Preliminary studies on haematological and plasmatic parameters in gilthead sea bream (Sparus aurata) held under day/night temperature variations.

This study aimed to evaluate to what extent diel water temperature oscillations over a short period of time (41 h) affected haematologic and plasmatic parameters of gilthead sea bream. Sea bream juveniles (160 fish; 177.2 ± 20.9 g) were divided in two homogenous groups, experimental and control. The experimental group was exposed to water at 22.3 ± 0.6 °C between 5 a.m. and 5 p.m. and to colder water from 5 p.m. to 5 a.m. (temperature variation of 8.6 ± 0.3 °C) during 41h period, whereas for the control group, water temperature was maintained constant (22.4 ± 0.7 °C) for the same period. Fish were fed three times per day (10:30 a.m., 2 p.m., 4 p.m.). Blood was collected from fish caudal veins (n = 6) at different time points (10 a.m., 1 p.m., 5 p.m., 6:30 p.m., 8 p.m. and 10 a.m. of the following day) from each group, for haematologic (haematocrit and haemoglobin) and plasmatic (glucose, cortisol, triglycerides and total protein) determinations. All parameters studied tended to vary throughout the study period, both for experimental and control groups. Significant differences were observed between the two groups for all the studied parameters at different sampling points. These differences were observed near the influence of the cold-water period or during the cooling-water period. Results suggested that colder water temperature cycles depressed fish metabolic activity and affected the levels of the analysed blood-circulating metabolites. Overall, results suggested that stress response was similar for experimental and control groups and that gilthead sea bream were able to cope with diel water temperature variations.

Hsp70 and Hsp110 Chaperones Promote Early Steps of Proteasome Assembly.

Whereas assembly of the 20S proteasome core particle (CP) in prokaryotes apparently occurs spontaneously, the efficiency of this process in eukaryotes relies on the dedicated assembly chaperones Ump1, Pba1-Pba2, and Pba3-Pba4. For mammals, it was reported that CP assembly initiates with formation of a complete α-ring that functions as a template for ß subunit incorporation. By contrast, we were not able to detect a ring composed only of a complete set of α subunits in S. cerevisiae. Instead, we found that the CP subunits α1, α2, and α4 each form independent small complexes. Purification of such complexes containing α4 revealed the presence of chaperones of the Hsp70/Ssa and Hsp110/Sse families. Consistently, certain small complexes containing α1, α2, and α4 were not formed in strains lacking these chaperones. Deletion of the SSE1 gene in combination with deletions of PRE9 (α3), PBA3, or UMP1 genes resulted in severe synthetic growth defects, high levels of ubiquitin-conjugates, and an accumulation of distinct small complexes with α subunits. Our study shows that Hsp70 and Hsp110 chaperones cooperate to promote the folding of individual α subunits and/or their assembly with other CP subunits, Ump1, and Pba1-Pba4 in subsequent steps.

A multi-camera and multimodal dataset for posture and gait analysis.

Monitoring gait and posture while using assisting robotic devices is relevant to attain effective assistance and assess the user's progression throughout time. This work presents a multi-camera, multimodal, and detailed dataset involving 14 healthy participants walking with a wheeled robotic walker equipped with a pair of affordable cameras. Depth data were acquired at 30 fps and synchronized with inertial data from Xsens MTw Awinda sensors and kinematic data from the segments of the Xsens biomechanical model, acquired at 60 Hz. Participants walked with the robotic walker at 3 different gait speeds, across 3 different walking scenarios/paths at 3 different locations. In total, this dataset provides approximately 92 minutes of total recording time, which corresponds to nearly 166.000 samples of synchronized data. This dataset may contribute to the scientific research by allowing the development and evaluation of: (i) vision-based pose estimation algorithms, exploring classic or deep learning approaches; (ii) human detection and tracking algorithms; (iii) movement forecasting; and (iv) biomechanical analysis of gait/posture when using a rehabilitation device.

Biphasic modulation of fatty acid synthase by hydrogen peroxide in Saccharomyces cerevisiae.

