Entropy and Barrier-Hopping Determine Conformational Viscoelasticity in Single Biomolecules

Biological macromolecules have complex and non-trivial energy landscapes, endowing them a unique conformational adaptability and diversity in function. Hence, understanding the processes of elasticity and dissipation at the nanoscale is important to molecular biology and also emerging fields such as nanotechnology. Here we analyse single molecule fluctuations in an atomic force microscope (AFM) experiment using a generic model of biopolymer viscoelasticity that importantly includes sources of local `internal' conformational dissipation. Comparing two biopolymers, dextran and cellulose, polysaccharides with and without the well-known `chair-to-boat' transition, reveals a signature of this simple conformational change as minima in both the elasticity and internal friction around a characteristic force. A calculation of two-state populations dynamics offers a simple explanation in terms of an elasticity driven by the entropy, and friction by barrier-controlled hopping, of populations on a landscape. The microscopic model, allows quantitative mapping of features of the energy landscape, revealing unexpectedly slow dynamics, suggestive of an underlying roughness to the free energy.Comment: 25 pages, 7 figures, naturemag.bst, modified nature.cls (naturemodified.cls

Effects of Soil Moisture on Photosynthesis and Fluorescence in Heteromeles arbutifolia

Some plants have evolved to increase their chances of survival by being drought-adapted. Among those plant species is Heteromeles arbutifolia, native to California. Logically, the fact that Heteromeles arbutifolia tolerates the low supply of water makes this plant more likely to be within environments where the level of sun exposure is high. Thus, we hypothesized that lowering soil moisture will cause an increase in xylem pressure, causing an increase in photo-protection and florescence, and a decrease in photosynthetic rate. This has not been tested before on a native chaparral plant such as Heteromeles arbutifolia. The experiment was held as following: six H. arbutifolia were evenly divided up into a treatment group and a control group. Both groups were given equal hydration at the beginning of this experiment. From then on the control group maintained a constant daily rate of hydration as the experimental groups received no water. Data of photosynthesis, photo-protection, florescence, xylem pressure, and soil moisture were taken for both groups throughout the entire experiment. Data collection showed statistically significant results of comparing non-photosynthetic quenching to photosynthetic quenching in water-stressed plants and water-saturated plants. Significant correlations were drawn between the following: the fraction of non-photosynthetic quenching to xylem pressure, the fraction of non-photosynthetic quenching to soil moisture, the fraction of non-photosynthetic quenching versus the time since the last event of irrigation, and xylem pressure and the time since the plant was last irrigated. It was concluded that photo-protection rate increases with water stress, which supported the initial hypothesis in part. However, the study also concluded that there was not a significant difference between the two groups regarding the florescence rate

The Role of Bovine Herpesvirus-1 Glycoprotein III in Molecular Pathogenesis and Immunomodulation.

Bovine herpesvirus-1 (BHV-1) is the pathogenic agent for infectious bovine rhinotracheitis. Current vaccines to protect against infection have serious disadvantages to producers and have been implicated in epizootic outbreaks. The virus has also been shown to cause suppression of the immune system. BHV-1 vaccines could be improved by identifying and genetically deleting those aspects of the virus responsible for immunosuppression and providing a marker within the genome which would distinguish the vaccine strain from wild types in the case of disease outbreaks. This study examined the role of gIII in viral infection and immunosuppression. This was accomplished by expressing full copy gIII and carboxy-terminal truncations of gIII in a transient eukaryotic expression system. The gene for gIII was then truncated to delete the portion of the protein showing homology to the MHC class II antigen constant domain. Glycoprotein III-derivatives were tested for their antigenic potential in lymphoproliferation assays using bovine peripheral blood mononuclear cells. Truncated versions of gIII lacking the MHC homologous region showed improved antigenic potential. Further studies using the full-copy version of gIII showed it suppressed the proliferative response to mitogen, UV-inactivated BHV-1, and other gIII derivatives. A gIII-null BHV-1 mutant, KB3305, was isolated to evaluate the effect of deleting the MHC homologous region on the virion and to provide a marker for strain identification. KB3305 replicated 10-fold less efficiently than the wild type and was approximately 15-fold more cell associated. Heparin reduced the attachment of the mutant to GBK cells by 65%, compared to 75% for the wild type. In addition, KB3305 failed to exhibit hemagglutinating activity, in contrast to the wild-type strain. In lymphoproliferation assays, KB3305 was able to stimulate an equivalent response to that induced by the wild type virus, indicating that the lack of gIII does not influence viral antigenicity

