Biochemical Characterization of the Release of FAK Autoinhibition

Focal adhesion kinase is an essential nonreceptor tyrosine kinase that plays an important role in development, in homeostasis and in the progression of human disease. Multiple stimuli activate FAK, which requires a change in structure from an autoinhibited to activated conformation. In the autoinhibited conformation the FERM domain associates with the catalytic domain of FAK and PI(4,5)P2 binding to the FERM domain plays a role in the release of autoinhibition, activating the enzyme. This work aims to better characterize the residues involved in the release of the autoinhibitory conformation. The first aim was to characterize the effects of point mutations found in the COSMIC database. None of these mutations had an impact on the FAK autoinhibited conformation, thus classifying FAK as a kinase in which overexpression, and not mutation, is the mechanism by which its activity is increased in cancer. The second aim was to assess the role of FERM domain basic residues at the interface of the FERM and kinase domains in the autoinhibited conformation, R184, K190, and K191. While mutation of these residues does not cause a significant shift in FAK conformation, the results do suggest a minor role in binding to the membrane through PI(4,5)P2 and interacting with the kinase domain to maintain the autoinhibited conformation. The third aim was to characterize residues on the catalytic domain that were modeled binding with the membrane in silico. Constructs with these residues mutated to alanine exhibited defects in phosphorylation and failed to completely rescue the phenotype associated with fak -/- phenotype fibroblasts demonstrating the importance of these residues in FAK function. The catalytic domain of FAK exhibited PI(4,5)P2 binding in vitro and binding activity was lost upon mutation of putative PI(4,5)P2 binding site basic residues. Collectively, these studies further characterize the structural elements that aid in maintenance of both the open and autoinhibited conformations of FAK

A co-production approach guided by the behaviour change wheel to develop an intervention for reducing sedentary behaviour after stroke

Background Stroke survivors are highly sedentary; thus, breaking up long uninterrupted bouts of sedentary behaviour could have substantial health benefit. However, there are no intervention strategies specifically aimed at reducing sedentary behaviour tailored for stroke survivors. The purpose of this study was to use co-production approaches to develop an intervention to reduce sedentary behaviour after stroke. Methods A series of five co-production workshops with stroke survivors, their caregivers, stroke service staff, exercise professionals, and researchers were conducted in parallel in two-stroke services (England and Scotland). Workshop format was informed by the behaviour change wheel (BCW) framework for developing interventions and incorporated systematic review and empirical evidence. Taking an iterative approach, data from activities and audio recordings were analysed following each workshop and findings used to inform subsequent workshops, to inform both the activities of the next workshop and ongoing intervention development. Findings Co-production workshop participants (n = 43) included 17 staff, 14 stroke survivors, six caregivers and six researchers. The target behaviour for stroke survivors is to increase standing and moving, and the target behaviour for caregivers and staff is to support and encourage stroke survivors to increase standing and moving. The developed intervention is primarily based on co-produced solutions to barriers to achieving the target behaviour. The developed intervention includes 34 behaviour change techniques. The intervention is to be delivered through stroke services, commencing in the inpatient setting and following through discharge into the community. Participants reported that taking part in intervention development was a positive experience. Conclusions To our knowledge, this is the first study that has combined the use of co-production and the BCW to develop an intervention for use in stroke care. In-depth reporting of how a co-production approach was combined with the BCW framework, including the design of bespoke materials for workshop activities, should prove useful to other researchers and practitioners involved in intervention development in stroke

Cells exhibiting strong p16INK4a promoter activation in vivo display features of senescence

The activation of cellular senescence throughout the lifespan promotes tumor suppression, whereas the persistence of senescent cells contributes to aspects of aging. This theory has been limited, however, by an inability to identify and isolate individual senescent cells within an intact organism. Toward that end, we generated a murine reporter strain by “knocking-in” a fluorochrome, tandem-dimer Tomato (tdTom), into exon 1α of the p16 INK4a locus. We used this allele (p16 tdTom ) for the enumeration, isolation, and characterization of individual p16 INK4a -expressing cells (tdTom + ). The half-life of the knocked-in transcript was shorter than that of the endogenous p16 INK4a mRNA, and therefore reporter expression better correlated with p16 INK4a promoter activation than p16 INK4a transcript abundance. The frequency of tdTom + cells increased with serial passage in cultured murine embryo fibroblasts from p16 tdTom/+ mice. In adult mice, tdTom + cells could be readily detected at low frequency in many tissues, and the frequency of these cells increased with aging. Using an in vivo model of peritoneal inflammation, we compared the phenotype of cells with or without activation of p16 INK4a and found that tdTom + macrophages exhibited some features of senescence, including reduced proliferation, senescence-associated β-galactosidase (SA-β-gal) activation, and increased mRNA expression of a subset of transcripts encoding factors involved in SA-secretory phenotype (SASP). These results indicate that cells harboring activation of the p16 INK4a promoter accumulate with aging and inflammation in vivo, and display characteristics of senescence

