The importance of specific tryptophans to UVR8 function: an intrinsic chromophore for a UV-B photoreceptor

Although sessile organisms, unable to run away from danger, plants are well adapted to the potential harmful effects of sunlight’s high energy photons within the UV-B wavelength range (280-315 nm). For instance they are able to, among other things; produce their own sunscreen to counter any damage to their proteins, lipids and DNA. Plants of course depend on light as a source of energy for photosynthesis but also use specific wavelengths within the electromagnetic spectrum in a number of ways to act as an informational signal, including UV-B wavelengths, which can induce photomorphogenic responses that allow adaptation and survival for plants in the ever-changing environmental conditions they inhabit. It is now well established in plants that there are more than two pathways operating in response to different wavelengths and fluence rates of UV-B. In response to high, potentially damaging UV-B levels plants utilize a non-specific pathway which overlaps with other stress pathways such as pathogen attack and wounding by, for example, herbivores. And in response to low non-damaging UV-B levels plants utilize the UV-B specific photomorphogenic pathways which bring about acclimation, preparing the plant for potential higher doses and actively promoting plant survival (Jenkins and Brown, 2007). A number of photoreceptors have been identified in plants which act throughout the electromagnetic spectrum, but only in the last year has one been discovered operating at UV-B wavelengths. In fact until then no UV-B- specific photoreceptor had been found in any organism and it was not known how plants perceive UV-B light to initiate photomorphogenic responses. Over the last decade evidence was mounting in favour of the most upstream component of the UV-B photomorphogenic pathway and the only UV-B specific component, UVR8 (UV-RESISTANCE LOCUS 8) as being a UV-B photoreceptor. Now it has been demonstrated in plants to be a bona fide UV-B photoreceptor and to perceive UV-B by a novel mechanism (Rizzini et al., 2011, Christie et al., 2012, Wu et al., 2012). It has been demonstrated upon UV-B irradiation that UVR8 can dissociate from a homodimer to a monomer in vivo and in vitro. And unlike other conventional photoreceptors, which use a chromophore to detect specific wavelengths of light, UVR8 uses tryptophan residues found within its protein structure to carry out photoperception. When UV-B is detected via specific tryptophan residues found within the dimeric UVR8 protein, the energy is captured and used to cause disruption and breakage of several salt bridges between adjacent homodimers causing monomerization and subsequently leading to interaction with COP1 (CONSTITUTIVELY PHOTOMORPHOGENIC 1), nuclear accumulation and signal transduction (Christie et al., 2012; Wu et al., 2012; Favory et al 2009; Kaiserli and Jenkins 2007; Brown et al., 2005). Once UVR8 is in its active form it can then regulate the transcription of a number of UV-B responsive photomorphogenic genes allowing the plant to acclimate to counteract any future potential damage, which in turn promotes the plant’s survival and reproduction (Brown et al., 2005; Oravecz et al., 2006; Favory et al., 2009). When I first started my studies UVR8 was implicated in UV-B responses but it was unknown if it functioned as a photoreceptor. The purpose of my Ph.D was to determine if UVR8 was a UV-B photoreceptor and if so how it perceives UV-B. And more specifically, to address the question: can tryptophan residues within its structure act as an intrinsic chromophore? To investigate this aim I firstly used site directed mutagenesis to mutate specific and multiple tryptophan residues of the 14 found within UVR8’s structure to alanine, phenylalanine and tyrosine. Then I carried out transient expression studies in Nicotiana benthamiana to determine if the mutant protein tagged to GFP was stable and to determine if its subcellular localisation was affected. These UVR8 Trp mutant variants were further analyzed using yeast 2-hybrid assays (Y2H) to test for interaction with COP1, RUP1/RUP2 (REPRESSOR OF UV-B PHOTOMORPHOGENESIS) and also homodimerization. This allowed me to identify Trp mutant candidates to introduce transgenically into Arabidopsis and test further for their ability to complement the null mutant uvr8-1. The mutants were tested using a number of assays to check for monomer/dimer status, subcellular localisation, protein stability, COP1 interaction, photomorphogenic gene expression, hypocotyl inhibition and chromatin binding. Herein I present in vivo data in yeast and plants which shows, as reported by Rizzini et al. (2011), Christie et al. (2012) and Wu et al. (2012), that specific Trps, mainly W285 and W233 within the triad W233, W285, W337 have key roles in photoreception. W337 has a lesser role. These triad Trps, which are all in the conserved motif GWRHT, have now been shown in the UVR8 crystal structure to be brought into close proximity (Christie et al., 2012, Wu et al., 2012). The W285A mutant did not complement uvr8-1 and the W233A mutant only partially complemented, whereas W337A substantially complemented uvr8-1. And although all three Trp mutants constitutively interact with COP1 in planta before and after UV-B irradiation, this is not sufficient to rescue the uvr8-1 mutant for W285A and W233A, suggesting that although COP1 interaction is required for UV-B specific photomorphogenic responses it is not sufficient to mediate a response. Furthermore, for each of the triad mutants their dimer/monomer status is affected, and W285A is constitutively monomeric without being functional. Therefore, similar to COP1 interaction, monomerization on its own is not sufficient for UVR8 activation. In addition, I show that of the remaining 11 trps left of the 14 in total found within UVR8, some (W39, W144, W352) are important for structure and hence function, and the others (W92, 94, 196, 198, 250, 300, 302, 400) are not essential for function and/or structure. To further support the intrinsic Trp chromophore model of UVR8 I also present an action spectrum for dimer to monomer conversion for pure UVR8 protein in vitro from samples expressed and purified from E.coli. The spectrum closely resembles the absorption spectra of UVR8 and Trp in solution, with a maximum response at 280 nm. Moreover, the action spectrum partially resembles the in vivo UVR8 dependent HY5 (ELONGATED HYPOCOTYL 5) expression action spectrum published previously (Brown et al., 2009), although the in vivo HY5 study shows a substantial response at 300 nm, which this in vitro study lacks. Overall I show the importance of specific Trps to the UV-B photoreceptor UVR8 in yeast and in planta and demonstrate that W285 and W233 in particular are important in allowing UVR8 to function as a photoreceptor by acting as intrinsic chromophores

