Publisher Correction: Blocking elevated p38 MAPK restores efferocytosis and inflammatory resolution in the elderly.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Blocking elevated p38 MAPK restores efferocytosis and inflammatory resolution in the elderly.

Increasing age alters innate immune-mediated responses; however, the mechanisms underpinning these changes in humans are not fully understood. Using a human dermal model of acute inflammation, we found that, although inflammatory onset is similar between young and elderly individuals, the resolution phase was substantially impaired in elderly individuals. This arose from a reduction in T cell immunoglobulin mucin receptor-4 (TIM-4), a phosphatidylserine receptor expressed on macrophages that enables the engulfment of apoptotic bodies, so-called efferocytosis. Reduced TIM-4 in elderly individuals was caused by an elevation in macrophage p38 mitogen-activated protein kinase (MAPK) activity. Administering an orally active p38 inhibitor to elderly individuals rescued TIM-4 expression, cleared apoptotic bodies and restored a macrophage resolution phenotype. Thus, inhibiting p38 in elderly individuals rejuvenated their resolution response to be more similar to that of younger people. This is the first resolution defect identified in humans that has been successfully reversed, thereby highlighting the tractability of targeting pro-resolution biology to treat diseases driven by chronic inflammation.

Identification of Phosphorylation Codes for Arrestin Recruitment by G Protein-Coupled Receptors.

G protein-coupled receptors (GPCRs) mediate diverse signaling in part through interaction with arrestins, whose binding promotes receptor internalization and signaling through G protein-independent pathways. High-affinity arrestin binding requires receptor phosphorylation, often at the receptor's C-terminal tail. Here, we report an X-ray free electron laser (XFEL) crystal structure of the rhodopsin-arrestin complex, in which the phosphorylated C terminus of rhodopsin forms an extended intermolecular ß sheet with the N-terminal ß strands of arrestin. Phosphorylation was detected at rhodopsin C-terminal tail residues T336 and S338. These two phospho-residues, together with E341, form an extensive network of electrostatic interactions with three positively charged pockets in arrestin in a mode that resembles binding of the phosphorylated vasopressin-2 receptor tail to ß-arrestin-1. Based on these observations, we derived and validated a set of phosphorylation codes that serve as a common mechanism for phosphorylation-dependent recruitment of arrestins by GPCRs.

Structural Basis of the Activation of Heterotrimeric Gs-Protein by Isoproterenol-Bound ß1-Adrenergic Receptor.

Cardiac disease remains the leading cause of morbidity and mortality worldwide. The ß1-adrenergic receptor (ß1-AR) is a major regulator of cardiac functions and is downregulated in the majority of heart failure cases. A key physiological process is the activation of heterotrimeric G-protein Gs by ß1-ARs, leading to increased heart rate and contractility. Here, we use cryo-electron microscopy and functional studies to investigate the molecular mechanism by which ß1-AR activates Gs. We find that the tilting of α5-helix breaks a hydrogen bond between the sidechain of His373 in the C-terminal α5-helix and the backbone carbonyl of Arg38 in the N-terminal αN-helix of Gαs. Together with the disruption of another interacting network involving Gln59 in the α1-helix, Ala352 in the ß6-α5 loop, and Thr355 in the α5-helix, these conformational changes might lead to the deformation of the GDP-binding pocket. Our data provide molecular insights into the activation of G-proteins by G-protein-coupled receptors.

Genetic prevalence and clinical relevance of canine Mendelian disease variants in over one million dogs.