Identity, Memory and Popular Politics in Sixteenth-Century Kent

This thesis is an investigation of social memory, the landscape, and identity in Tudor Kent, with the aim of obtaining a better understanding of popular politics in early modern England. By the end of Elizabeth's reign, England was in a very different position to the England of Henry VII. In 1603 England was a Protestant nation, the state's reach into the lives of its subjects was further than ever before, and popular rebellion on the scale of 1381 and 1549 had died out. Each of these changes would have had a significant impact on the commonalty. In order to understand the political lives of the common people of England, it is therefore important to explore their experiences. The way people remembered their lives, and the way that these memories were embedded in collective histories and written into the local landscape itself, were vital to the formation of identities, which in turn influenced their political associations and actions. The use of sociological and anthropological theories make it possible to look deeper into the way identity influenced political action, especially in an area where sources are scarce. This study of Kent, a county with a strong identity and a long history of popular rebellion, examines the overlapping identities available to its sixteenth century inhabitants. It looks at the way in which local layers of identity, based in customs, access to resources, and a relationship with the landscape, combined with external influences such as the Reformation or the Armada to form a variety of identities of different strengths in different contexts. The way in which these identities interacted with each other as well as with the political circumstances is shown to influence collective participation in protest action. Kent's rebellious traditions, with the associated rhetoric and physical spaces, form another layer of identity to be called upon at certain times. The strength of the superordinate and subordinate identities of the Kentish people in such contexts is explored in order to understand the engagement in rebellion, as well as the decline of such engagement in the second half of the sixteenth century. By combining this with the labels applied to the people of Kent in chronicles, plays and other texts, which served to undermine identities such as Invicta, this thesis looks at the political ramifications of identity and memory in Kent

Investigating the functionality of an OCT4-short response element in human induced pluripotent stem cells.

Pluripotent stem cells offer great therapeutic promise for personalized treatment platforms for numerous injuries, disorders, and diseases. Octamer-binding transcription factor 4 (OCT4) is a key regulatory gene maintaining pluripotency and self-renewal of mammalian cells. With site-specific integration for gene correction in cellular therapeutics, use of the OCT4 promoter may have advantages when expressing a suicide gene if pluripotency remains. However, the human OCT4 promoter region is 4 kb in size, limiting the capacity of therapeutic genes and other regulatory components for viral vectors, and decreasing the efficiency of homologous recombination. The purpose of this investigation was to characterize the functionality of a novel 967bp OCT4-short response element during pluripotency and to examine the OCT4 titer-dependent response during differentiation to human derivatives not expressing OCT4. Our findings demonstrate that the OCT4-short response element is active in pluripotency and this activity is in high correlation with transgene expression in vitro, and the OCT4-short response element is inactivated when pluripotent cells differentiate. These studies demonstrate that this shortened OCT4 regulatory element is functional and may be useful as part of an optimized safety component in a site-specific gene transferring system that could be used as an efficient and clinically applicable safety platform for gene transfer in cellular therapeutics

Communication interventions for medically unexplained symptom conditions in general practice : a systematic review and meta-analysis of randomised controlled trials

Background Medically unexplained symptoms (MUS) account for 3–50% of all General Practitioner (GP) consultations and are difficult to diagnose due to their unknown aetiology, symptom overlap between conditions, and lack of effective treatment options. MUS patients’ and primary care clinicians frequently face challenges during consultations, with GPs reporting difficulty identifying and classifying MUS, whilst patients report stigma and feeling illegitimised by clinicians. Communication interventions have been proposed as a method to facilitate the doctor-patient relationship and aid the management of MUS. Aim This systematic review aims to evaluate the effectiveness of primary care based communication interventions at improving MUS patients’ and/or clinician outcomes. Method Four electronic databases were searched from inception to November 2021. Two researchers independently undertook screening, data extraction and quality appraisal. Given the heterogeneous nature of the studies identified, narrative syntheses were conducted, along with meta-analyses where possible to pool data. Results 9 papers from 10 Randomised Controlled Trials were included. The included studies displayed considerable risk of bias and poor reporting. Some limited evidence suggests that communication interventions tailored to MUS and not following a pre-specified model (such as reattribution) could improve pain, mental and physical functioning whilst reattribution training may improve clinician confidence treating MUS. However, methodological limitations mean that these findings should be interpreted with caution. Conclusion A range of interventions for improving communication with MUS patients in primary care have been evaluated. However, the heterogeneous nature of existing evidence and poor study quality mean we cannot conclude whether these interventions are effective. Before considering further randomised controlled trials researchers should focus on developing a new or modified communication intervention for MUS patients and their clinicians. Trail registration The systematic review was prospectively registered with PROSPERO (registration record CRD42020206437)

