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Background: Asthmatic children present variable degrees of airway inflammation, remodeling, and resistance, which correlate with disease control and severity. The chronic inflammatory process of the airway triggers airway remodeling, which reflects the degree of airway resistance. Pro-inflammatory and pro-fibrotic mediators are centrally involved in this process., Objective: To investigate whether the levels of pulmonary and systemic pro-inflammatory and pro-fibrotic mediators present a correlation with the resistance of the respiratory system and of the proximal and distal airways., Methods: 39 Asthmatic children (persistent mild and moderate) and 39 non-asthmatic children (both between 6 and 13 years old) were evaluated for anthropometric characteristics, lung function and mechanics, and pulmonary and systemic immune responses., Results: Asthmatic children showed an increased number of blood eosinophils ( p  < 0.04), basophils ( p  < 0.04), monocytes ( p  < 0.002) and lymphocytes ( p  < 0.03). In addition, asthmatic children showed impaired lung function, as demonstrated by FEV1 ( p  < 0.0005) and FEV1/FVC ( p  < 0.004), decreased total resistance of the respiratory system (R5Hz; p  < 0.009), increased resistance of the proximal airways (R20Hz; p  < 0.02), increased elastance (Z5Hz; p  < 0.02) and increased reactance (X5Hz; p  < 0.002) compared to non-asthmatic children. Moreover, the following inflammatory factors were significantly higher in asthmatic than non-asthmatic children: GM-CSF in the breath condensate (BC) ( p  < 0.0001) and in the serum ( p  < 0.0001); TGF-beta in the BC ( p  < 0.0001) and in the serum ( p  < 0.004); IL-5 in the BC ( p  < 0.02) and in the serum ( p  < 0.01); IL-4 in the serum ( p  < 0.0002)., Conclusions: Impulse oscillometry is a sensitive method to detect airway resistance in persistent mild and moderate asthmatic children, an event followed by increased levels of pro-inflammatory and pro-fibrotic mediators.

Aim: Physical activity attenuates hypertension in older adults, but its impact on pulmonary function and mechanics in hypertensive older adults is unknown. The study seeks to understand whether a physically active lifestyle can improve respiratory capacity, the mechanical efficiency of the lungs, and, consequently, the quality of life of these individuals, comparing data between groups of active and sedentary hypertensive older adults., Methods: This is a cross-sectional study. We evaluated 731 older adults, stratified into two initial groups: hypertensive older adults (HE; n = 445) and non-hypertensive older adults (NHE; n = 286). For a secondary analysis, we used the International Physical Activity Questionnaire to sub-stratify HE and NHE into four groups: physically inactive hypertensive (PIH; n = 182), active hypertensive (AH; n = 110), physically inactive non-hypertensive (PINH; n = 104), and active non-hypertensive (ANH; n = 65). Lung function was measured by spirometry, and lung mechanics were assessed by impulse oscillometry., Results: Hypertensive older adults presented reduced lung function compared to non-hypertensive older adults, and physical inactivity accentuated this decline. Regarding pulmonary mechanics, hypertensive older adults had higher resistance of the entire respiratory system (R5 Hz), the central airways (R20 Hz), and peripheral airways (R5-20 Hz), which may trigger bronchoconstriction., Conclusions: Hypertension is associated with impaired lung function and mechanics in older adults, and a physically active lifestyle attenuates these dysfunctions.

Reputation systems promote cooperation and tie formation in social networks. But how reputations affect cooperation and the evolution of networks is less clear when societies are characterized by fundamental, identity-based, social divisions like those centered on politics in the contemporary U.S. Using a large web-based experiment with participants (N = 1073) embedded in networks where each tie represents the opportunity to play a dyadic iterated prisoners' dilemma, we investigate how cooperation and network segregation varies with whether and how reputation systems track behavior toward members of the opposing political party (outgroup members). As predicted, when participants know others' political affiliation, early cooperation patterns show ingroup favoritism. As a result, networks become segregated based on politics. However, such ingroup favoritism and network-level political segregation is reduced in conditions in which participants know how others behave towards participants from both their own party and participants from the other party. These findings have implications for our understanding of reputation systems in polarized contexts., (© 2023. The Author(s).)

Drosophila ovarian Follicle Stem Cells (FSCs) present a favorable paradigm for understanding how stem cell division and differentiation are balanced in communities where those activities are independent. FSCs also allow exploration of how this balance is integrated with spatial stem cell heterogeneity. Posterior FSCs become proliferative Follicle Cells (FCs), while anterior FSCs become quiescent Escort Cells (ECs) at about one fourth the frequency. A single stem cell can nevertheless produce both FCs and ECs because it can move between anterior and posterior locations. Studies based on EdU incorporation to approximate division rates suggested that posterior FSCs divide faster than anterior FSCs. However, direct measures of cell cycle times are required to ascertain whether FC output requires a net flow of FSCs from anterior to posterior. Here, by using live imaging and FUCCI cell-cycle reporters, we measured absolute division rates. We found that posterior FSCs cycle more than three times faster than anterior FSCs and produced sufficient new cells to match FC production. H2B-RFP dilution studies supported different cycling rates according to A/P location and facilitated live imaging, showing A/P exchange of FSCs in both directions, consistent with the dynamic equilibrium inferred from division rate measurements. Inversely graded Wnt and JAK-STAT pathway signals regulate FSC differentiation to ECs and FCs. JAK-STAT promotes both differentiation to FCs and FSC cycling, affording some coordination of these activities. When JAK-STAT signaling was manipulated to be spatially uniform, the ratio of posterior to anterior division rates was reduced but remained substantial, showing that graded JAK-STAT signaling only partly explains the graded cycling of FSCs. By using FUCCI markers, we found a prominent G2/M cycling restriction of posterior FSCs together with an A/P graded G1/S restriction, that JAK-STAT signaling promotes both G1/S and G2/M transitions, and that PI3 kinase signaling principally stimulates the G2/M transition., Competing Interests: No competing interests, (Copyright: © 2023 Melamed et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Recent results have shown that the human malaria-resistant hemoglobin S mutation originates de novo more frequently in the gene and in the population where it is of adaptive significance, namely, in the hemoglobin subunit beta gene compared to the nonresistant but otherwise identical 20A[Formula: see text]T mutation in the hemoglobin subunit delta gene, and in sub-Saharan Africans, who have been subject to intense malarial pressure for many generations, compared to northern Europeans, who have not. This finding raises a fundamental challenge to the traditional notion of accidental mutation. Here, we address this finding with the replacement hypothesis, according to which preexisting genetic interactions can lead directly and mechanistically to mutations that simplify and replace them. Thus, an evolutionary process under selection can gradually hone in on interactions of importance for the currently evolving adaptations, from which large-effect mutations follow that are relevant to these adaptations. We exemplify this hypothesis using multiple types of mutation, including gene fusion mutations, gene duplication mutations, A[Formula: see text]G mutations in RNA-edited sites and transcription-associated mutations, and place it in the broader context of a system-level view of mutation origination called interaction-based evolution. Potential consequences include that similarity of mutation pressures may contribute to parallel evolution in genetically related species, that the evolution of genome organization may be driven by mutational mechanisms, that transposable element movements may also be explained by replacement, and that long-term directed mutational responses to specific environmental pressures are possible. Such mutational phenomena need to be further tested by future studies in natural and artificial settings., (© 2023. The Author(s).)

