FKHR-L1 can act as a critical effector of cell death induced by cytokine withdrawal: protein kinase B–enhanced cell survival through maintenance of mitochondrial integrity

Survival signals elicited by cytokines include the activation of phosphatidylinositol 3-kinase (PI3K), which in turn promotes the activation of protein kinase B (PKB). Recently, PKB has been demonstrated to phosphorylate and inactivate forkhead transcription factor FKHR-L1, a potent inducer of apoptosis. To explore the mechanisms underlying the induction of apoptosis after cytokine withdrawal or FKHR-L1 activation, we used a cell line in which FKHR-L1 activity could be specifically induced. Both cytokine withdrawal and FKHR-L1 activation induced apoptosis, which was preceded by an upregulation in p27KIP1 and a concomitant decrease in cells entering the cell cycle. Induction of apoptosis by both cytokine withdrawal and activation of FKHR-L1 correlated with the disruption of mitochondrial membrane integrity and cytochrome c release. This was preceded by upregulation of the pro-apoptotic Bcl-2 family member Bim. Ectopic expression of an inhibitory mutant of FKHR-L1 substantially reduced the levels of apoptosis observed after cytokine withdrawal. Activation of PKB alone was sufficient to promote cell survival, as measured by maintenance of mitochondrial integrity and the resultant inhibition of effector caspases. Furthermore, hematopoietic stem cells isolated from Bim−/− mice exhibited reduced levels of apoptosis upon inhibition of PI3K/PKB signaling

Structural and Biophysical Insights into SPINK1 Bound to Human Cationic Trypsin

(1) The serine protease inhibitor Kazal type 1 (SPINK1) inhibits trypsin activity in zymogen granules of pancreatic acinar cells. Several mutations in the SPINK1 gene are associated with acute recurrent pancreatitis (ARP) and chronic pancreatitis (CP). The most common variant is SPINK1 p.N34S. Although this mutation was identified two decades ago, the mechanism of action has remained elusive. (2) SPINK1 and human cationic trypsin (TRY1) were expressed in E. coli, and inhibitory activities were determined. Crystals of SPINK1–TRY1 complexes were grown by using the hanging-drop method, and phases were solved by molecular replacement. (3) Both SPINK1 variants show similar inhibitory behavior toward TRY1. The crystal structures are almost identical, with minor differences in the mutated loop. Both complexes show an unexpected rotamer conformation of the His63 residue in TRY1, which is a member of the catalytic triad. (4) The SPINK1 p.N34S mutation does not affect the inhibitory behavior or the overall structure of the protein. Therefore, the pathophysiological mechanism of action of the p.N34S variant cannot be explained mechanistically or structurally at the protein level. The observed histidine conformation is part of a mechanism for SPINK1 that can explain the exceptional proteolytic stability of this inhibitor

Establishing cordgrass plants cluster their shoots to avoid ecosystem engineering

Vegetated coastal ecosystems such as salt marshes, dunes and seagrass meadows occur at the land–sea interface—a dynamic environment typified by harsh growing conditions. These ecosystems are known as biogeomorphic landscapes because their functioning depends on biophysical interactions by which organisms engineer landforms to their own benefit. The strength of such biogeomorphic feedbacks depends on plant traits, such as stem flexibility and shoot density. Recent work demonstrated that dune grasses with similar morphological traits can build contrasting landscapes due to differences in their spatial shoot organization. However, in contrast to dune grasses that trap and stabilize sand particles in aeolian landscapes, flow attenuation in aquatic environments can generate scouring around plant stems and cause uprooting, leading to establishment thresholds for young plants. Yet, it remains unknown how findings from aeolian landscapes translate to aquatic systems and how young clonally expanding plants in hydrodynamically exposed conditions overcome these establishment thresholds by optimizing shoot placement. Here, we measured shoot patterns of 90 establishing cordgrass patches Spartina anglica at 18 European field sites that cover a broad range of hydrodynamic conditions. Next, we carried out a field experiment to investigate how observed spatial shoot patterns affect plant–sediment feedbacks. Surprisingly, field survey analyses reveal highly consistent clustered shoot patterns, regardless of environmental conditions. Experimental results demonstrate that this clustered pattern minimizes scouring compared to densely clumped organizations typically observed in established patches. Synthesis. In contrast to earlier findings highlighting that establishing dune grasses optimize their landscape engineering capacity via a flexible shoot placement strategy, we find that cordgrass instead follows a fixed strategy that minimizes engineering effects in its early life stages. We suggest that marsh grasses avoid physical stress and associated establishment thresholds in their early life stage, and switch to an ecosystem engineering strategy once established. These findings shed new light on how plant traits interact with their environment to shape the landscape and pave the way for improved restoration designs by mimicking the natural shoot organization of establishing vegetation. Read the free Plain Language Summary for this article on the Journal blog.</p

Coding by Choice: A Transitional Analysis of Social Participation Patterns and Programming Contributions in the Online Scratch Community

