Establishing RNAi in a non-model organism: The Antarctic nematode Panagrolaimus sp. DAW1

The Antarctic nematode Panagrolaimus sp. DAW1 is one of the only organisms known to survive extensive intracellular freezing throughout its tissues. Although the physiological mechanisms of this extreme adaptation are partly understood, the molecular mechanisms remain largely unknown. RNAi is a method that allows the examination of gene function in a direct, targeted manner, by knocking out specific mRNAs and revealing the effects on the phenotype. In this study we have explored the viability of RNAi in Panagrolaimus sp. DAW1. In the first trial, nematodes were fed E. coli expressing Panagrolaimus sp. DAW1 dsRNA of the embryonic lethal genes rps-2 and dhc, and the blister gene duox. Pd-rps-2(RNAi)-treated nematodes showed a significant decrease in larval hatching. However, qPCR showed no significant decrease in the amount of rps-2 mRNA in Pd-rps-2(RNAi)-treated animals. Several soaking protocols for dsRNA uptake were investigated using the fluorescent dye FITC. Desiccation-enhanced soaking showed the strongest uptake of FITC and resulted in a significant and consistent decrease of mRNA levels of two of the four tested genes (rps-2 and tps-2a), suggesting effective uptake of dsRNA-containing solution by the nematode. These findings suggest that RNAi by desiccation-enhanced soaking is viable in Panagrolaimus sp. DAW1 and provide the first functional genomic approach to investigate freezing tolerance in this non-model organism. RNAi, in conjunction with qPCR, can be used to screen for candidate genes involved in intracellular freezing tolerance in Panagrolaimus sp. DAW1

Molecular analysis of the cold tolerant Antarctic nematode, Panagrolaimus davidi

Isolated and established in culture from the Antarctic in 1988, the nematode Panagrolaimus davidi has proven to be an ideal model for the study of adaptation to the cold. Not only is it the best-documented example of an organism surviving intracellular freezing but it is also able to undergo cryoprotective dehydration. As part of an ongoing effort to develop a molecular understanding of this remarkable organism, we have assembled both a transcriptome and a set of genomic scaffolds. We provide an overview of the transcriptome and a survey of genes involved in temperature stress. We also explore, in silico, the possibility that P. davidi will be susceptible to an environmental RNAi response, important for further functional studies

Molecular snapshot of an intracellular freezing event in an Antarctic nematode

The Antarctic nematode, Panagrolaimus sp. DAW1 (formerly called Panagrolaimus davidi), is the best documented example of an organism able to survive intracellular ice formation in all of its compartments. Not only is it able to survive such extreme physiological disruption, but it is able to produce progeny once thawed from such a state. In addition, under slower rates, or less extreme degrees, of cooling, its body remains unfrozen and the vapour pressure difference between the supercooled body fluids and the surrounding ice leads to a process termed cryoprotective dehydration. In contrast to a fairly large body of work in building up our molecular understanding of cryoprotective dehydration, no comparable work has been undertaken on intracellular freezing. This paper describes an experiment subjecting cultures of Panagrolaimus sp. DAW1 to a range of temperatures including a rapid descent to −10 °C, in a medium just prior to, and after, freezing. Through deep sequencing of RNA libraries we have gained a snapshot of which genes are highly abundant when P. sp. DAW1 is undergoing an intracellular freezing event. The onset of freezing correlated with a high production of genes involved in cuticle formation and subsequently, after 24 h in a frozen state, protease production. In addition to the mapping of RNA sequencing, we have focused on a select set of genes arising both from the expression profiles, as well as implicated from other cold tolerance studies, to undertake qPCR. Among the most abundantly represented transcripts in the RNA mapping is the zinc-metalloenzyme, neprilysin, which also shows a particularly strong upregulated signal through qPCR once the nematodes have frozen

Effects of laser shock peening on the mechanisms of fatigue short crack initiation and propagation of AA7075-T651

A laser shock peening (LSP) treatment was performed on AA7075-T651 for maximum fatigue improvement. Surface and microstructural characterisation techniques (micro-hardness, SEM-EBSD, contact-profilometry) showed LSP surface modification was limited, and LSP generated deep compressive residual stresses above -300MPa. Fatigue testing showed a two-order magnitude increase in overall life, due to the mechanism of crack initiation changing from surface second-phase particles to subsurface crack initiation dependent on the local stress field. Modelling highlights the sensitive balance between surface roughness (including LSP-induced pits) and residual stress on the micro-mechanism of crack initiation, and how this can be used to maximise fatigue life extension