We review an array of experimental methodological factors that either contribute to or detract from the measurement of pragmatic implicatures in child language. We carry out a truth value judgment task to measure children's interpretations of the Spanish existential quantifier algunos in implicature-consistent and implicature-inconsistent contexts. Independently, we take measures of children's inhibition, working memory, attention, approximate number ability, phrasal syntax, and lexicon. We model the interplay of these variables using a piecewise structural equation model (SEM), common in the life sciences, but not in the social and behavioral sciences. By 6 years of age, the children in our sample were not statistically different from adults in their interpretations. Syntax, lexicon, and inhibition significantly predict implicature generation, each accounting for unique variance. The approximate number system and inhibition significantly predict lexical development. The statistical power of the piecewise SEM components, with a sample of 64 children, is high, in comparison to a traditional, globally estimated SEM of the same data.

Cases of parallel or recurrent gene fusions in evolution as well as in genetic disease and cancer are difficult to explain, because unlike point mutations, they can require the repetition of a similar configuration of multiple breakpoints rather than the repetition of a single point mutation. The used-together-fused-together hypothesis holds that genes that are used together repeatedly and persistently in a specific context are more likely to undergo fusion mutation in the course of evolution for mechanistic reasons. This hypothesis offers to explain gene fusion in both evolution and disease under one umbrella. Using bioinformatic data, we tested this hypothesis against alternatives, including that all gene pairs can fuse by random mutation, but among pairs thus fused, those that had interacted previously are more likely to be favored by selection. Results show that across multiple measures of gene interaction, human genes whose orthologs are fused in one or more species are more likely to interact with each other than random pairs of genes of the same genomic distance between pair members; that an overlap exists between genes that fused in the course of evolution in non-human species and genes that undergo fusion in human cancers; and that across six primate species studied, fusions predominate over fissions and exhibit substantial evolutionary parallelism. Together, these results support the used-together-fused-together hypothesis over its alternatives. Multiple implications are discussed, including the relevance of mutational mechanisms to the evolution of genome organization, to the distribution of fitness effects of mutation, to evolutionary parallelism and more., Supplementary Information: The online version contains supplementary material available at 10.1007/s11692-022-09579-9., Competing Interests: Conflict of InterestThe authors declare that they have no conflict of interest., (© The Author(s) 2022.)

Objectives: The mRNA-based COVID-19 vaccines are now employed globally and have shown high efficacy in preventing SARS-CoV-2 infection. However, less is known about the vaccine efficacy in immune-suppressed individuals. This study sought to explore whether humoral immunity to the COVID-19 vaccine BNT162b2 is altered in RA patients treated with Janus kinase inhibitors by analysing their antibodies titre, neutralization activity and B cell responses., Methods: We collected plasma samples from 12 RA patients who were treated with Janus kinase inhibitors and received two doses of the BNT162b2 vaccine, as well as 26 healthy individuals who were vaccinated with the same vaccine. We analysed the quantity of the anti-spike IgG and IgA antibodies that were elicited following the BNT162b2 vaccination, the plasma neutralization capacity and the responsiveness of the B-lymphocytes. We used ELISA to quantify the antibody titres, and a plasma neutralization assay was used to determine the virus neutralization capacity. Alteration in expression of the genes that are associated with B cell activation and the germinal centre response were analysed by quantitative PCR., Results: Reduced levels of anti-spike IgG antibodies and neutralization capacity were seen in the RA patients who were treated with JAK inhibitors in comparison with healthy individuals. Furthermore, B cell responsiveness to the SARS-CoV-2 spike protein was reduced in the RA patients., Conclusion: RA patients who are treated with JAK inhibitors show a suppressed humoral response following BNT162b2 vaccination, as revealed by the quantity and quality of the anti-spike antibodies., (© The Author(s) 2021. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Social networks are fundamental to the broad scale cooperation observed in human populations. But by structuring the flow of benefits from cooperation, networks also create and sustain macro-level inequalities. Here we ask how two aspects of inequality shape the evolution of cooperation in dynamic social networks. Results from a crowdsourced experiment (N = 1080) show that inequality alters the distribution of cooperation within networks such that participants engage in more costly cooperation with their wealthier partners in order to maintain more valuable connections to them. Inequality also influences network dynamics, increasing the tendency for participants to seek wealthier partners, resulting in structural network change. These processes aggregate to alter network structures and produce greater system-level inequality. The findings thus shed critical light on how networks serve as both boon and barrier to macro-level human flourishing., (© 2022. The Author(s).)

Macrophages are some of the most important immune cells in the organism and are responsible for creating an inflammatory immune response in order to inhibit the passage of microscopic foreign bodies into the blood stream. Sometimes, their activation can be responsible for chronic inflammatory diseases such as asthma, tuberculosis, hepatitis, sinusitis, inflammatory bowel disease, and viral infections. Prolonged inflammation can damage the organs or may lead to death in serious conditions. In the present study, RAW264.7 macrophages were exposed to lipopolysaccharide (LPS; 20 ng/mL) and simultaneously treated with 20 µg/mL of natural-based formulation (NBF), mushroom-cannabidiol extract). Pro-inflammatory cytokines, chemokines, and other inflammatory markers were analyzed. The elevations in the presence of interleukin-6 (IL-6), cycloxygenase-2 (COX-2), C-C motif ligand-5 (CCL5), and nitrite response, following exposure to LPS, were completely inhibited by NBF administration. IL-1β and tumor necrosis factor alpha (TNF-α) release were inhibited by 3.9-fold and 1.5-fold, respectively. No toxic effect of NBF, as assessed by lactate dehydrogenase (LDH) release, was observed. Treatment of the cells with NBF significantly increased the mRNA levels of TLR2, and TLR4, but not NF-κB. Thus, it appears that the NBF possesses anti-inflammatory and immunomodulatory effects which can attenuate the release of pro-inflammatory markers. NBF may be a candidate for the treatment of acute and chronic inflammatory diseases and deserves further investigation.

Although it is known that the mutation rate varies across the genome, previous estimates were based on averaging across various numbers of positions. Here, we describe a method to measure the origination rates of target mutations at target base positions and apply it to a 6-bp region in the human hemoglobin subunit beta ( HBB ) gene and to the identical, paralogous hemoglobin subunit delta ( HBD ) region in sperm cells from both African and European donors. The HBB region of interest (ROI) includes the site of the hemoglobin S (HbS) mutation, which protects against malaria, is common in Africa, and has served as a classic example of adaptation by random mutation and natural selection. We found a significant correspondence between de novo mutation rates and past observations of alleles in carriers, showing that mutation rates vary substantially in a mutation-specific manner that contributes to the site frequency spectrum. We also found that the overall point mutation rate is significantly higher in Africans than in Europeans in the HBB region studied. Finally, the rate of the 20A→T mutation, called the "HbS mutation" when it appears in HBB , is significantly higher than expected from the genome-wide average for this mutation type. Nine instances were observed in the African HBB ROI, where it is of adaptive significance, representing at least three independent originations; no instances were observed elsewhere. Further studies will be needed to examine mutation rates at the single-mutation resolution across these and other loci and organisms and to uncover the molecular mechanisms responsible., (© 2022 Melamed et al.; Published by Cold Spring Harbor Laboratory Press.)