While massive online communities have drawn the attention of researchers and educators on their potential to support active collaborative work, knowledge sharing, and user-generated content, few studies examine participation in these communities at scale. The little research that does exist attends almost solely to adults rather than communities to support youths’ learning and identity development. In this chapter, we tackle two challenges related to understanding social practices that support learning in massive social networking forums where users engage in design. We examined a youth programmer community, called Scratch.mit.edu, that garners the voluntary participation of millions of young people worldwide. We report on site-wide distributions and patterns of participation that illuminate the relevance of different online social practices to ongoing involvement in the online community. Drawing on a random sample of more than 5000 active users of Scratch.mit.edu over a 3-month time period in early 2012, we examine log files that captured the frequency of three types of social practices that contribute to enduring participation: DIY participatory activities, socially supportive actions, and socially engaging interactions. Using latent transition analysis, we found (1) distinct patterns of participation (classes) across three time points (e.g., high networkers who are generally active, commenters who focus mainly on social participation, downloaders engaging in DIY participatory activities), (2) unique migration changes in class membership across time, (3) relatively equal gender representation across these classes, and (4) importance of membership length (or age) in terms of class memberships. In the discussion, we review our approach to analysis and outline implications for the design and study of online communities and tools for youth

Restoration of impaired intestinal barrier function by the hydrolysed casein diet contributes to the prevention of type 1 diabetes in the diabetes-prone BioBreeding rat

Aims/hypothesis Impaired intestinal barrier function is observed in type I diabetes patients and animal models of the disease. Exposure to diabetogenic antigens from the intestinal milieu due to a compromised intestinal barrier is considered essential for induction of the autoimmune process leading to type I diabetes. Since a hydrolysed casein (HC) diet prevents autoimmune diabetes onset in diabetes-prone (DP)-BioBreeding (BB) rats, we studied the role of the HC diet on intestinal barrier function and, therefore, prevention of autoimmune diabetes onset in this animal model. Methods DP-BB rats were fed the HC diet from weaning onwards and monitored for autoimmune diabetes development. Intestinal permeability was assessed in vivo by lactulose mannitol test and ex vivo by measuring trans-epithelial electrical resistance (TEER). Levels of serum zonulin, a physiological tight junction modulator, were measured by ELISA. heal mRNA expression of Myo9b, Cldn1, Cldn2 and Ocln (which encode the tight junction-related proteins myosin IXb, claudin-1, claudin-2 and occludin) and Il-10, Tgf-beta (also known as Il10 and Tgfb, respectively, which encode regulatory cytokines) was analysed by quantitative PCR. Results The HC diet reduced autoimmune diabetes by 50% in DP-BB rats. In DP-BB rats, prediabetic gut permeability negatively correlated with the moment of autoimmune diabetes onset. The improved intestinal barrier function that was induced by HC diet in DP-BB rats was visualised by decreasing lactulose:mannitol ratio, decreasing serum zonulin levels and increasing ileal TEER. The HC diet modified ileal mRNA expression of Myo9b, and Cldn1 and Cldn2, but left Ocln expression unaltered. Conclusions/interpretation Improved intestinal barrier function might be an important intermediate in the prevention of autoimmune diabetes by the HC diet in DP-BB rats. Effects on tight junctions, ileal cytokines and zonulin production might be important mechanisms for this effect

Gaia Early Data Release 3:Modelling and calibration of Gaia's point and line spread functions

Context: The unprecedented astrometric precision of the Gaia mission relies on accurate estimates of the locations of sources in the Gaia data stream. This is ultimately performed by point spread function (PSF) fitting, which in turn requires an accurate reconstruction of the PSF. Gaia Early Data Release 3 (EDR3) will, for the first time, use a PSF calibration that models several of the strongest dependences, leading to signficantly reduced systematic errors. Aims: We describe the PSF model and calibration pipeline implemented for Gaia EDR3, including an analysis of the calibration results over the 34 months of data. We include a discussion of the limitations of the current pipeline and directions for future releases. This will be of use both to users of Gaia data and as a reference for other precision astrometry missions. Methods: We develop models of the 1D line spread function (LSF) and 2D PSF profiles based on a linear combination of basis components. We fit the models to selected primary sources in independent time ranges, using simple parameterisations for the colour and other dependences. Variation in time is smoothed by merging the independent calibrations in a square root information filter, with resets at certain mission events that induce a discontinuous change in the PSF. Results: The PSF calibration shows strong time and colour dependences that accurately reproduce the varying state of the Gaia astrometric instrument. Analysis of the residuals reveals both the performance and the limitations of the current models and calibration pipeline, and indicates the directions for future development. Conclusions: The PSF modelling and calibration carried out for Gaia EDR3 represents a major step forwards in the data processing and will lead to reduced systematic errors in the core mission data products. Further significant improvements are expected in the future data releases.Comment: Accepted by A&A for inclusion in Gaia EDR3 special issu