected cells. CONCLUSION: KIM-2 cells are a valuable addition, therefore, to currently available lines of mammary epithelial cells. Their capacity for extensive differentiation in the absence of exogenously added basement membrane, and ability to undergo apoptosis in response to physiological signals will provide an invaluable model system for the study of signal transduction pathways and transcriptional regulatory mechanisms that control differentiation and involution in the mammary gland

Fructosamine; is the current interest in alternative glycaemic markers justified?

This is an accepted manuscript of an article published by Wiley in Diabetic Medicine on 27/03/2015, available online: https://doi.org/10.1111/dme.12767 The accepted version of the publication may differ from the final published version.Published versio

Vaccines for COVID-19

Since the emergence of COVID-19, caused by the SARS-CoV-2 virus, at the end of 2019 there has been an explosion of vaccine development. By the 1st September 2020, a staggering number of vaccines (over 200) had started pre-clinical development of which 39 had entered clinical trials, including some approaches that have not previously been licensed for human vaccines. Vaccines have been widely considered as part of the exit strategy to enable the return to previous patterns of working, schooling and socialising. Importantly, to effectively control the COVID-19 pandemic, production needs to be scaled up from a small number of pre-clinical doses to enough filled vials to immunise the world's population, which requires close engagement with manufacturers and regulators. It will require a global effort to control the virus, necessitating equitable access for all countries to effective vaccines. This review explores the immune responses required to protect against SARS-CoV-2 and the potential for vaccine-induced immunopathology. It describes the profile of the different platforms and the advantages and disadvantages of each approach. The review also addresses the critical steps between promising pre-clinical leads and manufacturing at scale. The issues faced during this pandemic and the platforms being developed to address it will be invaluable for future outbreak control. Nine months after the outbreak began, we are at a point where pre-clinical and early clinical data is being generated for the vaccines, an overview of this important area will help our understanding of the next phases

Investigating the consequences of asymmetric endoplasmic reticulum inheritance in Saccharomyces cerevisiae under stress using a combination of single cell measurements and mathematical modelling

Adaptation allows organisms to maintain a constant internal environment, which is optimised for growth. The unfolded protein response (UPR) is an example of a feedback loop that maintains endoplasmic reticulum (ER) homeostasis, and is characteristic of how adaptation is often mediated by transcriptional networks. The more recent discovery of asymmetric division in maintaining ER homeostasis, however, is an example of how alternative non-transcriptional pathways can exist, but are overlooked by gold standard transcriptomic or proteomic population-based assays. In this study, we have used a combination of fluorescent reporters, flow cytometry and mathematical modelling to explore the relative roles of asymmetric cell division and the UPR in maintaining ER homeostasis. Under low ER stress, asymmetric division leaves daughter cells with an ER deficiency, necessitating activation of the UPR and prolonged cell cycle during which they can recover ER functionality before growth. Mathematical analysis of and simulation results from our mathematical model reinforce the experimental observations that low ER stress primarily impacts the growth rate of the daughter cells. These results demonstrate the interplay between homeostatic pathways and the importance of exploring sub-population dynamics to understand population adaptation to quantitatively different stresses

From venture idea to venture formation:The role of sensemaking, sensegiving and sense receiving

This article explores the sensemaking processes entrepreneurs use when transitioning between venture ideas and venture formation. Adopting a sensemaking/sensegiving approach and utilising an interpretivist methodology, we use sensemaking to analyse the entrepreneurial journey of four diverse entrepreneurs. In so doing, we make three contributions: first, we locate the early stages of the entrepreneurial context as a primary site where sensemaking occurs as entrepreneurs deal with the differences between expectations and reality. Second, we show how sensemaking occurs when entrepreneurs build a causal map of the problem they wish to address and how social exchanges are crucial as entrepreneurs then refine that idea with other sensegivers. Finally, we extend scholarly understanding through explaining the ways in which sensemaking, sensegiving and sense receiving contribute to the entrepreneurs' decision to act and create a new venture