Taking into account published contradictory results concerning the regulation of fatty acid synthase (Fas) by H(2)O(2), we carried out a systematic study where two methods of H(2)O(2) delivery (steady-state and bolus addition) and the effect of a wide range of H(2)O(2) concentrations were investigated. A decrease in Fas activity was observed for cells exposed to 100 and 150µM H(2)O(2) in a steady-state, while a bolus addition of the same H(2)O(2) concentrations did not alter Fas activity. Similar results were observed for the mRNA levels of FAS1, the gene that encodes Fas subunit ß. However, the exposure to a steady-state 50µM H(2)O(2) dose lead to an increase in FAS1 mRNA levels, showing a biphasic modulation of Fas by H(2)O(2). The results obtained emphasize that cellular effects of H(2)O(2) can vary over a narrow range of concentrations. Therefore, a tight control of H(2)O(2) exposure, which can be achieved by exposing H(2)O(2) in a steady-state, is important for cellular studies of H(2)O(2)-dependent redox regulation.

Chaperone-assisted assembly of the proteasome core particle.

The 26S proteasome is a non-lysosomal protease in the cytosol and nucleus of eukaryotic cells. Its main function is to mediate ubiquitin-dependent proteolysis. The 26S proteasome is a multimeric complex composed by the 20S proteasome CP (core particle) and the 19S RPs (regulatory particles). Although the atomic structure of the 26S proteasome has not yet been determined, high-resolution structures are available for its CP. Studies on the complicated assembly pathway of the proteasome have revealed that it involves an unprecedented number of dedicated chaperones. Assembly of the CP alone involves three conserved proteasome-assembly chaperones [PAC1-PAC2, PAC3-PAC4 and UMP1 (ubiquitin-mediated proteolysis 1)]. Whereas the two heterodimeric PACs have been implicated in the formation of rings of the seven distinct alpha subunits, UMP1 is important for the formation and dimerization of proteasome precursor complexes containing beta subunits. Dimerization coincides with the incorporation of the last beta subunit (beta7). Additional modules important for the assembly of precursor complexes and their dimerization reside in the beta subunits themselves, either as transient or as permanent extensions. Particularly important domains are the propeptide of beta5 and the C-terminal extensions of beta2 and beta7. Upon maturation of the active sites by autocatalytic processing, UMP1 is degraded by the native proteasome.

Toxicological effects of the rare earth element neodymium in Mytilus galloprovincialis.

The wide range of applications of rare earth elements (REE) is leading to their occurrence in worldwide aquatic environments. Among the most popular REE is Neodymium (Nd), being widely used in permanent magnets, lasers, and glass additives. Neodymium-iron-boron (NdFeB) magnets is the main application of Nd since they are used in electric motors, hard disk drives, speakers and generators for wind turbines. Recent studies have already evaluated the toxic potential of different REE, but no information is available on the effects of Nd towards marine bivalves. Thus, the present study evaluated the biochemical alterations caused by Nd in the mussel Mytilus galloprovincialis exposed to this element for 28 days. The results obtained clearly demonstrated that Nd was accumulated by mussels, leading to mussel's metabolic capacity increase and GLY expenditure, in an attempt to fuel up defense mechanisms. Antioxidant and biotransformation defenses were insufficient in the elimination of ROS excess, resulting from the presence of Nd and increased electron transport system activity, which caused cellular damages (measured by lipid peroxidation) and loss of redox balance (assessed by the ratio between reduced and oxidized glutathione). The results obtained clearly highlight the potential toxicity of REEs and, in particular of Nd, with impacts at cellular level, which may have consequences in mussel's survival, growth and reproduction, affecting mussel's population.

H2O2 induces rapid biophysical and permeability changes in the plasma membrane of Saccharomyces cerevisiae.

In Saccharomyces cerevisiae, the diffusion rate of hydrogen peroxide (H2O2) through the plasma membrane decreases during adaptation to H2O2 by means of a mechanism that is still unknown. Here, evidence is presented that during adaptation to H2O2 the anisotropy of the plasma membrane increases. Adaptation to H2O2 was studied at several times (15min up to 90min) by applying the steady-state H2O2 delivery model. For wild-type cells, the steady-state fluorescence anisotropy increased after 30min, or 60min, when using 2-(9-anthroyloxy) stearic acid (2-AS), or diphenylhexatriene (DPH) membrane probe, respectively. Moreover, a 40% decrease in plasma membrane permeability to H2O2 was observed at 15min with a concomitant two-fold increase in catalase activity. Disruption of the ergosterol pathway, by knocking out either ERG3 or ERG6, prevents the changes in anisotropy during H2O2 adaptation. H2O2 diffusion through the plasma membrane in S. cerevisiae cells is not mediated by aquaporins since the H2O2 permeability constant is not altered in the presence of the aquaporin inhibitor mercuric chloride. Altogether, these results indicate that the regulation of the plasma membrane permeability towards H2O2 is mediated by modulation of the biophysical properties of the plasma membrane.

