Comparison of the most demanding scenarios during different in-season training sessions and official matches in professional basketball players

The purpose of this study was to compare physical demands during the most demanding scenarios (MDS) of different training sessions and official matches in professional basketball players across playing positions. Thirteen professional basketball players were monitored over a 9-week competitive season using a local positioning system. Peak physical demands included total distance, distance covered at > 18 km·h^-1, distance and number of accelerations (≥ 2 m∙s^-2) and decelerations (≤ -2 m∙s^-2) over a 60-second epoch. Analysis of variance for repeated measures, Bonferroni post-hoc tests and standardised Cohen’s effect size (ES) were calculated. Overall, almost all physical demands during the MDS of training were lower (-6.2% to -35.4%) compared to official matches. The only variable that surpassed competition demands was distance covered at > 18 km·h^-1, which presented moderate (ES = 0.61, p = 0.01) and small (ES = 0.48, p > 0.05) increases during training sessions four and three days before a competition, respectively. Conversely, the two previous practices before match day presented trivial to very large decreases (ES = 0.09–2.66) in all physical demands. Furthermore, centres achieved the lowest peak value in total distance covered during matches, forwards completed the greatest peak distance at > 18 km·h^-1, and guards performed the greatest distance and number of high-intensity accelerations and decelerations. In conclusion, physical demands during the MDS of different training sessions across the microcycle failed to match or surpass peak values during official matches, which should be considered when prescribing a training process intended to optimise the MDS of match play.     

Inducible recruitment of Cdc42 or WASP to a cell-surface receptor triggers actin polymerization and filopodium formation

BACKGROUND: Cdc42, a GTP-binding protein of the Rho family, controls actin cytoskeletal organization and helps to generate actin-based protruding structures, such as filopodia. In vitro, Cdc42 regulates actin polymerization by facilitating the creation of free barbed ends - the more rapidly growing ends of actin filaments - and subsequent elongation at these ends. The Wiskott- Aldrich syndrome protein, WASP, which has a pleckstrin-homology domain and a Cdc42/Rac-binding motif, has been implicated in cell signaling and cytoskeleton reorganization. We have investigated the consequences of local recruitment of activated Cdc42 or WASP to the plasma membrane. RESULTS: We used an activated Cdc42 protein that could be recruited to an engineered membrane receptor by adding rapamycin as a bridge, and added antibody-coupled beads to aggregate these receptors. Inducible recruitment of Cdc42 to clusters of receptors stimulated actin polymerization, resulting in the formation of membrane protrusions. Cdc42-induced protrusions were enriched in the vasodilator-stimulated phosphoprotein VASP and the focal-adhesion-associated proteins zyxin and ezrin. The Cdc42 effector WASP could also induce the formation of protrusions, albeit of different morphology. CONCLUSIONS: This is the first demonstration that the local recruitment of activated Cdc42 or its downstream effector, WASP, to a membrane receptor in whole cells is sufficient to trigger actin polymerization that results in the formation of membrane protrusions. Our data suggest that Cdc42-induced actin-based protrusions result from the local and serial recruitment of cytoskeletal proteins including zyxin, VASP, and ezrin.     

Elucidation of a PTS-carbohydrate chemotactic signal pathway in Escherichia coli using a time-resolved behavioral assay

Chemotaxis of Escherichia coli toward phosphotransferase systems (PTSs)-carbohydrates requires phosphoenolpyruvate-dependent PTSs as well as the chemotaxis response regulator CheY and its kinase, CheA. Responses initiated by flash photorelease of a PTS substrates D-glucose and its nonmetabolizable analog methyl alpha-D-glucopyranoside were measured with 33-ms time resolution using computer-assisted motion analysis. This, together with chemotactic mutants, has allowed us to map out and characterize the PTS chemotactic signal pathway. The responses were absent in mutants lacking the general PTS enzymes EI or HPr, elevated in PTS transport mutants, retarded in mutants lacking CheZ, a catalyst of CheY autodephosphorylation, and severely reduced in mutants with impaired methyl-accepting chemotaxis protein (MCP) signaling activity. Response kinetics were comparable to those triggered by MCP attractant ligands over most of the response range, the most rapid being 11.7 +/- 3.1 s-1. The response threshold was <10 nM for glucose. Responses to methyl alpha-D-glucopyranoside had a higher threshold, commensurate with a lower PTS affinity, but were otherwise kinetically indistinguishable. These facts provide evidence for a single pathway in which the PTS chemotactic signal is relayed rapidly to MCP-CheW-CheA signaling complexes that effect subsequent amplification and slower CheY dephosphorylation. The high sensitivity indicates that this signal is generated by transport-induced dephosphorylation of the PTS rather than phosphoenolpyruvate consumption.     

