The coordination of cell growth during fission yeast mating requires Ras1-GTP hydrolysis

The spatial and temporal control of polarity is fundamental to the survival of all organisms. Cells define their polarity using highly conserved mechanisms that frequently rely upon the action of small GTPases, such as Ras and Cdc42. Schizosaccharomyces pombe is an ideal system with which to study the control of cell polarity since it grows from defined tips using Cdc42-mediated actin remodeling. Here we have investigated the importance of Ras1-GTPase activity for the coordination of polarized cell growth during fission yeast mating. Following pheromone stimulation, Ras1 regulates both a MAPK cascade and the activity of Cdc42 to enable uni-directional cell growth towards a potential mating partner. Like all GTPases, when bound to GTP, Ras1 adopts an active conformation returning to an inactive state upon GTP-hydrolysis, a process accelerated through interaction with negative regulators such as GAPs. Here we show that, at low levels of pheromone stimulation, loss of negative regulation of Ras1 increases signal transduction via the MAPK cascade. However, at the higher concentrations observed during mating, hyperactive Ras1 mutations promote cell death. We demonstrate that these cells die due to their failure to coordinate active Cdc42 into a single growth zone resulting in disorganized actin deposition and unsustainable elongation from multiple tips. These results provide a striking demonstration that the deactivation stage of Ras signaling is fundamentally important in modulating cell polarity

How much do disasters cost? A comparison of disaster cost estimates in Australia

Extreme weather events in Australia are common and a large proportion of the population are exposed to such events. Therefore, there is great interest as to how these events impact Australia's society and economy, which requires understanding the current and historical impact of disasters. Despite global efforts to record and cost disaster impacts, no standardised method of collecting and recording data retrospectively yet exists. The lack of standardisation in turn results in a range of different estimates of economic impacts. This paper examines five examples of aggregate disaster loss and impacts of natural disasters in Australia, and comparisons between them reveal significant data shortcomings. The reliability of data sources, and the methodology employed to analyse them can have significant impacts on conclusions regarding the overall cost of disasters, the relative costs of different hazards (disaster types), and the distribution of losses across Australian states. We highlight difficulties with time series comparisons, further complicated by the interdependencies of the databases. We reiterate the need for consistent and comparable data collection and analysis, to respond to the increasing frequency and severity of disasters in Australia

The role of the RACK1 ortholog Cpc2p in modulating pheromone-induced cell cycle arrest in fission yeast

The detection and amplification of extracellular signals requires the involvement of multiple protein components. In mammalian cells the receptor of activated C kinase (RACK1) is an important scaffolding protein for signal transduction networks. Further, it also performs a critical function in regulating the cell cycle by modulating the G1/S transition. Many eukaryotic cells express RACK1 orthologs, with one example being Cpc2p in the fission yeast Schizosaccharomyces pombe. In contrast to RACK1, Cpc2p has been described to positively regulate, at the ribosomal level, cells entry into M phase. In addition, Cpc2p controls the stress response pathways through an interaction with Msa2p, and sexual development by modulating Ran1p/Pat1p. Here we describe investigations into the role, which Cpc2p performs in controlling the G protein-mediated mating response pathway. Despite structural similarity to Gβ-like subunits, Cpc2p appears not to function at the G protein level. However, upon pheromone stimulation, cells overexpressing Cpc2p display substantial cell morphology defects, disorientation of septum formation and a significantly protracted G1 arrest. Cpc2p has the potential to function at multiple positions within the pheromone response pathway. We provide a mechanistic interpretation of this novel data by linking Cpc2p function, during the mating response, with its previous described interactions with Ran1p/Pat1p. We suggest that overexpressing Cpc2p prolongs the stimulated state of pheromone-induced cells by increasing ste11 gene expression. These data indicate that Cpc2p regulates the pheromone-induced cell cycle arrest in fission yeast by delaying cells entry into S phase

Remote measurement of surface temperature distributions in laser heated samples

SIGLEAvailable from British Library Document Supply Centre- DSC:DX179865 / BLDSC - British Library Document Supply CentreGBUnited Kingdo