Investigating trehalose synthesis genes after cold acclimation in the Antarctic nematode Panagrolaimus sp. DAW1

Panagrolaimus sp. DAW1 is a freeze-tolerant Antarctic nematode which survives extensive intracellular ice formation. The molecular mechanisms of this extreme adaptation are still poorly understood. We recently showed that desiccation-enhanced RNAi-soaking can be used - in conjunction with qPCR - to screen for phenotypes associated with reduced expression of candidate genes in Panagrolaimus sp. DAW1. Here we present the use of this approach to investigate the role of trehalose synthesis genes in this remarkable organism. Previous studies have showed that acclimating Panagrolaimus sp. DAW1 at 5 °C before freezing or desiccation substantially enhances survival. In this study the expression of tps-2 and other genes associated with trehalose metabolism, as well as lea-1, hsp-70 and gpx-1 in cold-acclimated and non-acclimated nematodes was analyzed using qPCR. Pd-tps-2 and Pd-lea-1 were significantly up-regulated after cold-acclimation, indicating an inducible expression in the cold-adaptation of Panagrolaimus sp. DAW1. The role of trehalose synthesis genes in Panagrolaimus sp. DAW1 was further investigated by RNAi. Compared to the controls, Pd-tps-2a(RNAi)-treated and cold-acclimated nematodes showed a significant decrease in mRNA, but no change in trehalose content or freezing survival. The involvement of two other trehalose synthesis genes (tps-2b and gob-1) was also investigated. These findings provide the first functional genomic investigation of trehalose synthesis genes in the non-model organism Panagrolaimus sp. DAW1. The presence of several trehalose synthesis genes with different RNAi sensitivities suggests the existence of multiple backup systems in Panagrolaimus sp. DAW1, underlining the importance of this sugar in preparation for freezing

Horseplay, care and hands on hard work: gendered strategies of a project manager on a construction site

The discourse of managerial expertise favours rational analysis and masculine ideals but contemporary management literature also recognises the value of well-being and employee voice in the workplace. Drawing upon narrative analysis of interview data, we share unique insights into the lived experiences of Laura, one female project manager who recently managed a construction site in the Midlands in the UK. In contrast to previous research which indicates that female managers tend to conform to quite a traditional set of gender behaviours, Laura embraces a range of workplace appropriate gendered strategies, such as hard work and horseplay, together with sensitivity and caring. She draws from this mix of gendered strategies in negotiating between two different discourses of construction; one professional and one tough and practical. Her behaviour both reproduces the masculine ideals (through horseplay and heroic management) and opens up possibilities for modernising construction management (by caring). It is this combination of strategies that is at the heart of tacit expertise for Laura. Theoretically, the discussion adds to the development of a more nuanced understanding of management expertise as situated and person specific knowledge that draws on both the explicit and tacit. Specifically, the centrality of gendered strategies beyond the masculine ideals to success on site is highlighted

Erratum to: Search for diboson resonances in hadronic final states in 139 fb−1 of pp collisions at s√ = 13 TeV with the ATLAS detector

A mistake was identified for the paper [1] in the treatment of the radion [2] cross-sections, which resulted in multiple changes.(undefined)info:eu-repo/semantics/publishedVersio

Measurement of the cross section for isolated-photon plus jet production in pp collisions at √s=13 TeV using the ATLAS detector

The dynamics of isolated-photon production in association with a jet in proton–proton collisions at a centre-of-mass energy of 13 TeV are studied with the ATLAS detector at the LHC using a dataset with an integrated luminosity of 3.2 fb−1. Photons are required to have transverse energies above 125 GeV. Jets are identified using the anti- algorithm with radius parameter and required to have transverse momenta above 100 GeV. Measurements of isolated-photon plus jet cross sections are presented as functions of the leading-photon transverse energy, the leading-jet transverse momentum, the azimuthal angular separation between the photon and the jet, the photon–jet invariant mass and the scattering angle in the photon–jet centre-of-mass system. Tree-level plus parton-shower predictions from Sherpa and Pythia as well as next-to-leading-order QCD predictions from Jetphox and Sherpa are compared to the measurements