Computer-tailored physical activity behavior change interventions targeting adults: a systematic review

<p>Abstract</p> <p>Background</p> <p>Increasing physical activity is important in the promotion of better health. Computer-tailored behavior change programs have shown promise in changing lifestyle risk factors.</p> <p>Purpose</p> <p>To provide a narrative systematic review describing the range of evidence on 'second' and 'third' generation computer-tailored primary prevention interventions for physical activity, to determine their effectiveness and key characteristics of success. Unlike previous reviews, this review used specific criteria to measure the external validity of studies, was exclusive to primary prevention interventions in which tailoring was generated through an expert system, and excluded first generation computer-tailored interventions.</p> <p>Methods</p> <p>Computer-tailored intervention studies published from January 1996–2008 were identified through a search of five databases: Medline; Embase; PsycINFO; CINAHL; and All EBM Reviews and by examining reference lists of relevant articles.</p> <p>Results</p> <p>Seventeen articles were included, describing the evaluation of 16 interventions, ten of which found significant positive effects of the computer-tailored interventions on physical activity or weight reduction outcomes.</p> <p>Conclusion</p> <p>The evidence of effectiveness for computer-tailored physical activity interventions is inconclusive. They have potential to reach large groups of people however there is uncertainty whether reported effects are generalizable and sustained.</p

Your money or your life: Comparing judgements in trolley problems involving economic and emotional Harms, injury and death: Natalie gold et al

There is a long-standing debate in philosophy about whether it is morally permissible to harm one person in order to prevent a greater harm to others and, if not, what is the moral principle underlying the prohibition. Hypothetical moral dilemmas are used in order to probe moral intuitions. Philosophers use them to achieve a reflective equilibrium between intuitions and principles, psychologists to investigate moral decision-making processes. In the dilemmas, the harms that are traded off are almost always deaths. However, the moral principles and psychological processes are supposed to be broader than this, encompassing harms other than death. Further, if the standard pattern of intuitions is preserved in the domain of economic harm, then that would open up the possibility of studying behaviour in trolley problems using the tools of experimental economics. We report the results of two studies designed to test whether the standard patterns of intuitions are preserved when the domain and severity of harm are varied. Our findings show that the difference in moral intuitions between bystander and footbridge scenarios is replicated across different domains and levels of physical and non-physical harm, including economic harms

Engineering strong magnetoelectricity using a hexagonal 2D material on electron-doped hexagonal LuFeO3_3

Cubic perovskite-structure ABO$_3$ and A$_{1-x}$A$^{\prime}$$_x$BO$_3$-type oxides have been investigated extensively while their hexagonal-structure versions have received minimal attention, even though they are multiferroic and can form heterostructures with the manifold hexagonal two-dimensional materials. Hexagonal ferrites of the form RFeO$_3$, where R is yttrium or a rare-earth element such as Lu, Yb, etc., feature coupled ferroelectricity (FE) and weak-ferromagnetism (wFM), exhibiting linear magnetoelectricity. Their only drawback is weak ferromagnetism. In this paper, we employ density-functional-theory (DFT) calculations on hexagonal LuFeO$_3$ ($h$-LFO), targeting its magnetic ordering by electron doping,anticipating spin-disproportionation of the Fe sublattices. Indeed, we show that spin-disproportionation in heavily-electron-doped versions Lu$_{1-x}$Hf$_x$FeO$_3$ ($h$-LHFO), especially for x=1/3 and 1/2, leads to robust out-of-plane collinear ferrimagnetism that is stable at room temperature. Furthermore, the robust ferroelectricity of $h$-LFO persists via a Jahn-Teller metal-to-insulator transition. Finally, we construct a $h$-LHFO/$h$-2D heterostructure, where $h$-2D stands for the FE/FM monolayer MnSTe, and demonstrate strong magnetoelectric coupling, namely manipulation of magnetic skyrmions in MnSTe by an external electric field through the $h$-LHFO polarization, opening up a new realm for magnetoelectric applications