This study aimed to determine the free radical scavenging and antioxidant potential of hot water extracts prepared from different combinations and ratios of submerged cultivated mycelial biomass of medicinal mushrooms. Total phenolic compounds, flavonoid content, and antioxidant activity were evaluated for combined crude hot water extracts from medicinal higher Basidiomycetes mushrooms belonging to ten genera. The results demonstrate that almost all tested combinations were good sources of phenolic compounds and flavonoids, ranging between 16.42 and 18.83 gallic acid equivalents/g and 1.5 and 4.34 mg rutin equivalents/g, respectively. Moreover, free radical scavenging properties were evaluated with the DPPH and ABTS assays and metal chelating effects were investigated. All tested samples and/or extracts demonstrated significant free radical scavenging properties and antioxidant potential.

Loss of B lymphocyte regeneration in the bone marrow (BM) is an immunologic hallmark of advanced age, which impairs the replenishment of peripheral B-cell subsets and results in impaired humoral responses, thereby contributing to immune system dysfunction associated with aging. A better understanding of the mechanism behind this loss may suggest ways to restore immune competence and promote healthy aging. In this study, we uncover an immune-endocrine regulatory circuit that mediates cross-talk between peripheral B cells and progenitors in the BM, to balance B-cell lymphopoiesis in both human and mouse aging. We found that tumor necrosis factor α (TNF-α), which is increasingly produced by peripheral B cells during aging, stimulates the production of insulin-like growth factor-binding protein 1 (IGFBP-1), which binds and sequesters insulin-like growth factor 1 (IGF-1) in the circulation, thereby restraining its activity in promoting B-cell lymphopoiesis in the BM. Upon B-cell depletion in aging humans and mice, circulatory TNF-α decreases, resulting in increased IGF-1 and reactivation of B-cell lymphopoiesis. Perturbation of this circuit by administration of IGF-1 to old mice or anti-TNF-α antibodies to human patients restored B-cell lymphopoiesis in the BM. Thus, we suggest that in both human and mouse aging, peripheral B cells use the TNF-α/IGFBP-1/IGF-1 axis to repress B-cell lymphopoiesis. This trial was registered at www.clinicaltrials.govas#NCT00863187., (© 2021 by The American Society of Hematology.)

The ability to design a protein to bind specifically to a target RNA enables numerous applications, with the modular architecture of the PUF domain lending itself to new RNA-binding specificities. For each repeat of the Pumilio-1 PUF domain, we generate a library that contains the 8,000 possible combinations of amino acid substitutions at residues critical for RNA contact. We carry out yeast three-hybrid selections with each library against the RNA recognition sequence for Pumilio-1, with any possible base present at the position recognized by the randomized repeat. We use sequencing to score the binding of each variant, identifying many variants with highly repeat-specific interactions. From these data, we generate an RNA binding code specific to each repeat and base. We use this code to design PUF domains against 16 RNAs, and find that some of these domains recognize RNAs with two, three or four changes from the wild type sequence., (© 2021. The Author(s).)

This research describes the investigation of submerged cultivated mycelial biomass and hot water extracts prepared from different combinations and ratios of medicinal mushroom (MM) dry powders, comprising various biologically active compounds/secondary metabolites. In particular, it was evaluated the proximate composition (moisture, ash, crude protein, fat, total carbohydrates, and total energy), γ-aminobutyric acid (GABA) and ergothioneine (ERG), amino acid content of mycelia of 16 higher Basidiomycetes MM species. The results obtained demonstrate that almost all tested combinations were found to be good sources of polysaccharides, with content varying in the ranges of 4.73 ± 1.33% and 58.46 ± 4.15%. Total protein contents varied in 1.97 ± 0.40% - 5.37 ± 0.40% range. ERG was detected in all tested samples, while GABA existed only in eight samples out of 15 and varied from 0.03 ± < 0.01 to 0.61 ± 0.03 mg/g, and from 0.16 ± 0.03 to 5.69 ± 0.41 mg/g respectively. Analyses of total phenolic and flavonoid contents demonstrate considerable content in all samples (15.53 ± 0.23 - 18.88 ± 0.34 mg gallic acid equivalents/g and 1.23 ± 0.04 - 4.34 ± 0.73 mg rutin equivalents/g respectively). In present research the complexity of samples/extracts were evaluated by multiple antioxidant assays to verify their antioxidant capacity. Determination of in vitro antioxidant activity was successfully carried out by several different methods such as 2,2-diphenyl-1-picrylhydrazyl scavenging activity, reducing power, chelating ability, hydroxyl radical scavenging activity, and 2,2'-azino-bis(3-ethylbenzothi-azoline-6-sulfonic acid scavenging activity. Therefore, all tested samples confirm the capable antioxidant activities of bioactive compounds extracted from MMs.

The phosphatidylinositol 3-kinase (PI3K) signaling cascade downstream of the B cell receptor (BCR) signalosome is essential for B cell maturation. Proper signaling strength is maintained through the PI3K negative regulator phosphatase and tensin homolog (PTEN). Although a role for microRNA (miRNA)-dependent control of the PTEN-PI3K axis has been described, the contribution of individual miRNAs to the regulation of this crucial signaling modality in mature B lymphocytes remains to be elucidated. Our analyses reveal that ablation of miR-29 specifically in B lymphocytes results in an increase in PTEN expression and dampening of the PI3K pathway in mature B cells. This dysregulation has a profound impact on the survival of B lymphocytes and results in increased class switch recombination and decreased plasma cell differentiation. Furthermore, we demonstrate that ablation of one copy of Pten is sufficient to ameliorate the phenotypes associated with miR-29 loss. Our data suggest a critical role for the miR-29-PTEN-PI3K regulatory axis in mature B lymphocytes., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Many adult stem cell communities are maintained by population asymmetry, where stochastic behaviors of multiple individual cells collectively result in a balance between stem cell division and differentiation. We investigated how this is achieved for Drosophila Follicle Stem Cells (FSCs) by spatially-restricted niche signals. FSCs produce transit-amplifying Follicle Cells (FCs) from their posterior face and quiescent Escort Cells (ECs) to their anterior. We show that JAK-STAT pathway activity, which declines from posterior to anterior, dictates the pattern of divisions over the FSC domain, promotes more posterior FSC locations and conversion to FCs, while opposing EC production. Wnt pathway activity declines from the anterior, promotes anterior FSC locations and EC production, and opposes FC production. The pathways combine to define a stem cell domain through concerted effects on FSC differentiation to ECs and FCs at either end of opposing signaling gradients, and impose a pattern of proliferation that matches derivative production., Competing Interests: DM, DK No competing interests declared, (© 2020, Melamed and Kalderon.)