Hundreds of genetic variants implicated in Mendelian disease have been characterized in dogs and commercial screening is being offered for most of them worldwide. There is typically limited information available regarding the broader population frequency of variants and uncertainty regarding their functional and clinical impact in ancestry backgrounds beyond the discovery breed. Genetic panel screening of disease-associated variants, commercially offered directly to the consumer or via a veterinary clinician, provides an opportunity to establish large-scale cohorts with phenotype data available to address open questions related to variant prevalence and relevance. We screened the largest canine cohort examined in a single study to date (1,054,293 representative dogs from our existing cohort of 3.5 million; a total of 811,628 mixed breed dogs and 242,665 purebreds from more than 150 countries) to examine the prevalence and distribution of a total of 250 genetic disease-associated variants in the general population. Electronic medical records from veterinary clinics were available for 43.5% of the genotyped dogs, enabling the clinical impact of variants to be investigated. We provide detailed frequencies for all tested variants across breeds and find that 57% of dogs carry at least one copy of a studied Mendelian disease-associated variant. Focusing on a subset of variants, we provide evidence of full penetrance for 10 variants, and plausible evidence for clinical significance of 22 variants, on diverse breed backgrounds. Specifically, we report that inherited hypocatalasia is a notable oral health condition, confirm that factor VII deficiency presents as subclinical bleeding propensity and verify two genetic causes of reduced leg length. We further assess genome-wide heterozygosity levels in over 100 breeds, and show that a reduction in genome-wide heterozygosity is associated with an increased Mendelian disease variant load. The accumulated knowledge represents a resource to guide discussions on genetic test relevance by breed.

A cholesterol switch controls phospholipid scrambling by G protein-coupled receptors.

Class A G protein-coupled receptors (GPCRs), a superfamily of cell membrane signaling receptors, moonlight as constitutively active phospholipid scramblases. The plasma membrane of metazoan cells is replete with GPCRs yet has a strong resting trans-bilayer phospholipid asymmetry, with the signaling lipid phosphatidylserine confined to the cytoplasmic leaflet. To account for the persistence of this lipid asymmetry in the presence of GPCR scramblases, we hypothesized that GPCR-mediated lipid scrambling is regulated by cholesterol, a major constituent of the plasma membrane. We now present a technique whereby synthetic vesicles reconstituted with GPCRs can be supplemented with cholesterol to a level similar to that of the plasma membrane and show that the scramblase activity of two prototypical GPCRs, opsin and the ß1-adrenergic receptor, is impaired upon cholesterol loading. Our data suggest that cholesterol acts as a switch, inhibiting scrambling above a receptor-specific threshold concentration to disable GPCR scramblases at the plasma membrane.

Broadly neutralizing humanized SARS-CoV-2 antibody binds to a conserved epitope on Spike and provides antiviral protection through inhalation-based delivery in non-human primates.

The COVID-19 pandemic represents a global challenge that has impacted and is expected to continue to impact the lives and health of people across the world for the foreseeable future. The rollout of vaccines has provided highly anticipated relief, but effective therapeutics are required to further reduce the risk and severity of infections. Monoclonal antibodies have been shown to be effective as therapeutics for SARS-CoV-2, but as new variants of concern (VoC) continue to emerge, their utility and use have waned due to limited or no efficacy against these variants. Furthermore, cumbersome systemic administration limits easy and broad access to such drugs. As well, concentrations of systemically administered antibodies in the mucosal epithelium, a primary site of initial infection, are dependent on neonatal Fc receptor mediated transport and require high drug concentrations. To reduce the viral load more effectively in the lung, we developed an inhalable formulation of a SARS-CoV-2 neutralizing antibody binding to a conserved epitope on the Spike protein, ensuring pan-neutralizing properties. Administration of this antibody via a vibrating mesh nebulization device retained antibody integrity and resulted in effective distribution of the antibody in the upper and lower respiratory tract of non-human primates (NHP). In comparison with intravenous administration, significantly higher antibody concentrations can be obtained in the lung, resulting in highly effective reduction in viral load post SARS-CoV-2 challenge. This approach may reduce the barriers of access and uptake of antibody therapeutics in real-world clinical settings and provide a more effective blueprint for targeting existing and potentially emerging respiratory tract viruses.

Social dilemmas among unequals.