Dual positive and negative regulation of GPCR signaling by GTP hydrolysis

G protein-coupled receptors (GPCRs) regulate a variety of intracellular pathways through their ability to promote the binding of GTP to heterotrimeric G proteins. Regulator of G protein signaling (RGS) proteins increase the intrinsic GTPase activity of G-subunits and are widely regarded as negative regulators of G protein signaling. Using yeast we demonstrate that GTP hydrolysis is not only required for desensitization, but is essential for achieving a high maximal (saturated level) response. Thus RGS-mediated GTP hydrolysis acts as both a negative (low stimulation) and positive (high stimulation) regulator of signaling. To account for this we generated a new kinetic model of the G protein cycle where GGTP enters an inactive GTP-bound state following effector activation. Furthermore, in vivo and in silico experimentation demonstrates that maximum signaling output first increases and then decreases with RGS concentration. This unimodal, non-monotone dependence on RGS concentration is novel. Analysis of the kinetic model has revealed a dynamic network motif that shows precisely how inclusion of the inactive GTP-bound state for the G produces this unimodal relationship

Folding-competent and folding-defective forms of Ricin A chain have different fates following retrotranslocation from the endoplasmic reticulum

We report that a toxic polypeptide retaining the potential to refold upon dislocation from the endoplasmic reticulum (ER) to the cytosol (ricin A chain; RTA) and a misfolded version that cannot (termed RTAΔ), follow ER-associated degradation (ERAD) pathways in Saccharomyces cerevisiae that substantially diverge in the cytosol. Both polypeptides are dislocated in a step mediated by the transmembrane Hrd1p ubiquitin ligase complex and subsequently degraded. Canonical polyubiquitylation is not a prerequisite for this interaction because a catalytically inactive Hrd1p E3 ubiquitin ligase retains the ability to retrotranslocate RTA, and variants lacking one or both endogenous lysyl residues also require the Hrd1p complex. In the case of native RTA, we established that dislocation also depends on other components of the classical ERAD-L pathway as well as an ongoing ER–Golgi transport. However, the dislocation pathways deviate strikingly upon entry into the cytosol. Here, the CDC48 complex is required only for RTAΔ, although the involvement of individual ATPases (Rpt proteins) in the 19S regulatory particle (RP) of the proteasome, and the 20S catalytic chamber itself, is very different for the two RTA variants. We conclude that cytosolic ERAD components, particularly the proteasome RP, can discriminate between structural features of the same substrate

Government liabilities for disaster risk in industrialized countries: a case study of Australia

This paper explores sovereign risk preferences against direct and indirect natural disasters losses in industrialized countries. Using Australia as a case study, the analysis compares expected disaster losses and government capacity to finance losses. Utilizing a national disaster loss dataset, extreme value theory is applied to estimate an all-hazard annual loss distribution. Unusually but critically, the dataset includes direct as well as indirect losses, allowing for the analysis to consider the oft-ignored issue of indirect losses. Expected annual losses (direct, and direct plus indirect) are overlaid with a risk-layer approach, to distinguish low, medium and extreme loss events. Each risk layer is compared to available fiscal resources for financing losses, grounded in the political reality of Australian disaster financing. When considering direct losses only, we find support for a risk-neutral preference on the part of the Australian government for low and medium loss levels, and a risk-averse preference in regard to extreme losses. When indirect losses are also estimated, we find that even medium loss levels are expected to overwhelm available fiscal resources, thereby violating the available resources assumption underlying arguments for sovereign risk neutrality. Our analysis provides empirical support for the assertion that indirect losses are a major, under-recognised concern for industrialized countries. A risk-averse preference in regard to medium and extreme loss events recommends enhanced investment in both corrective and prospective risk reduction in relation to these risks level, in particular to reduce indirect losses

Feedback activation of neurofibromin terminates growth factor-induced Ras activation.