Should I stay or should I go? - 2:Monitoring influences on NHS staff retention in the post COVID-19 world Winter 2020 to spring 2023

The issue of staff retention in the NHS is not new but has been brought into sharp relief in the post-COVID-19 era of unpreceded staff shortage. While steps have been taken to train new health professionals and recruit from overseas, net gains to the NHS staff complement are at risk of being significantly blunted or defeated in the absence of finding ways to stabilise and enhance the retention of established staff. At the institutional level, what has been widely characterised as a pandemic of early­ exit risks a spiral of inter-related losses becoming endemic. Foreseeable impacts include loss of expertise and institutional memory, degraded capacity to deliver patient care, degraded workforce and work-team stability, loss of return on investment in health professional training, and increased human resource costs to employers (e.g. recruitment and employment of bank/agency staff). All have implications for standards of patient care and the potential to negatively impact on the well-being of staff in-post to the extent that it risks degrading their disposition and/or capacity to remain.The foundation research on which this report is based, 'Should I stay or should I go? NHS staff retention in the post COVID-19 world: Challenges and prospects', was funded by the Economic and Social Research Council, in response to the UKRI open­ call for COVID-19 relevant social research in spring 2020, supplemented by follow-on funding from the health sector. At its inception the research aims were to provide human resource strategy and policy relevant insight into:•the impact of the COVID-19 experiences and its legacy on employees' strength of attachment, commitment and capacity to remain in NHS employment;•the relative salience and strength of push and pull variables on staff stay versus leave intentions and behaviour;•what might need to change to motivate/enable current employees to remain in NHS employment; and•the need, nature and scope for intervention to maintain/enhance retention rates.At Wave Four of our survey, the scope of data gathering broadened from its initial focus on primary impacts arising from COVID-19 in 2020/21 and its legacy to include other features of the post-pandemic work environment, including: staff shortages, workload, job-demands, working conditions, pay and other background climate factors on staff resilience, capacity and disposition to remain.This report provides an overview of headline findings from the NHS employee survey component of our research. The survey sample covered all staff types, however a central focus was on health professionals and associated care staff, due to the more restricted scope for personnel substitution.The current report is focused on findings from Wave Four of the survey, conducted in spring 2023, and represents an update on and point of comparison with findings from the three earlier waves (winter 2020/21, summer/autumn 2021 and spring/summer 2022), published in the University of Bath Institute for Policy Research report series in January 2023 (Weyman et al. 2023).In common with Wave Three, the Wave Four survey was completed by a sample of NHS employees in England. Waves One and Two were completed by UK-wide samples. However, the close alignment of the response profiles across the devolved nation samples in Waves One and Two gives confidence to considering Waves Three and Four findings to have UK-wide generalisability and relevance

Genomic and proteomic profiling of responses to toxic metals in human lung cells.

Examining global effects of toxic metals on gene expression can be useful for elucidating patterns of biological response, discovering underlying mechanisms of toxicity, and identifying candidate metal-specific genetic markers of exposure and response. Using a 1,200 gene nylon array, we examined changes in gene expression following low-dose, acute exposures of cadmium, chromium, arsenic, nickel, or mitomycin C (MMC) in BEAS-2B human bronchial epithelial cells. Total RNA was isolated from cells exposed to 3 M Cd(II) (as cadmium chloride), 10 M Cr(VI) (as sodium dichromate), 3 g/cm2 Ni(II) (as nickel subsulfide), 5 M or 50 M As(III) (as sodium arsenite), or 1 M MMC for 4 hr. Expression changes were verified at the protein level for several genes. Only a small subset of genes was differentially expressed in response to each agent: Cd, Cr, Ni, As (5 M), As (50 M), and MMC each differentially altered the expression of 25, 44, 31, 110, 65, and 16 individual genes, respectively. Few genes were commonly expressed among the various treatments. Only one gene was altered in response to all four metals (hsp90), and no gene overlapped among all five treatments. We also compared low-dose (5 M, noncytotoxic) and high-dose (50 M, cytotoxic) arsenic treatments, which surprisingly, affected expression of almost completely nonoverlapping subsets of genes, suggesting a threshold switch from a survival-based biological response at low doses to a death response at high doses

Determining the sustainable irregular condition : an analysis of an irregular mixed-species selection stand in Scotland based on recurrent inventories at six-year intervals over 24 years

Funding This study was internally funded by the University of Aberdeen. Acknowledgements The authors wish to thank Mr Charles Taylor and Mr Mark Brazendale of Tay Forest District, Forestry Commission Scotland for their support in carrying out this study.Peer reviewedPostprin