Down-regulation of fatty acid synthase increases the resistance of Saccharomyces cerevisiae cells to H2O2.

Changes in plasma membrane permeability caused by H2O2 were recently found to be involved in the adaptation to H2O2, but the mechanism responsible for this change remains largely unknown. Here this mechanism was addressed and two lines of evidence showed for the first time that fatty acid synthase (Fas) plays a key role during the cellular response of Saccharomyces cerevisiae to H2O2: (1) adaptation was associated with a decrease in both Fas expression and activity; (2) more importantly, decreasing Fas activity by 50% through deletion of one of the FAS alleles increased the resistance to lethal doses of H2O2. The mechanism by which a decrease of Fas expression causes a higher resistance to H2O2 was not fully elucidated. However, the fas1Delta strain plasma membrane had large increases in the levels of lignoceric acid (C24:0) (40%) and cerotic acid (C26:0) (50%), suggesting that alterations in the plasma membrane composition are involved. Very-long-chain fatty acids (VLCFA) through interdigitation or by modulating formation of lipid rafts may decrease the overall or localized plasma membrane permeability to H2O2, respectively, thus conferring a higher resistance to H2O2.

Compacting proteins: pros and cons of osmolyte-induced folding.

Biomedical applications of osmolytes, including stabilization of protein-based pharmaceutics, preservation of living biological material and potential therapeutic prescription in vivo, are intimately related to the fact that osmolytes favour the native structure of proteins. The shift towards the native structure is associated to the compaction of the protein by a non-specific mechanism. This compaction is observed mostly for the unfolded state but also for the transition state ensemble and even for the native state. In addition, more stable three-dimensional structures are more stabilized by osmolytes if the overall protein fold is the same indicating that point mutations and osmolytes should share a similar mechanism for protein stabilization. A synergistic effect to increase protein stability between accumulation of osmolytes and protein engineering strategies seems to have operated during evolution. However, the conformational pre-organization of the unfolded state (compaction) induced by osmolytes which increases the folding rate, might lead to the accumulation of off-folding pathway intermediates with non-native structure that delay folding. Also, osmolytes favor protein aggregation as an alternative way to shield protein surfaces from the solvent. The sometimes observed effect of osmolytes on the prevention of protein aggregation is apparent as they only decrease the accumulation of aggregation-competent partially unfolded states.

Modulation of plasma membrane lipid profile and microdomains by H2O2 in Saccharomyces cerevisiae.

In Saccharomyces cerevisiae, the rate of hydrogen peroxide (H(2)O(2)) diffusion through the plasma membrane decreases during adaptation to H(2)O(2) by a still unknown mechanism. Here, adaptation to H(2)O(2) was observed to modulate rapidly the expression of genes coding for enzymes involved in ergosterol and lipid metabolism. Adaptation to H(2)O(2) also alters plasma membrane lipid composition. The main changes were the following: (a) there was a decrease in oleic acid (30%) and in the ratio between unsaturated and saturated long-chain fatty acids; (b) the phosphatidylcholine:phosphatidylethanolamine ratio increased threefold; (c) sterol levels were unaltered but there was an increased heterogeneity of sterol-rich microdomains and increased ordered domains; (d) the levels of the sterol precursor squalene increased twofold, in agreement with ERG1 gene down-regulation; and (e) C26:0 became the major very long chain fatty acid owing to an 80% decrease in 2-hydroxy-C26:0 levels and a 50% decrease in C20:0 levels, probably related to the down-regulation of fatty acid elongation (FAS1, FEN1, SUR4) and ceramide synthase (LIP1, LAC1) genes. Therefore, H(2)O(2) leads to a reorganization of the plasma membrane microdomains, which may explain the lower permeability to H(2)O(2), and emerges as an important regulator of lipid metabolism and plasma membrane lipid composition.