Prosocial behavior is paradoxical because it often entails a cost to one's own welfare to benefit others. Theoretical models suggest that prosociality is driven by several forms of reciprocity. Although we know a great deal about how each of these forms operates in isolation, they are rarely isolated in the real world. Rather, the topological features of human social networks are such that people are often confronted with multiple types of reciprocity simultaneously. Does our current understanding of human prosociality break down if we account for the fact that the various forms of reciprocity tend to co-occur in nature? Results of a large experiment show that each basis of human reciprocity is remarkably robust to the presence of other bases. This lends strong support to existing models of prosociality and puts theory and research on firmer ground in explaining the high levels of prosociality observed in human social networks., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

In this paper, we extend the logic of existing sociological theory on status to explain how status processes can inform selection in competitive choice situations. We argue that in the absence of knowledge about the specific abilities of others and assuming a desire to win, when given the opportunity to "pick their battles," people will draw on overt status differences as a basis for selecting a competitor from a pool of possible competitors. Results from three studies indicate that, as predicted, status differences affect competitor selection, with individuals choosing to compete against those who are relatively lower status based on diffuse characteristics. Moreover, consistent with expectation state theories, results from two studies show that the expectations that people form for their potential competitors based on status differences mediate this relationship. We conclude by discussing the implications of this research., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Forecasting extremely rare events is a pressing problem, but efforts to model such outcomes are often limited by the presence of multiple causes within classes of events, insufficient observations of the outcome to assess fit, and biased estimates due to insufficient observations of the outcome. We introduce a novel approach for analyzing rare event data that addresses these challenges by turning attention to the conditions under which rare outcomes do not occur. We detail how configurational methods can be used to identify conditions or sets of conditions that would preclude the occurrence of a rare outcome. Results from Monte Carlo experiments show that our approach can be used to systematically preclude up to 78.6% of observations, and application to ground-truth data coupled with a bootstrap inferential test illustrates how our approach can also yield novel substantive insights that are obscured by standard statistical analyses., Competing Interests: The authors have declared that no competing interests exist.

Aging is associated with increasing prevalence and severity of infections caused by a decline in bone marrow (BM) lymphopoiesis and reduced B-cell repertoire diversity. The current study proposes a strategy to enhance immune responsiveness in aged mice and humans, through rejuvenation of the B lineage upon B-cell depletion. We used hCD20Tg mice to deplete peripheral B cells in old and young mice, analyzing B-cell subsets, repertoire and cellular functions in vitro, and immune responsiveness in vivo. Additionally, elderly patients, previously treated with rituximab healthy elderly and young individuals, were vaccinated against hepatitis B (HBV) after undergoing a detailed analysis for B-cell compartments. B-cell depletion in old mice resulted in rejuvenated B-cell population that was derived from de novo synthesis in the bone marrow. The rejuvenated B cells exhibited a "young"-like repertoire and cellular responsiveness to immune stimuli in vitro. Yet, mice treated with B-cell depletion did not mount enhanced antibody responses to immunization in vivo, nor did they survive longer than control mice in "dirty" environment. Consistent with these results, peripheral B cells from elderly depleted patients showed a "young"-like repertoire, population dynamics, and cellular responsiveness to stimulus. Nevertheless, the response rate to HBV vaccination was similar between elderly depleted and nondepleted subjects, although antibody titers were higher in depleted patients. This study proposes a proof of principle to rejuvenate the peripheral B-cell compartment in aging, through B-cell depletion. Further studies are warranted in order to apply this approach for enhancing humoral immune responsiveness among the elderly population., (© 2019 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.)

microRNAs (miRNAs) down-modulate the levels of proteins by sequence-specific binding to their respective target mRNAs, causing translational repression or mRNA degradation. The miR-17∼92 cluster encodes for six miRNAs whose target recognition specificities are determined by their distinct sequence. In mice, the four miRNA families generated from the miR-17∼92 cluster coordinate to allow for proper lymphocyte development and effective adaptive immune responses following infection or immunization. Lymphocyte development and homeostasis rely on tight regulation of PI3K signaling to avoid autoimmunity or immunodeficiency, and the miR-17∼92 miRNAs appear as key mediators to appropriately tune PI3K activity. On the other hand, in lymphoid tumors overexpression of the miR-17∼92 miRNAs is a common oncogenic event. In this review, we touch on what we have learned so far about the miR-17∼92 miRNAs, particularly with respect to their role in lymphocyte development, homeostasis and pathology., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: The aim of this study was to report the prevalence and mortality associated with anticoagulant-related nephropathy (ARN) through a systematic review of the literature., Methods: Electronic searches were conducted in the Medline and EMBASE databases, and manual searches were performed in the reference lists of the identified studies. The studies were selected by two independent researchers, first by evaluating the titles and abstracts and then by reading the complete texts of the identified studies. Case series, cross-sectional studies, cohort studies and case-control studies reporting the prevalence and factors associated with ARN were selected. The methodological quality was assessed using the Newcastle-Ottawa scale. Meta-analyses of the prevalence of ARN and 5-year mortality using the random effects model were performed when possible. Heterogeneity was assessed using the I 2 statistic., Results: Five studies were included. Prevalence of ARN ranged from 19% to 63% among the four included cohort studies. Meta-analysis of these resulted in high heterogeneity [ I 2 96%, summary effect 31%; 95% confidence interval (CI) 22-42%]. Subgroup meta-analysis yielded an ARN prevalence of 20% among studies that included patients with fewer comorbidities ( I 2 12%; 95% CI 19-22%). In a direct comparison, meta-analysis of the 5-year mortality rate between anticoagulated patients who had experienced ARN and anticoagulated patients without ARN, patients with ARN were 91% more likely to die (risk ratio = 1.91; 95% CI 1.22-3; I 2 87%). Risk factors for ARN that were reported in the literature included initial excessive anticoagulation, chronic kidney disease, age, diabetes, hypertension, cardiovascular disease and heart failure., Conclusions: ARN studies are scarce and heterogeneous, and present significant methodological limitations. The high prevalence of ARN reported herein suggests that this entity is underdiagnosed in clinical practice. Mortality in patients with ARN seems to be high compared with patients without this condition in observational studies.

Dynamic networks, where ties can be shed and new ties can be formed, promote the evolution of cooperation. Yet, past research has only compared networks where all ties can be severed to those where none can, confounding the benefits of fully dynamic networks with the presence of some dynamic ties within the network. Further, humans do not live in fully dynamic networks. Instead, in real-world networks, some ties are subject to change, while others are difficult to sever. Here, we consider whether and how cooperation evolves in networks containing both static and dynamic ties. We argue and find that the presence of dynamic ties in networks promotes cooperation even in static ties. Consistent with previous work demonstrating that cooperation cascades in networks, our results show that cooperation is enhanced in networks with both tie types because the higher rate of cooperation that occurs following the dynamics process "spills over" to those relations that are more difficult to alter. Thus, our findings demonstrate the critical role that dynamic ties play in promoting cooperation by altering behavioral outcomes even in non-dynamic relations.