Direct reciprocity is a powerful mechanism for the evolution of cooperation on the basis of repeated interactions1-4. It requires that interacting individuals are sufficiently equal, such that everyone faces similar consequences when they cooperate or defect. Yet inequality is ubiquitous among humans5,6 and is generally considered to undermine cooperation and welfare7-10. Most previous models of reciprocity do not include inequality11-15. These models assume that individuals are the same in all relevant aspects. Here we introduce a general framework to study direct reciprocity among unequal individuals. Our model allows for multiple sources of inequality. Subjects can differ in their endowments, their productivities and in how much they benefit from public goods. We find that extreme inequality prevents cooperation. But if subjects differ in productivity, some endowment inequality can be necessary for cooperation to prevail. Our mathematical predictions are supported by a behavioural experiment in which we vary the endowments and productivities of the subjects. We observe that overall welfare is maximized when the two sources of heterogeneity are aligned, such that more productive individuals receive higher endowments. By contrast, when endowments and productivities are misaligned, cooperation quickly breaks down. Our findings have implications for policy-makers concerned with equity, efficiency and the provisioning of public goods.

Genetic epidemiology of blood type, disease and trait variants, and genome-wide genetic diversity in over 11,000 domestic cats.

In the largest DNA-based study of domestic cats to date, 11,036 individuals (10,419 pedigreed cats and 617 non-pedigreed cats) were genotyped via commercial panel testing elucidating the distribution and frequency of known disease, blood type, and physical trait associated genetic variants across cat breeds. This study provides allele frequencies for many disease-associated variants for the first time and provides updates on previously reported information with evidence suggesting that DNA testing has been effectively used to reduce disease associated variants within certain pedigreed cat populations over time. We identified 13 disease-associated variants in 47 breeds or breed types in which the variant had not previously been documented, highlighting the relevance of comprehensive genetic screening across breeds. Three disease-associated variants were discovered in non-pedigreed cats only. To investigate the causality of nine disease-associated variants in cats of different breed backgrounds our veterinarians conducted owner interviews, reviewed clinical records, and invited cats to have follow-up clinical examinations. Additionally, genetic variants determining blood types A, B and AB, which are relevant clinically and in cat breeding, were genotyped. Appearance-associated genetic variation in all cats is also discussed. Lastly, genome-wide SNP heterozygosity levels were calculated to obtain a comparable measure of the genetic diversity in different cat breeds. This study represents the first comprehensive exploration of informative Mendelian variants in felines by screening over 10,000 pedigreed cats. The results qualitatively contribute to the understanding of feline variant heritage and genetic diversity and demonstrate the clinical utility and importance of such information in supporting breeding programs and the research community. The work also highlights the crucial commitment of pedigreed cat breeders and registries in supporting the establishment of large genomic databases, that when combined with phenotype information can advance scientific understanding and provide insights that can be applied to improve the health and welfare of cats.

Interrogating l-fuconate dehydratase with tartronate and 3-hydroxypyruvate reveals subtle differences within the mandelate racemase-subgroup of the enolase superfamily.

Enzymes of the enolase superfamily share a conserved structure and a common partial reaction (i.e., metal-assisted, Brønsted base-catalyzed enol(ate) formation). The architectures of the enolization apparatus at the active sites of the mandelate racemase (MR)-subgroup members MR and l-fuconate dehydratase (FucD) are almost indistinguishable at the structural level. Tartronate and 3-hydroxypyruvate (3-HP) recognize the enolization apparatus and can be used to interrogate the active sites for differences that may not be apparent from structural data. We report a circular dichroism-based assay of FucD activity that monitors the change in ellipticity at 216 nm (Δ[Θ]S-P = 8985 ± 87 deg cm2 mol-1) accompanying the conversion of l-fuconate to 2-keto-3-deoxy-l-fuconate. Tartronate was a linear mixed-type inhibitor of FucD (Ki = 8.4 ± 0.7 mM, αKi = 63 ± 11 mM), binding 18-fold weaker than l-fuconate, compared with 2-fold weaker binding of tartronate by MR relative to mandelate. 3-HP irreversibly inactivated FucD (kinact/KI = 0.018 ± 0.002 M-1s-1) with an efficiency that was ∼4.6 × 103-fold less than that observed with MR. The inactivation arose predominantly from modifications at multiple sites and Tris-HCl, but not l-fuconate, afforded protection against inactivation. Similar to the reaction of 3-HP with MR, 3-HP modified the Brønsted base catalyst (Lys 220) at the active site of FucD, which was facilitated by the Brønsted acid catalyst His 351. Thus, the interactions of tartronate and 3-HP with MR and FucD revealed differences in binding affinity and reactivity that differentiated between the enzymes' enolization apparatuses.