Disruption of Thyroid Hormone Activation in Type 2 Deiodinase Knockout Mice Causes Obesity With Glucose Intolerance and Liver Steatosis Only at Thermoneutrality

Objective: Thyroid hormone accelerates energy expenditure; thus, hypothyroidism is intuitively associated with obesity. However, studies failed to establish such a connection. In brown adipose tissue (BAT), thyroid hormone activation via type 2 deiodinase (D2) is necessary for adaptive thermogenesis, such that mice lacking D2 (D2KO) exhibit an impaired thermogenic response to cold. Here we investigate whether the impaired thermogenesis of D2KO mice increases their susceptibility to obesity when placed on a high-fat diet. Research Design and Methods: To test this, D2KO mice were admitted to a comprehensive monitoring system acclimatized to room temperature (22$^\circ$C) or thermoneutrality (30$^\circ$C) and kept either on chow or high-fat diet for 60 days. Results: At 22$^\circ$C, D2KO mice preferentially oxidize fat, have a similar sensitivity to diet-induced obesity, and are supertolerant to glucose. However, when thermal stress is eliminated at thermoneutrality (30$^\circ$C), an opposite phenotype is encountered, one that includes obesity, glucose intolerance, and exacerbated hepatic steatosis. We suggest that a compensatory increase in BAT sympathetic activation of the D2KO mice masks metabolic repercussions that they would otherwise exhibit. Conclusions: Thus, upon minimization of thermal stress, high-fat feeding reveals the defective capacity of D2KO mice for diet-induced thermogenesis, provoking a paradigm shift in the understanding of the role of the thyroid hormone in metabolism

Implicit and Explicit Values as a Predictor of Ethical Decision-Making and Ethical Behavior

The present study uses measures of implicit and explicit values to predict moral behaviors. Implicit value measures based on a word-fragment completion tasks were developed in this study to assess implicit values. Because values and moral processes are believed to operate at both explicit and implicit levels, it was hypothesized that both implicit and explicit values would predict moral behaviors. Results from a laboratory study show that both implicit and explicit values predicted actual moral behavior, consistent with dual process theories of morality. Chronic collective identity moderated the relation of both implicit and explicit values to ethical behavior. Theoretical and practical implications for the use of both explicit and implicit value measures in research and applied settings are discussed

Reimagining Restitution: New Approaches to Support Youth and Communities

Courts began ordering youth restitution in the 1960s as a less restrictive sanction than probation or incarceration for mostly white youth. Since then, restitution has been linked to higher recidivism rates and heightened racial and economic disparities in the juvenile justice system. This report provides an overview of the historical and current landscape of restitution imposed on youth, the impact of restitution on youth, victims and communities, and provides recommendations for how jurisdictions can reimagine restitution

The intrinsic primary bioreceptivity of concrete in the coastal environment – A review

The proliferation of artificial concrete structures (ACSs) in the marine environment causes intertidal habitat loss and is a poor surrogate for natural rocky shores in terms of species richness, abundance, and community composition. As hard engineered coastlines increase, there is growing interest in how new concrete structures can facilitate improved habitat and biodiversity compared to existing concrete structures. Experiments that have substituted cement binder and aggregates in varying proportions and combinations have demonstrated that it is possible to enhance the primary bioreceptivity of concrete, either chemically or via microtopographical texture. This review synthesises key literature and identifies which concrete formulas prove most effective at enhancing bioreceptivity and those that have limited value, providing recommendations for coastal practitioners and for formulas that warrant further study. It is evident that the efficacy of chemical bioreceptivity of concrete is likely to be spatio-temporally limited (months) and enhancing surface roughness should be prioritised as a way to enhance colonisation. However, both chemical and physical methods require further investigation in within in situ marine settings for longer durations (>12 months)

Temperature Drives Epidemics in a Zooplankton-Fungus Disease System: A Trait-Driven Approach Points to Transmission via Host Foraging