Appropriate PI3K signals generated by the antigen receptor are essential to promote B cell development. Regulation of recombination activating gene (RAG)-1 and RAG-2 expression is one key process that is mediated by PI3K to ensure developmental progression and selection. When PI3K signals are too high or too low, expression of RAGs does not turn off and B cell development is impaired or blocked. Yet, the mechanism which tunes PI3K activity to control RAG expression during B cell development in the bone marrow is unknown. Recently we showed that a c-Myc/miR17-92/PTEN axis regulates PI3K activity for positive and negative selection of immature B cells. Here, we show that the c-Myc/miR17-92/PTEN axis tunes PI3K activity to control the expression of RAGs in proB cells. Using different genetically engineered mouse models we show that impaired function of the c-Myc/miR17-92/PTEN axis alters the PI3K/Akt/Foxo1 pathway to result in dis-regulated expression of RAG and a block in B cell development. Studies using 38c-13 B lymphoma cells, where RAGs are constitutively expressed, suggest that this regulatory effect is mediated post-translationally through Foxo1.

Background: Psychotic experiences might represent non-specific markers of poor mental health in adolescence. However, only a few predominantly cross-sectional studies have tested their association with disordered eating behaviours in adolescent and adult populations. The aim of this study was to explore the association between psychotic experiences at age 13 years, and disordered eating behaviours and body-mass index (BMI) at age 18 years., Methods: We used data from the Avon Longitudinal Study of Parents and Children, a longitudinal birth cohort based in Avon (England, UK) including mothers with an expected delivery date between April 1, 1991, and Dec 31, 1992, and their children. Psychotic experiences (such as delusions and hallucinations) and BMI were measured at clinical assessments when children were nearly aged 13 years, and data on disordered eating behaviours (ie, presence of binge eating, purging, fasting, or excessive exercise for weight loss; any of these behaviours [included to increase statistical power]; and number of behaviours [included to investigate severity]) were obtained via a postal questionnaire that used adapted questions from the Youth Risk Behaviour Surveillance System questionnaire at approximately age 18 years. For each outcome, we ran a univariable model and four multivariable models (logistic, linear [for BMI], or negative binomial [for the number of behaviours] regression), progressively adjusting for child and maternal sociodemographic, physical, and mental health characteristics (including child's sex, and maternal age, marital status, and highest academic qualification); autistic traits at age 7 years (measured with the Social and Communication Disorder Checklist); baseline BMI at age 13 years, and depressive symptoms at baseline (ie, at age 13 years when psychotic experiences were measured: childs' symptoms measured with the Moods and feelings Questionnaire, and maternal symptoms measured at 32 weeks' gestation with the Edinburgh Postnatal Depression Scale). We imputed missing outcome and covariate data., Findings: Our sample included 6361 children, of whom 734 (12%) reported psychotic experiences at age 13 years. In univariable models, psychotic experiences were associated with greater odds of reporting any disordered eating behaviours (odds ratio [OR] 1·92, 95% CI 1·46-2·52; p<0·0001), and more severe symptoms (as measured by the number of disordered eating behaviours: 0·58, 0·32-0·84; p<0·0001) at age 18 years. These associations were slightly attenuated by adjustment for maternal and child characteristics (any disordered eating behaviours OR 1·82, 95% CI 1·35-2·44, p<0·0001; number of disordered eating behaviours 0·49, 95% CI 0·23-0·75, p<0·00001), autistic traits at age 7 years (any disordered eating behaviours OR 1·80, 95% CI 1·34-2·41, p<0·0001; number of disordered eating behaviours 0·48, 95% CI 0·22-0·74, p<0·00001), and BMI (any disordered-eating behaviours OR 1·83, 95% CI 1·36-2·46, p<0·0001; number of disordered-eating behaviours 0·32, 95% CI 0·06-0·57, p<0·00001) Adjusting for baseline depressive symptoms attenuated, but not removed, these associations (any disordered eating OR 1·50, 95% CI 1·10-2·03, p=0·010; more severe symptoms 0·32, 0·06-0·57, p=0·017). Psychotic experiences were also associated with greater binge eating, purging, and fasting behaviours, although some associations weakened after controlling for depressive symptoms. We noted no associations between psychotic experiences and excessive exercise or BMI in any of the models., Interpretation: Our findings suggested that psychotic experiences are markers of increased risk for several disordered eating behaviours in late adolescence, possibly by indicating more severe psychopathology in early adolescence. More research investigating shared risk factors for psychotic experiences and eating disorders is warranted to elucidate shared and specific causal pathways., Funding: Wellcome Trust, the Royal Society, University College London Hospitals National Institute for Health Research Biomedical Research Centre, UK Medical Research Council, and the University of Bristol., (Copyright © 2018 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.)

Cancer-initiating gatekeeper mutations that arise in stem cells would be especially potent if they stabilize and expand an affected stem cell lineage. It is therefore important to understand how different stem cell organization strategies promote or prevent variant stem cell amplification in response to different types of mutation, including those that activate proliferation. Stem cell numbers can be maintained constant while producing differentiated products through individually asymmetrical division outcomes or by population asymmetry strategies in which individual stem cell lineages necessarily compete for niche space. We considered alternative mechanisms underlying population asymmetry and used quantitative modeling to predict starkly different consequences of altering proliferation rate: A variant, faster proliferating mutant stem cell should compete better only when stem cell division and differentiation are independent processes. For most types of stem cells, it has not been possible to ascertain experimentally whether division and differentiation are coupled. However, Drosophila follicle stem cells (FSCs) provided a favorable system with which to investigate population asymmetry mechanisms and also for measuring the impact of altered proliferation on competition. We found from detailed cell lineage studies that division and differentiation of an individual FSC are not coupled. We also found that FSC representation, reflecting maintenance and amplification, was highly responsive to genetic changes that altered only the rate of FSC proliferation. The FSC paradigm therefore provides definitive experimental evidence for the general principle that relative proliferation rate will always be a major determinant of competition among stem cells specifically when stem cell division and differentiation are independent., Competing Interests: The authors declare no conflict of interest.

Humans' propensity to cooperate is driven by our embeddedness in social networks. A key mechanism through which networks promote cooperation is clustering. Within clusters, conditional cooperators are insulated from exploitation by noncooperators, allowing them to reap the benefits of cooperation. Dynamic networks, where ties can be shed and new ties formed, allow for the endogenous emergence of clusters of cooperators. Although past work suggests that either reputation processes or network dynamics can increase clustering and cooperation, existing work on network dynamics conflates reputations and dynamics. Here we report results from a large-scale experiment (total n = 2,675) that embedded participants in clustered or random networks that were static or dynamic, with varying levels of reputational information. Results show that initial network clustering predicts cooperation in static networks, but not in dynamic ones. Further, our experiment shows that while reputations are important for partner choice, cooperation levels are driven purely by dynamics. Supplemental conditions confirmed this lack of a reputation effect. Importantly, we find that when participants make individual choices to cooperate or defect with each partner, as opposed to a single decision that applies to all partners (as is standard in the literature on cooperation in networks), cooperation rates in static networks are as high as cooperation rates in dynamic networks. This finding highlights the importance of structured relations for sustained cooperation, and shows how giving experimental participants more realistic choices has important consequences for whether dynamic networks promote higher levels of cooperation than static networks., Competing Interests: The authors declare no conflict of interest.