En Route to a Molecular Terminal Tin Oxide.

In the pursuit of terminal tin chalcogenides, heteroleptic stannylenes bearing terphenyl- and hexamethyldisilazide ligands were reacted with carbodiimides to yield the respective guanidinato complexes. Further supported by quantum chemical calculations, this revealed that the iso-propyl-substituted derivative provides the maximum steric protection achievable. Oxidation with elemental selenium produced monomeric terminal tin selenides with four-coordinate tin centers. In reactions with N2O as oxygen transfer reagent, silyl migration toward putative terminal tin oxide intermediates gave rise to tin complexes with terminal ─OSiMe3 functionality. To prevent silyl migration, the silyl groups were substituted with cyclohexyl moieties. This analogue exhibited distinctively different reactivities toward selenium and N2O, yielding a 1,2,3,4,5-tetraselenastannolane and chalcogenide-bridged dimeric compounds, respectively.

Biomechanics of predator-prey arms race in lion, zebra, cheetah and impala.

The fastest and most manoeuvrable terrestrial animals are found in savannah habitats, where predators chase and capture running prey. Hunt outcome and success rate are critical to survival, so both predator and prey should evolve to be faster and/or more manoeuvrable. Here we compare locomotor characteristics in two pursuit predator-prey pairs, lion-zebra and cheetah-impala, in their natural savannah habitat in Botswana. We show that although cheetahs and impalas were universally more athletic than lions and zebras in terms of speed, acceleration and turning, within each predator-prey pair, the predators had 20% higher muscle fibre power than prey, 37% greater acceleration and 72% greater deceleration capacity than their prey. We simulated hunt dynamics with these data and showed that hunts at lower speeds enable prey to use their maximum manoeuvring capacity and favour prey survival, and that the predator needs to be more athletic than its prey to sustain a viable success rate.

Structure of the glucagon receptor in complex with a glucagon analogue.

Class B G-protein-coupled receptors (GPCRs), which consist of an extracellular domain (ECD) and a transmembrane domain (TMD), respond to secretin peptides to play a key part in hormonal homeostasis, and are important therapeutic targets for a variety of diseases. Previous work has suggested that peptide ligands bind to class B GPCRs according to a two-domain binding model, in which the C-terminal region of the peptide targets the ECD and the N-terminal region of the peptide binds to the TMD binding pocket. Recently, three structures of class B GPCRs in complex with peptide ligands have been solved. These structures provide essential insights into peptide ligand recognition by class B GPCRs. However, owing to resolution limitations, the specific molecular interactions for peptide binding to class B GPCRs remain ambiguous. Moreover, these previously solved structures have different ECD conformations relative to the TMD, which introduces questions regarding inter-domain conformational flexibility and the changes required for receptor activation. Here we report the 3.0 Å-resolution crystal structure of the full-length human glucagon receptor (GCGR) in complex with a glucagon analogue and partial agonist, NNC1702. This structure provides molecular details of the interactions between GCGR and the peptide ligand. It reveals a marked change in the relative orientation between the ECD and TMD of GCGR compared to the previously solved structure of the inactive GCGR-NNC0640-mAb1 complex. Notably, the stalk region and the first extracellular loop undergo major conformational changes in secondary structure during peptide binding, forming key interactions with the peptide. We further propose a dual-binding-site trigger model for GCGR activation-which requires conformational changes of the stalk, first extracellular loop and TMD-that extends our understanding of the previously established two-domain peptide-binding model of class B GPCRs.

Publisher Correction: Cryo-EM structure of human rhodopsin bound to an inhibitory G protein.

In the PDF version of this Article, owing to a typesetting error, an incorrect figure was used for Extended Data Fig. 5; the correct figure was used in the HTML version. This has been corrected online.