Conformational study of proteins by SAXS and EDXD: the case of trypsin and trypsinogen

The radius of gyration (Rg) of bovine trypsinogen and beta-trypsin was measured by an energy-dispersive X-ray technique (EDXD) and by small-angle X-ray scattering (SAXS), under different solvent conditions. Both techniques gave superimposable results. The experimental evidence demonstrated that: (1) no structural modifications and/or damage occurred during the data acquisition by EDXD; (2) at pH 4 the active enzyme has one class of chloride binding sites in common with the zymogen, whereas the latter protease shows an additional class able to reverse the effects on Rg induced by chloride at low concentration; and (3) the pH profile of the Rg of both proteases does not resemble at all the pH effect on beta-trypsin activity, a result in line with the finding that the electrical potentials induced by surface charge are small in absolute magnitude and produce no gradient across the active site.     

Comparative XANES study of serotransferrin and ovotransferrin at Cu K-edge: evidence of interactions among the metal sites

The Cu site structure of human serotransferrin and hen ovotransferrin using XANES spectroscopy has been investigated. Although the transferrin family proteins have been extensively studied, the results reported herein are the first concerning the structure of the metal site in C-terminal and N-terminal in the whole protein. Our structural data show that these proteins differ with regard to the independence of the two binding sites and the geometry of copper coordination, ranging from a poorly to a significantly distorted octahedron.     

Functional analysis of RPS27 mutations and expression in melanoma

Next‐generation sequencing has enabled genetic and genomic characterization of melanoma to an unprecedent depth. However, the high mutational background plus the limited depth of coverage of whole‐genome sequencing performed on cutaneous melanoma samples make the identification of novel driver mutations difficult. We sought to explore the somatic mutation portfolio in exonic and gene regulatory regions in human melanoma samples, for which we performed targeted sequencing of tumors and matched germline DNA samples from 89 melanoma patients, identifying known and novel recurrent mutations. Two recurrent mutations found in the RPS27 promoter associated with decreased RPS27 mRNA levels in vitro. Data mining and IHC analyses revealed a bimodal pattern of RPS27 expression in melanoma, with RPS27‐low patients displaying worse prognosis. In vitro characterization of RPS27‐high and RPS27‐low melanoma cell lines, as well as loss‐of‐function experiments, demonstrated that high RPS27 status provides increased proliferative and invasive capacities, while low RPS27 confers survival advantage in low attachment and resistance to therapy. Additionally, we demonstrate that 10 other cancer types harbor bimodal RPS27 expression, and in those, similarly to melanoma, RPS27‐low expression associates with worse clinical outcomes. RPS27 promoter mutation could thus represent a mechanism of gene expression modulation in melanoma patients, which may have prognostic and predictive implications.     

The Computational Mechanisms of Mate Choice

In many species, mating signals encode information important for both species recognition and mate quality assessment. I investigate how the computational mechanisms used by females to integrate the two sources of information can affect their mating decisions. First, I present a sequential analysis model of decision making based on a two‐component signal, in which the first component encodes information important for mate quality assessment and the second component for species recognition. When the components interact additively, the ability of females to discriminate between signals that differ in only the component important for mate‐quality assessment is the same independently of the value of the species‐recognition component. In contrast, when the components interact multiplicatively, discrimination in mate quality depends also on the species‐recognition component, which can either amplify or attenuate the perceived differences in mate quality. In the second part of the paper, I show results of a two‐choice phonotaxis experiment on female Italian treefrogs, Hyla intermedia, that confirm the model predictions by showing that directional preferences for call rate (important for mate‐quality assessment) are stronger when pulse‐rate and fundamental frequency (important for species recognition) are close to the preferred population mean than when they are either higher or lower than the mean.