Adult stem cells provide a renewable source of differentiated cells for a wide variety of tissues and generally give rise to multiple cell types. Basic principles of stem cell organization and regulation underlying this behaviour are emerging. Local niche signals maintain stem cells, while different sets of signals act outside the niche to diversify initially equivalent stem cell progeny. Here we show that Drosophila ovarian follicle stem cells (FSCs) produced two distinct cell types directly. This cell fate choice was determined by the anterior-posterior position of an FSC and by the magnitude of spatially graded Wnt pathway activity. These findings reveal a paradigm of immediate diversification of stem cell derivatives according to stem cell position within a larger population, guided by a graded niche signal. We also found that FSCs strongly resemble mammalian intestinal stem cells in many aspects of their organization, including population asymmetry and dynamic heterogeneity.

Dynamic networks have been shown to increase cooperation, but prior findings are compatible with two different mechanisms for the evolution of cooperation. It may be that dynamic networks promote cooperation even in networks composed entirely of egoists, who strategically cooperate to attract and maintain profitable interaction partners. Alternatively, drawing on recent insights into heterogeneous social preferences, we expect that dynamic networks will increase cooperation only when nodes are occupied by persons with more prosocial preferences, who tend to attract and keep more cooperative partners relative to egoists. Our experiment used a standard procedure to classify participants a priori as egoistic or prosocial and then embedded them in homogeneous networks of all prosocials or all egoists, or in heterogeneous networks (50/50). Participants then interacted in repeated prisoner's dilemma games with alters in both static and dynamic networks. In both heterogeneous and homogeneous networks, we find dynamic networks only promote cooperation among prosocials. Resulting from their greater cooperation, prosocials' relations are more stable, yielding substantially higher fitness compared to egoists in both heterogeneous and homogeneous dynamic networks. Our results suggest that a key to the evolution and stability of cooperation is the ability of those with prosocial preferences to alter their networks.

The extracellular-regulated kinase (Erk) pathway is a major determinant in the control of diverse cellular processes, such as proliferation, differentiation, survival, and motility. The pathway executes its effects through kinases of the Erk family. Erks are not only critical for a variety of physiological processes, but are also associated with neurodegenerative diseases, cardiovascular diseases, diabetes and a large number of human cancers. However, the exact role of each Erk molecule in these biological and pathological processes is not fully determined. An efficient strategy for revealing these roles is to activate each Erk isoform individually, in a signal independent manner, and to monitor the molecular, physiological, and pathological effects. This could be achieved by developing intrinsically active variants for each Erk isoform and splicing variant and expressing these molecules individually in biological systems. A screening method that selects for relevant and useful active mutants of Erks is described in this chapter. The main principle of the method is to screen for mutants of Erk that function in the total absence of their relevant MEKs. Another principle is that the screen should be unbiased toward particular domains or mechanisms of action. We describe how these principles are combined into a screen that takes advantage of the yeast Mpk1/Erk pathway. Following the description of how intrinsically active Mpk1 molecules are isolated, we provide comprehensive and detailed descriptions of the methods used to characterize their catalytic activity, autophosphorylation capabilities, and phosphorylation status, as well as the methods used to determine the precise phosphorylated sites. The principles of the screen and the methods described here could be easily adapted for any Erk molecule in any organism.

Justice research has evolved by elucidating the factors that affect justice evaluations, as well as their consequences. Unfortunately, few researchers have paid attention to the pattern of rewards over time as a predictor of justice evaluations. There are two main objectives of this research. First, it aims to test the effect of reward stability on justice evaluations. Based on justice theory and prospect theory, we assume that an under-reward at one time cannot be fully offset by an equivalent over-reward at another time. Therefore, in unstable reward systems the asymmetry of the effect of unjust rewards with opposite directions will produce a lower level of justice evaluations over time. The second objective of this research is to show the moderating effect of the presentation order (primacy vs. recency) of unstable rewards on justice evaluations. The results from a controlled experiment with five conditions, which presents the instability of rewards in different orders, confirm both the negative effect of unstable rewards and the stronger effect of primacy on justice evaluations., Competing Interests: The authors have declared that no competing interests exist.

B cell development is a tightly regulated process dependent on sequential rearrangements of immunoglobulin loci that encode the antigen receptor. To elucidate the role of microRNAs (miRNAs) in the orchestration of B cell development, we ablated all miRNAs at the earliest stage of B cell development by conditionally targeting the enzymes critical for RNAi in early B cell precursors. Absence of any one of these enzymes led to a block at the pro- to pre-B cell transition due to increased apoptosis and a failure of pre-B cells to proliferate. Expression of a Bcl2 transgene allowed for partial rescue of B cell development, however, the majority of the rescued B cells had low surface immunoglobulin expression with evidence of ongoing light chain editing. Our analysis revealed that miRNAs are critical for the regulation of the PTEN-AKT-FOXO1 pathway that in turn controls Rag expression during B cell development., (Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.)

MAP kinases of the ERK family are conserved from yeast to humans. Their catalytic activity is dependent on dual phosphorylation of their activation loop's TEY motif, catalyzed by MAPK kinases (MEKs). Here we studied variants of Mpk1, a yeast orthologue of Erk, which is essential for cell wall integrity. Cells lacking MPK1, or the genes encoding the relevant MEKs, MKK1 and MKK2, do not proliferate under cell wall stress, imposed, for example, by caffeine. Mutants of Mpk1, Mpk1(Y268C) and Mpk1(Y268A), function independently of Mkk1 and Mkk2. We show that these variants are phosphorylated at their activation loop in mkk1∆mkk2∆ and mkk1∆mkk2∆pbs2∆ste7∆ cells, suggesting that they autophosphorylate. However, strikingly, when Y268C/A mutations were combined with the kinase-dead mutation, K54R, or mutations at the TEY motif, T190A+Y192F, the resulting proteins still allowed mkk1∆mkk2∆ cells to proliferate under caffeine stress. Mutating the equivalent residue, Tyr-280/Tyr-261, in Erk1/Erk2 significantly impaired Erk1/2's catalytic activity. This study describes the first case in which a MAPK, Erk/Mpk1, imposes a phenotype via a mechanism that is independent of TEY phosphorylation and an unusual case in which an equivalent mutation in a highly conserved domain of yeast and mammalian Erks causes an opposite effect., (© 2016 Goshen-Lago, Goldberg-Carp, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).)