Cryo-EM structure of human rhodopsin bound to an inhibitory G protein.

G-protein-coupled receptors comprise the largest family of mammalian transmembrane receptors. They mediate numerous cellular pathways by coupling with downstream signalling transducers, including the hetrotrimeric G proteins Gs (stimulatory) and Gi (inhibitory) and several arrestin proteins. The structural mechanisms that define how G-protein-coupled receptors selectively couple to a specific type of G protein or arrestin remain unknown. Here, using cryo-electron microscopy, we show that the major interactions between activated rhodopsin and Gi are mediated by the C-terminal helix of the Gi α-subunit, which is wedged into the cytoplasmic cavity of the transmembrane helix bundle and directly contacts the amino terminus of helix 8 of rhodopsin. Structural comparisons of inactive, Gi-bound and arrestin-bound forms of rhodopsin with inactive and Gs-bound forms of the ß2-adrenergic receptor provide a foundation to understand the unique structural signatures that are associated with the recognition of Gs, Gi and arrestin by activated G-protein-coupled receptors.

A new Finnish flavor of feline coat coloration, "salmiak," is associated with a 95-kb deletion downstream of the KIT gene.

Cats with a distinctive white hair pattern of unknown molecular cause have been discovered in the Finnish domestic cat population. Based on the unique appearance of these cats, we have named this phenotype salmiak ("salty licorice"). The use of a commercially available panel test to genotype four salmiak-colored cats revealed the absence of all known variants associated with white-haired phenotypic loci: full White (W), Spotting (Ws) and the Birman white Gloves associated (wg) allele of the KIT proto-oncogene (KIT) gene. Whole-genome sequencing on two salmiak-colored cats was conducted to search for candidate causal variants in the KIT gene. Despite a lack of coding variants, visual inspection of the short read alignments revealed a large ~95 kb deletion located ~65 kb downstream of the KIT gene in the salmiak cats. Additional PCR genotyping of 180 domestic cats and three salmiak-colored cats confirmed the homozygous derived variant genotype fully concordant with the salmiak phenotype. We suggest the newly identified variant be designated as wsal for "w salmiak".

Future time perspective: Factor structure, age pattern, and implications for psychological well-being and life satisfaction.

OBJECTIVE: Emerging research challenges the one-factor model of the future time perspective (FTP) scale by demonstrating two- and three-factor models of the FTP scale. METHOD: Three samples (i.e., Switzerland and the United States; N = 2022) explored the factor structure, age pattern differences, tested the link between FTP factors, psychological well-being, and life satisfaction, as well as age as a moderator. RESULTS: We distinguished FTP into opportunities, extension, and constraint factors that corroborated previous research. We did not find a replicable curvilinear age pattern difference on any of the FTP factors. The association between extension and life satisfaction was stronger among younger than older adults. Alternatively, the association between constraint and life satisfaction was stronger among younger than older adults in Samples A and C, and the pattern was reversed in Sample B. Lastly, the association between constraint and environmental mastery was stronger among older adults than younger adults in Sample B and the pattern was reversed in Sample C. CONCLUSION: This variation in the perception of the future is different for people at different periods in life and has an important consequence for living life well, especially a focus on extension and lack of constraint.

Fluid professional boundaries: ethnographic observations of co-located chiropractors, osteopaths and physiotherapists.