Randomised controlled feasibility trial of a web-based weight management intervention with nurse support for obese patients in primary care

<b>Background</b><p></p> There is a need for cost-effective weight management interventions that primary care can deliver to reduce the morbidity caused by obesity. Automated web-based interventions might provide a solution, but evidence suggests that they may be ineffective without additional human support. The main aim of this study was to carry out a feasibility trial of a web-based weight management intervention in primary care, comparing different levels of nurse support, to determine the optimal combination of web-based and personal support to be tested in a full trial.<p></p> <b>Methods</b><p></p> This was an individually randomised four arm parallel non-blinded trial, recruiting obese patients in primary care. Following online registration, patients were randomly allocated by the automated intervention to either usual care, the web-based intervention only, or the web-based intervention with either basic nurse support (3 sessions in 3 months) or regular nurse support (7 sessions in 6 months). The main outcome measure (intended as the primary outcome for the main trial) was weight loss in kg at 12 months. As this was a feasibility trial no statistical analyses were carried out, but we present means, confidence intervals and effect sizes for weight loss in each group, uptake and retention, and completion of intervention components and outcome measures.<p></p> <b>Results</b><p></p> All randomised patients were included in the weight loss analyses (using Last Observation Carried Forward). At 12 months mean weight loss was: usual care group (n = 43) 2.44 kg; web-based only group (n = 45) 2.30 kg; basic nurse support group (n = 44) 4.31 kg; regular nurse support group (n = 47) 2.50 kg. Intervention effect sizes compared with usual care were: d = 0.01 web-based; d = 0.34 basic nurse support; d = 0.02 regular nurse support. Two practices deviated from protocol by providing considerable weight management support to their usual care patients.<p></p> <b>Conclusions</b><p></p> This study demonstrated the feasibility of delivering a web-based weight management intervention supported by practice nurses in primary care, and suggests that the combination of the web-based intervention with basic nurse support could provide an effective solution to weight management support in a primary care context

Restoring Ureagenesis in Hepatocytes by CRISPR/Cas9-mediated Genomic Addition to Arginase-deficient Induced Pluripotent Stem Cells.

Urea cycle disorders are incurable enzymopathies that affect nitrogen metabolism and typically lead to hyperammonemia. Arginase deficiency results from a mutation in Arg1, the enzyme regulating the final step of ureagenesis and typically results in developmental disabilities, seizures, spastic diplegia, and sometimes death. Current medical treatments for urea cycle disorders are only marginally effective, and for proximal disorders, liver transplantation is effective but limited by graft availability. Advances in human induced pluripotent stem cell research has allowed for the genetic modification of stem cells for potential cellular replacement therapies. In this study, we demonstrate a universally-applicable CRISPR/Cas9-based strategy utilizing exon 1 of the hypoxanthine-guanine phosphoribosyltransferase locus to genetically modify and restore arginase activity, and thus ureagenesis, in genetically distinct patient-specific human induced pluripotent stem cells and hepatocyte-like derivatives. Successful strategies restoring gene function in patient-specific human induced pluripotent stem cells may advance applications of genetically modified cell therapy to treat urea cycle and other inborn errors of metabolism

Genotype imputation in winter wheat using first-generation haplotype map SNPs improves genome-wide association mapping and genomic prediction of traits