PI3K activity determines positive and negative selection of B cells, a key process for immune tolerance and B cell maturation. Activation of PI3K is balanced by phosphatase and tensin homolog (Pten), the PI3K's main antagonistic phosphatase. Yet, the extent of feedback regulation between PI3K activity and Pten expression during B cell development is unclear. Here, we show that PI3K control of this process is achieved post-transcriptionally by an axis composed of a transcription factor (c-Myc), a microRNA (miR17-92), and Pten. Enhancing activation of this axis through overexpression of miR17-92 reconstitutes the impaired PI3K activity for positive selection in CD19-deficient B cells and restores most of the B cell developmental impairments that are evident in CD19-deficient mice. Using a genetic approach of deletion and complementation, we show that the c-Myc/miR17-92/Pten axis critically controls PI3K activity and the sensitivity of immature B cells to negative selection imposed by activation-induced cell death., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

With two experiments the authors test and find support for the argument that in small, collectively oriented task groups, status affects social influence the most when the distribution of opinions reduces the least uncertainty. Moreover, they demonstrate that people use the distribution of both status and opinions to reduce uncertainty about the task on which they are working and that this, in turn, promotes social influence. Experiment 1 illustrates that, regardless of the group's sex composition, basis for status differentiation, or size of the group, uncertainty reduction mediates a significant share of the effect of status and opinions on social influence. Experiment 2 confirms that the effect of the distribution of both status and opinions on social influence is weaker as the task becomes more certain. These findings inform discussion about how status affects certainty in task groups and what this potentially means for organizational settings and sociological theory more generally.

Many enzymes are self-regulated and can either inhibit or enhance their own catalytic activity. Enzymes that do both are extremely rare. Many protein kinases autoactivate by autophosphorylating specific sites at their activation loop and are inactivated by phosphatases. Although mitogen-activated protein kinases (MAPKs) are usually activated by dual phosphorylation catalyzed by MAPK kinases (MAPKKs), the MAPK p38β is exceptional and is capable of self-activation by cis autophosphorylation of its activation loop residue T180. We discovered that p38β also autophosphorylates in trans two previously unknown sites residing within a MAPK-specific structural element known as the MAPK insert: T241 and S261. Whereas phosphorylation of T180 evokes catalytic activity, phosphorylation of S261 reduces the activity of T180-phosphorylated p38β, and phosphorylation of T241 reduces its autophosphorylation in trans Both phosphorylations do not affect the activity of dually phosphorylated p38β. T241 of p38β is found phosphorylated in vivo in bone and muscle tissues. In myogenic cell lines, phosphorylation of p38β residue T241 is correlated with differentiation to myotubes. T241 and S261 are also autophosphorylated in intrinsically active variants of p38α, but in this protein, they probably play a different role. We conclude that p38β is an unusual enzyme that automodulates its basal, MAPKK-independent activity by several autophosphorylation events, which enhance and suppress its catalytic activity., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

Cellular homeostasis in the B cell compartment is strictly imposed to balance cell production and cell loss. However, it is not clear whether B cell development in the bone marrow is an autonomous process or subjected to regulation by the peripheral B cell compartment. To specifically address this question, we used mice transgenic for human CD20, where effective depletion of B lineage cells is obtained upon administration of mouse anti-human CD20 antibodies, in the absence of any effect on other cell lineages and/or tissues. We followed the kinetics of B cell return to equilibrium by BrdU labeling and flow cytometry and analyzed the resulting data by mathematical modeling. Labeling was much faster in depleted mice. Compared to control mice, B cell-depleted mice exhibited a higher proliferation rate in the pro-/pre-B compartment, and higher cell death and lower differentiation in the immature B cell compartment. We validated the first result by analysis of the expression of Ki67, the nuclear protein expressed in proliferating cells, and the second using Annexin V staining. Collectively, our results suggest that B lymphopoiesis is subjected to homeostatic feedback mechanisms imposed by mature B cells in the peripheral compartment.

The receptor-tyrosine kinase (RTK)/Ras/Raf pathway is an essential cascade for mediating growth factor signaling. It is abnormally overactive in almost all human cancers. The downstream targets of the pathway are members of the extracellular regulated kinases (Erk1/2) family, suggesting that this family is a mediator of the oncogenic capability of the cascade. Although all oncogenic mutations in the pathway result in strong activation of Erks, activating mutations in Erks themselves were not reported in cancers. Here we used spontaneously active Erk variants to check whether Erk's activity per se is sufficient for oncogenic transformation. We show that Erk1(R84S) is an oncoprotein, as NIH3T3 cells that express it form foci in tissue culture plates, colonies in soft agar, and tumors in nude mice. We further show that Erk1(R84S) and Erk2(R65S) are intrinsically active due to an unusual autophosphorylation activity they acquire. They autophosphorylate the activatory TEY motif and also other residues, including the critical residue Thr-207 (in Erk1)/Thr-188 (in Erk2). Strikingly, Erk2(R65S) efficiently autophosphorylates its Thr-188 even when dually mutated in the TEY motif. Thus this study shows that Erk1 can be considered a proto-oncogene and that Erk molecules possess unusual autoregulatory properties, some of them independent of TEY phosphorylation., (© 2016 Smorodinsky-Atias, Goshen-Lago, et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).)

The elderly immune system is characterized by reduced responses to infections and vaccines, and an increase in the incidence of autoimmune diseases and cancer. Age-related deficits in the immune system may be caused by peripheral homeostatic pressures that limit bone marrow B-cell production or migration to the peripheral lymphoid tissues. Studies of peripheral blood B-cell receptor spectratypes have shown that those of the elderly are characterized by reduced diversity, which is correlated with poor health status. In the present study, we performed for the first time high-throughput sequencing of immunoglobulin genes from archived biopsy samples of primary and secondary lymphoid tissues in old (74 ± 7 years old, range 61-89) versus young (24 ± 5 years old, range 18-45) individuals, analyzed repertoire diversities and compared these to results in peripheral blood. We found reduced repertoire diversity in peripheral blood and lymph node repertoires from old people, while in the old spleen samples the diversity was larger than in the young. There were no differences in somatic hypermutation characteristics between age groups. These results support the hypothesis that age-related immune frailty stems from altered B-cell homeostasis leading to narrower memory B-cell repertoires, rather than changes in somatic hypermutation mechanisms., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Recent findings indicate that the ubiquitin-proteasome system is involved in the pathogenesis of cancer as well as autoimmune and several neurodegenerative diseases, and is thus a target for novel therapeutics. One disease that is related to aberrant protein degradation is multiple sclerosis, an autoimmune disorder involving the processing and presentation of myelin autoantigens that leads to the destruction of axons. Here, we show that brain-derived proteasomes from SJL mice with experimental autoimmune encephalomyelitis (EAE) in an ubiquitin-independent manner generate significantly increased amounts of myelin basic protein peptides that induces cytotoxic lymphocytes to target mature oligodendrocytes ex vivo. Ten times enhanced release of immunogenic peptides by cerebral proteasomes from EAE-SJL mice is caused by a dramatic shift in the balance between constitutive and β1i(high) immunoproteasomes in the CNS of SJL mice with EAE. We found that during EAE, β1i is increased in resident CNS cells, whereas β5i is imported by infiltrating lymphocytes through the blood-brain barrier. Peptidyl epoxyketone specifically inhibits brain-derived β1i(high) immunoproteasomes in vitro (kobs/[I] = 240 M(-1)s(-1)), and at a dose of 0.5 mg/kg, it ameliorates ongoing EAE in vivo. Therefore, our findings provide novel insights into myelin metabolism in pathophysiologic conditions and reveal that the β1i subunit of the immunoproteasome is a potential target to treat autoimmune neurologic diseases., (© The Author(s).)