BACKGROUND: Chiropractors, osteopaths and physiotherapists (COPs) can assess and manage musculoskeletal conditions with similar manual or physical therapy techniques. This overlap in scope of practice raises questions about the boundaries between the three professions. Clinical settings where they are co-located are one of several possible influences on professional boundaries and may provide insight into the nature of these boundaries and how they are managed by clinicians themselves. OBJECTIVES: To understand the nature of professional boundaries between COPs within a co-located clinical environment and describe the ways in which professional boundaries may be reinforced, weakened, or navigated in this environment. METHODS: Drawing from an interpretivist paradigm, we used ethnographic observations to observe interactions between 15 COPs across two clinics. Data were analysed using reflexive thematic analysis principles. RESULTS: We identified various physical and non-physical 'boundary objects' that influenced the nature of the professional boundaries between the COPs that participated in the study. These boundary objects overall seemed to increase the fluidity of the professional boundaries, at times simultaneously reinforcing and weakening them. The boundary objects were categorised into three themes: physical, including the clinic's floor plan, large and small objects; social, including identities and discourse; and organisational, including appointment durations and fees, remuneration policies and insurance benefits. CONCLUSIONS: Physical, social, organisational related factors made the nature of professional boundaries between COPs in these settings fluid; meaning that they were largely not rigid or fixed but rather flexible, responsive and subject to change. These findings may challenge patients, clinicians and administrators to appreciate that traditional beliefs of distinct boundaries between COPs may not be so in co-located clinical environments. Both clinical practice and future research on professional boundaries between COPs may need to further consider some of these broader factors.

Ideology selectively shapes attention to inequality.

Contemporary debates about addressing inequality require a common, accurate understanding of the scope of the issue at hand. Yet little is known about who notices inequality in the world around them and when. Across five studies (N = 8,779) employing various paradigms, we consider the role of ideological beliefs about the desirability of social equality in shaping individuals' attention to-and accuracy in detecting-inequality across the class, gender, and racial domains. In Study 1, individuals higher (versus lower) on social egalitarianism were more likely to naturalistically remark on inequality when shown photographs of urban scenes. In Study 2, social egalitarians were more accurate at differentiating between equal versus unequal distributions of resources between men and women on a basic cognitive task. In Study 3, social egalitarians were faster to notice inequality-relevant changes in images in a change detection paradigm indexing basic attentional processes. In Studies 4 and 5, we varied whether unequal treatment adversely affected groups at the top or bottom of society. In Study 4, social egalitarians were, on an incentivized task, more accurate at detecting inequality in speaking time in a panel discussion that disadvantaged women but not when inequality disadvantaged men. In Study 5, social egalitarians were more likely to naturalistically point out bias in a pattern detection hiring task when the employer was biased against minorities but not when majority group members faced equivalent bias. Our results reveal the nuances in how our ideological beliefs shape whether we accurately notice inequality, with implications for prospects for addressing it.

Overcoming barriers to equality, diversity, inclusivity, and sense of belonging in healthcare education: the Underrepresented Groups' Experiences in Osteopathic Training (UrGEnT) mixed methods study.

BACKGROUND: Individuals from minority groups have historically faced social injustices. Those from underrepresented groups have been less likely to access both healthcare services and higher education. Little is known about the experiences of underrepresented students during their undergraduate studies in osteopathy in the UK. The aim of this project was to explore awareness of cultural diversity and beliefs about patients from underrepresented groups in current osteopathic educational environments and evaluate students' preparedness to manage patients from diverse groups. The project also aimed to investigate the educational experiences of students from underrepresented backgrounds during their training and their opinions on changes that could support better levels of recruitment and achievement. The findings were discussed with stakeholders in interactive workshops with the aim to develop recommendations for action and change. METHODS: A transformative action research paradigm informed this mixed methods project. It included: 1/ a survey of students from all seven osteopathic educational providers in the UK using the Multidimensional Cultural Humility Scale (MCHS); 2/ a series of focus groups with students from underrepresented groups (women, students with disabilities, students from minority ethnic backgrounds, and students identifying as LGBTQIA+); and 3/ a workshop forum to discuss findings. RESULTS: A total of 202 participants completed the MCHS and demographic questionnaire and seven focus groups were conducted. A model was developed to describe participants' training experiences comprising two main themes: institutional contextual obstacles (with four sub-themes) and underrepresented students' conceptual understanding of Equity, Diversity and Inclusion (EDI). Recommendations for change identified in the workshops were based on three topics: institutions, staff, and students. CONCLUSION: Our findings confirm conclusions from other institutions that staff education is urgently needed to create and maintain equitable, inclusive environments in osteopathic educational institutions in the UK to support all students, particularly those from underrepresented groups. Institutional EDI processes and policies also need to be clarified or modified to ensure their usefulness, accessibility, and implementation.