BACKGROUND: Growth factors induce a characteristically short-lived Ras activation in cells emerging from quiescence. Extensive work has shown that transient as opposed to sustained Ras activation is critical for the induction of mitogenic programs. Mitogen-induced accumulation of active Ras-GTP results from increased nucleotide exchange driven by the nucleotide exchange factor Sos. In contrast, the mechanism accounting for signal termination and prompt restoration of basal Ras-GTP levels is unclear, but has been inferred to involve feedback inhibition of Sos. Remarkably, how GTP-hydrolase activating proteins (GAPs) participate in controlling the rise and fall of Ras-GTP levels is unknown. RESULTS: Monitoring nucleotide exchange of Ras in permeabilized cells we find, unexpectedly, that the decline of growth factor-induced Ras-GTP levels proceeds in the presence of unabated high nucleotide exchange, pointing to GAP activation as a major mechanism of signal termination. Experiments with non-hydrolysable GTP analogues and mathematical modeling confirmed and rationalized the presence of high GAP activity as Ras-GTP levels decline in a background of high nucleotide exchange. Using pharmacological and genetic approaches we document a raised activity of the neurofibromatosis type I tumor suppressor Ras-GAP neurofibromin and an involvement of Rsk1 and Rsk2 in the down-regulation of Ras-GTP levels. CONCLUSIONS: Our findings show that, in addition to feedback inhibition of Sos, feedback stimulation of the RasGAP neurofibromin enforces termination of the Ras signal in the context of growth-factor signaling. These findings ascribe a precise role to neurofibromin in growth factor-dependent control of Ras activity and illustrate how, by engaging Ras-GAP activity, mitogen-challenged cells play safe to ensure a timely termination of the Ras signal irrespectively of the reigning rate of nucleotide exchange.We acknowledge funding by the German research council (DFG), grant # RU 860/4-1 (AH), by the Federal Ministry of Education and Research (BMBF), Germany, FKZ: 01EO1002 (I.R., R.M.), by the BBSRC and through the BBSRC Midlands Interdisciplinary BioSciences Training Partnership (MAE-F) (GL - BB/G01227X/1 and BB/M00015X/1) and the National Council on Science and Technology of Mexico (CONACYT) (MAE-F).This is the final published version. It first appeared at http://biosignaling.biomedcentral.com/articles/10.1186/s12964-016-0128-z

Creating functional groups of marine fish from categorical traits

Background Functional groups serve two important functions in ecology: they allow for simplification of ecosystem models and can aid in understanding diversity. Despite their important applications, there has not been a universally accepted method of how to define them. A common approach is to cluster species on a set of traits, validated through visual confirmation of resulting groups based primarily on expert opinion. The goal of this research is to determine a suitable procedure for creating and evaluating functional groups that arise from clustering nominal traits. Methods To do so, we produced a species by trait matrix of 22 traits from 116 fish species from Tasman Bay and Golden Bay, New Zealand. Data collected from photographs and published literature were predominantly nominal, and a small number of continuous traits were discretized. Some data were missing, so the benefit of imputing data was assessed using four approaches on data with known missing values. Hierarchical clustering is utilised to search for underlying data structure in the data that may represent functional groups. Within this clustering paradigm there are a number of distance matrices and linkage methods available, several combinations of which we test. The resulting clusters are evaluated using internal metrics developed specifically for nominal clustering. This revealed the choice of number of clusters, distance matrix and linkage method greatly affected the overall within- and between- cluster variability. We visualise the clustering in two dimensions and the stability of clusters is assessed through bootstrapping. Results Missing data imputation showed up to 90% accuracy using polytomous imputation, so was used to impute the real missing data. A division of the species information into three functional groups was the most separated, compact and stable result. Increasing the number of clusters increased the inconsistency of group membership, and selection of the appropriate distance matrix and linkage method improved the fit. Discussion We show that the commonly used methodologies used for the creation of functional groups are fraught with subjectivity, ultimately causing significant variation in the composition of resulting groups. Depending on the research goal dictates the appropriate strategy for selecting number of groups, distance matrix and clustering algorithm combination

Deciphering the Agonist Binding Mechanism to the Adenosine A1 Receptor.

Despite being among the most characterized G protein-coupled receptors (GPCRs), adenosine receptors (ARs) have always been a difficult target in drug design. To date, no agonist other than the natural effector and the diagnostic regadenoson has been approved for human use. Recently, the structure of the adenosine A1 receptor (A1R) was determined in the active, Gi protein complexed state; this has important repercussions for structure-based drug design. Here, we employed supervised molecular dynamics simulations and mutagenesis experiments to extend the structural knowledge of the binding of selective agonists to A1R. Our results identify new residues involved in the association and dissociation pathway, they suggest the binding mode of N6-cyclopentyladenosine (CPA) related ligands, and they highlight the dramatic effect that chemical modifications can have on the overall binding mechanism, paving the way for the rational development of a structure-kinetics relationship of A1R agonists.Leverhulme Trus