Many protein interactions are conserved among organisms despite changes in the amino acid sequences that comprise their contact sites, a property that has been used to infer the location of these sites from protein homology. In an inter-species complementation experiment, a sequence present in a homologue is substituted into a protein and tested for its ability to support function. Therefore, substitutions that inhibit function can identify interaction sites that changed over evolution. However, most of the sequence differences within a protein family remain unexplored because of the small-scale nature of these complementation approaches. Here we use existing high throughput mutational data on the in vivo function of the RRM2 domain of the Saccharomyces cerevisiae poly(A)-binding protein, Pab1, to analyze its sites of interaction. Of 197 single amino acid differences in 52 Pab1 homologues, 17 reduce the function of Pab1 when substituted into the yeast protein. The majority of these deleterious mutations interfere with the binding of the RRM2 domain to eIF4G1 and eIF4G2, isoforms of a translation initiation factor. A large-scale mutational analysis of the RRM2 domain in a two-hybrid assay for eIF4G1 binding supports these findings and identifies peripheral residues that make a smaller contribution to eIF4G1 binding. Three single amino acid substitutions in yeast Pab1 corresponding to residues from the human orthologue are deleterious and eliminate binding to the yeast eIF4G isoforms. We create a triple mutant that carries these substitutions and other humanizing substitutions that collectively support a switch in binding specificity of RRM2 from the yeast eIF4G1 to its human orthologue. Finally, we map other deleterious substitutions in Pab1 to inter-domain (RRM2-RRM1) or protein-RNA (RRM2-poly(A)) interaction sites. Thus, the combined approach of large-scale mutational data and evolutionary conservation can be used to characterize interaction sites at single amino acid resolution.

Cells respond to changes in their environment through mechanisms that often necessitate reprogramming of the translation machinery. The fastest and strongest of all tested responses is the translation inhibition observed following abrupt depletion of glucose from the media of yeast cells. The speed of the response suggests a post-translational modification of a key component of the translation machinery. This translation factor is as yet unknown. A cAMP-dependent protein kinase mutant yeast strain (tpk1(w)) that does not respond properly to glucose depletion and maintains translation was described previously. We hypothesized that the inability of tpk1(w) to arrest translation results from abnormal expression of key translation mediators. Genome-wide analysis of steady-state mRNA levels in tpk1(w) revealed underexpression of several candidates. Elevating the cellular levels of eukaryotic initiation factor (eIF) 5 by overexpression rescued the translational defect of tpk1(w). Restoring ribosomal dissociation by eIF5 necessitated an active GAP domain and multiple regions throughout this protein. Phosphoproteomics analysis of wild-type cells overexpressing eIF5 revealed increased phosphorylation in a novel site (Thr191) upon glucose depletion. Mutating this residue and introducing it into tpk1(w) abolished the ability of eIF5 to rescue the translational defect. Intriguingly, introducing this mutation into the wild-type strain did not hamper its translational response. We further show that Thr191 is phosphorylated in vitro by Casein Kinase II (CKII), and yeast cells with a mutated CKII have a reduced response to glucose depletion. These results implicate phosphorylation of eIF5 at Thr191 by CKII as one of the pathways for regulating translation upon glucose depletion., (© 2014 FEBS.)

Protein kinases are regulated by a large number of mechanisms that vary from one kinase to another. However, a fundamental activation mechanism shared by all protein kinases is phosphorylation of a conserved activation loop threonine residue. This is achieved in many cases via autophosphorylation. The mechanism and structural basis for autophosphorylation are not clear and are in fact enigmatic because this phosphorylation occurs when the kinase is in its inactive conformation. Unlike most protein kinases, MAP kinases are not commonly activated by autophosphorylation but rather by MEK-dependent phosphorylation. Here we show that p38β, a p38 isoform that is almost identical to p38α, is exceptional and spontaneously autoactivates by autophosphorylation. We identified a 13-residue-long region composed of part of the αG-helix and the MAPK insert that triggers the intrinsic autophosphorylation activity of p38β. When inserted into p38α, this fragment renders it spontaneously active in vitro and in mammalian cells. We further found that an interaction between the N terminus and a particular region of the C-terminal extension suppresses the intrinsic autophosphorylation of p38β in mammalian cells. Thus, this study identified the structural motif responsible for the unique autophosphorylation capability of p38β and the motif inhibiting this activity in living cells. It shows that the MAPK insert and C-terminal extension, structural motifs that are unique to MAPKs, play a critical role in controlling autophosphorylation., (© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.)

Identifying communities or clusters in networked systems has received much attention across the physical and social sciences. Most of this work focuses on single layer or one-mode networks, including social networks between people or hyperlinks between websites. Multilayer or multi-mode networks, such as affiliation networks linking people to organizations, receive much less attention in this literature. Common strategies for discovering the community structure of multi-mode networks identify the communities of each mode simultaneously. Here I show that this combined approach is ineffective at discovering community structures when there are an unequal number of communities between the modes of a multi-mode network. I propose a dual-projection alternative for detecting communities in multi-mode networks that overcomes this shortcoming. The evaluation of synthetic networks with known community structures reveals that the dual-projection approach outperforms the combined approach when there are a different number of communities in the various modes. At the same time, results show that the dual-projection approach is as effective as the combined strategy when the number of communities is the same between the modes.

The RNA recognition motif (RRM) is the most common RNA-binding domain in eukaryotes. Differences in RRM sequences dictate, in part, both RNA and protein-binding specificities and affinities. We used a deep mutational scanning approach to study the sequence-function relationship of the RRM2 domain of the Saccharomyces cerevisiae poly(A)-binding protein (Pab1). By scoring the activity of more than 100,000 unique Pab1 variants, including 1246 with single amino acid substitutions, we delineated the mutational constraints on each residue. Clustering of residues with similar mutational patterns reveals three major classes, composed principally of RNA-binding residues, of hydrophobic core residues, and of the remaining residues. The first class also includes a highly conserved residue not involved in RNA binding, G150, which can be mutated to destabilize Pab1. A comparison of the mutational sensitivity of yeast Pab1 residues to their evolutionary conservation reveals that most residues tolerate more substitutions than are present in the natural sequences, although other residues that tolerate fewer substitutions may point to specialized functions in yeast. An analysis of ∼40,000 double mutants indicates a preference for a short distance between two mutations that display an epistatic interaction. As examples of interactions, the mutations N139T, N139S, and I157L suppress other mutations that interfere with RNA binding and protein stability. Overall, this study demonstrates that living cells can be subjected to a single assay to analyze hundreds of thousands of protein variants in parallel.