Climatic warming will likely have idiosyncratic impacts on infectious diseases, causing some to increase while others decrease or shift geographically. A mechanistic framework could better predict these different temperature-disease outcomes. However, such a framework remains challenging to develop, due to the nonlinear and (sometimes) opposing thermal responses of different host and parasite traits and due to the difficulty of validating model predictions with observations and experiments. We address these challenges in a zooplanktonfungus (Daphnia dentifera–Metschnikowia bicuspidata) system. We test the hypothesis that warmer temperatures promote disease spread and produce larger epidemics. In lakes, epidemics that start earlier and warmer in autumn grow much larger. In a mesocosm experiment, warmer temperatures produced larger epidemics. A mechanistic model parameterized with trait assays revealed that this pattern arose primarily from the temperature dependence of transmission rate (b), governed by the increasing foraging (and, hence, parasite exposure) rate of hosts ( f ). In the trait assays, parasite production seemed sufficiently responsive to shape epidemics as well; however, this trait proved too thermally insensitive in the mesocosm experiment and lake survey to matter much. Thus, in warmer environments, increased foraging of hosts raised transmission rate, yielding bigger epidemics through a potentially general, exposure-based mechanism for ectotherms. This mechanistic approach highlights how a trait-based framework will enhance predictive insight into responses of infectious disease to a warmer world

Inconsistent bioreceptivity of three mortar mixes in subtidal sites.

Concrete is extensively used in coastal engineering and development which, in addition to its high carbon footprint, threatens intertidal habitats and ecosystems. Eco-engineering addresses this by designing habitat features into coastal infrastructure. The chemical bioreceptivity of cement has been shown to vary, but ordinary Portland cement is generally considered to be the least bioreceptive. In this study, we compare two low carbon mortars (a natural, single source cement (VP), and an ordinary Portland cement/ ground granulated blast furnace slag blend (GGBS)) with an ordinary Portland cement-based control mix (OPC). The three mortars were made into smooth blocks which were secured to crates and deployed subtidally in two estuary sites on the south UK coast for 1 year. At 3-, 6- and 12-months intervals a crate was recovered from each site and species abundance, biomass and assemblage composition were determined. After 12 months, the VP mortar was significantly more species rich than both the OPC control and GGBS mortar, and organisms were significantly more abundant (numeric counts only), though this varied by mortar and site. However, OPC controls showed significantly higher percentage cover of biota than both low carbon mixes in both harbours. Overall, the GGBS mortar showed the least bioreceptivity of all three mortars. It is evident that the primary chemical bioreceptivity of OPC, GGBS and VP is inconsistent between ecological metrics and study sites and that using lower carbon cements does not necessarily enhance colonisation. The primary chemical bioreceptivity of these mortars may therefore perform inconsistently and other intrinsic factors that impact bioreceptivity and primary succession, such as rugosity, should be prioritised when designing ecological enhancements. Sustainability of materials, such as opting for low carbon cements, should also be a priority

Habitat, predators, and hosts regulate disease in Daphnia through direct and indirect pathways

Community ecology can link habitat to disease via interactions among habitat, focal hosts, other hosts, their parasites, and predators. However, complicated food web interactions (i.e., trophic interactions among predators and their impacts on host density and diversity) often obscure the important pathways regulating disease. Here, we disentangle community drivers in a case study of planktonic disease, using a two‐step approach. In step one, we tested univariate field patterns linking community interactions directly to two disease metrics. Density of focal hosts (Daphnia dentifera) was related to density but not prevalence of fungal (Metschnikowia bicuspidata) infections. Both disease metrics appeared to be driven by selective predators that cull infected hosts (fish, e.g., Lepomis macrochirus), sloppy predators that spread parasites while feeding (midges, Chaoborus punctipennis), and spore predators that reduce contact between focal hosts and parasites (other zooplankton, especially small‐bodied Ceriodaphnia sp.). Host diversity also negatively correlated with disease, suggesting a dilution effect. However, several of these univariate patterns were initially misleading, due to confounding ecological links among habitat, predators, host density, and host diversity. In step two, path models uncovered and explained these misleading patterns, and grounded them in habitat structure (refuge size). First, rather than directly reducing infection prevalence, fish predation drove disease indirectly through changes in density of midges and frequency of small spore predators (which became more frequent in lakes with small refuges). Second, small spore predators drove the two disease metrics through fundamentally different pathways: they directly reduced infection prevalence, but indirectly reduced density of infected hosts by lowering density of focal hosts (likely via competition). Third, the univariate diversity–disease pattern (signaling a dilution effect) merely reflected the confounding direct effects of these small spore predators. Diversity per se had no effect on disease, after accounting for the links between small spore predators, diversity, and infection prevalence. In turn, these small spore predators were regulated by both size‐selective fish predation and refuge size. Thus, path models not only explain each of these surprising results, but also trace their origins back to habitat structure.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/134436/1/ecm1222_am.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/134436/2/ecm1222-sup-0001-AppendixS1.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/134436/3/ecm1222.pd