Genome-wide single nucleotide polymorphism (SNP) variation allows for the capture of haplotype structure in populations and prediction of unobserved genotypes based on inferred regions of identity-by-descent (IBD). Here we have used a first-generation wheat haplotype map created by targeted re-sequencing of low-copy genomic regions in the reference panel of 62 lines to impute marker genotypes in a diverse panel of winter wheat cultivars from the U.S. Great Plains. The IBD segments between the reference population and winter wheat cultivars were identified based on SNP genotyped using the 90K iSelect wheat array and genotyping by sequencing (GBS). A genome-wide association study and genomic prediction of resistance to stripe rust in winter wheat cultivars showed that an increase in marker density achieved by imputation improved both the power and precision of trait mapping and prediction. The majority of the most significant marker-trait associations belonged to imputed genotypes. With the vast amount of SNP variation data accumulated for wheat in recent years, the presented imputation framework will greatly improve prediction accuracy in breeding populations and increase resolution of trait mapping hence, facilitate cross-referencing of genotype datasets available across different wheat populations

An internet-based intervention with brief nurse support to manage obesity in primary care (POWeR+): a pragmatic, parallel-group, randomised controlled trial

Background The obesity epidemic has major public health consequences. Expert dietetic and behavioural counselling with intensive follow-up is effective, but resource requirements severely restrict widespread implementation in primary care, where most patients are managed. We aimed to estimate the effectiveness and cost-effectiveness of an internet-based behavioural intervention (POWeR+) combined with brief practice nurse support in primary care. Methods We did this pragmatic, parallel-group, randomised controlled trial at 56 primary care practices in central and south England. Eligible adults aged 18 years or older with a BMI of 30 kg/m2 or more (or ≥28 kg/m2 with hypertension, hypercholesterolaemia, or diabetes) registered online with POWeR+—a 24 session, web-based, weight management intervention lasting 6 months. After registration, the website automatically randomly assigned patients (1:1:1), via computer-generated random numbers, to receive evidence-based dietetic advice to swap foods for similar, but healthier, choices and increase fruit and vegetable intake, in addition to 6 monthly nurse follow-up (control group); web-based intervention and face-to-face nurse support (POWeR+Face-to-face [POWeR+F]; up to seven nurse contacts over 6 months); or web-based intervention and remote nurse support (POWeR+Remote [POWeR+R]; up to five emails or brief phone calls over 6 months). Participants and investigators were masked to group allocation at the point of randomisation; masking of participants was not possible after randomisation. The primary outcome was weight loss averaged over 12 months. We did a secondary analysis of weight to measure maintenance of 5% weight loss at months 6 and 12. We modelled the cost-effectiveness of each intervention. We did analysis by intention to treat, with multiple imputation for missing data. This trial is registered as an International Standard Randomised Controlled Trial, number ISRCTN21244703. Findings Between Jan 30, 2013, and March 20, 2014, 818 participants were randomly assigned to the control group (n=279), the POWeR+F group (n=269), or the POWeR+R group (n=270). Weight loss averaged over 12 months was recorded in 666 (81%) participants. The control group lost almost 3 kg over 12 months (crude mean weight: baseline 104·38 kg [SD 21·11; n=279], 6 months 101·91 kg [19·35; n=136], 12 months 101·74 kg [19·57; n=227]). The primary imputed analysis showed that compared with the control group, patients in the POWeR+F group achieved an additional weight reduction of 1·5 kg (95% CI 0·6–2·4; p=0·001) averaged over 12 months, and patients in the POWeR+R group achieved an additional 1·3 kg (0·34–2·2; p=0·007). 21% of patients in the control group had maintained a clinically important 5% weight reduction at month 12, compared with 29% of patients in the POWeR+F group (risk ratio 1·56, 0·96–2·51; p=0·070) and 32% of patients in the POWeR+R group (1·82, 1·31–2·74; p=0·004). The incremental overall cost to the health service per kg weight lost with the POWeR+ interventions versus the control strategy was £18 (95% CI −129 to 195) for POWeR+F and –£25 (−268 to 157) for POWeR+R; the probability of being cost-effective at a threshold of £100 per kg lost was 88% and 98%, respectively. No adverse events were reported. Interpretation Weight loss can be maintained in some individuals by use of novel written material with occasional brief nurse follow-up. However, more people can maintain clinically important weight reductions with a web-based behavioural program and brief remote follow-up, with no increase in health service costs. Future research should assess the extent to which clinically important weight loss can be maintained beyond 1 year