In vivo spectroscopy and NMR metabolite fingerprinting approaches to connect the dynamics of photosynthetic and metabolic phenotypes in resurrection plant Haberlea rhodopensis during desiccation and recovery.

International audienceThe resurrection plant Haberlea rhodopensis was used to study dynamics of drought response of photosynthetic machinery parallel with changes in primary metabolism. A relation between leaf water content and photosynthetic performance was established, enabling us to perform a non-destructive evaluation of the plant water status during stress. Spectroscopic analysis of photosynthesis indicated that, at variance with linear electron flow (LEF) involving photosystem (PS) I and II, cyclic electron flow around PSI remains active till almost full dry state at the expense of the LEF, due to the changed protein organization of photosynthetic apparatus. We suggest that, this activity could have a photoprotective role and prevent a complete drop in adenosine triphosphate (ATP), in the absence of LEF, to fuel specific energy-dependent processes necessary for the survival of the plant, during the late states of desiccation. The NMR fingerprint shows the significant metabolic changes in several pathways. Due to the declining of LEF accompanied by biosynthetic reactions during desiccation, a reduction of the ATP pool during drought was observed, which was fully and quickly recovered after plants rehydration. We found a decline of valine accompanied by lipid degradation during stress, likely to provide alternative carbon sources for sucrose accumulation at late stages of desiccation. This accumulation, as well as the increased levels of glycerophosphodiesters during drought stress could provide osmoprotection to the cells

Increased Phosphorylation of Vimentin in Noninfiltrative Meningiomas

International audienceBACKGROUND: Tissue invasion or tissue infiltration are clinical behaviors of a poor-prognosis subset of meningiomas. We carried out proteomic analyses of tissue extracts to discover new markers to accurately distinguish between infiltrative and noninfiltrative meningiomas. METHODOLOGY/PRINCIPAL FINDINGS: Protein lysates of 64 different tissue samples (including two brain-invasive and 32 infiltrative tumors) were submitted to SELDI-TOF mass spectrometric analysis. Mass profiles were used to build up both unsupervised and supervised hierarchical clustering. One marker was found at high levels in noninvasive and noninfiltrative tumors and appeared to be a discriminative marker for clustering infiltrative and/or invasive meningiomas versus noninvasive meningiomas in two distinct subsets. Sensitivity and specificity were 86.7% and 100%, respectively. This marker was purified and identified as a multiphosphorylated form of vimentin, a cytoskeletal protein expressed in meningiomas. CONCLUSIONS/SIGNIFICANCE: Specific forms of vimentin can be surrogate molecular indicators of the invasive/infiltrative phenotype in tumors

Derrière le masque du consensus… Analyse des divergences syndicales autour des enjeux de santé mentale au travail

International audienceThis article questions the dissenting position of trade union organisations for employees on the challenges of mental health in the workplace by referring to the primary results of a sociological survey that was led, at a confederal level, on leaders of occupational health and on the formation of approximately ten trade unions, as well on a study of documents (congress documents, trade union reports, websites). It shows that there are also structural “traditional” fracture lines concerning economic challenges, such as employment and employees. These divisions prove to be minimally effective for understanding the differences and agreements in opinion with regard to mental health, both in the means of noting the factors explaining these health risks – notably the role played by the organisation of work and management – and in identifying the risks themselves. However, these rifts clearly resurface with regard to defining the means of action and, above all, the purposes for which these are taken. The trade union positions are organised according to a timeline from trying to foster agreement with employers to the need to build strong relationships to make meaningful changes in this area.Cet article questionne le positionnement revendicatif des organisations syndicales de salariés sur les enjeux de santé mentale au travail en mobilisant les principaux résultats d’une enquête sociologique menée, au niveau confédéral, auprès des responsables de la santé au travail et de la formation d’une dizaine de syndicats, ainsi que sur une étude documentaire (documents de congrès, presse syndicale, sites internet). Il montre qu’aussi structurantes que puissent être les lignes de fracture « traditionnelles » instituées sur les enjeux économiques comme l’emploi ou les salaires, ces oppositions s’avèrent peu opérantes pour comprendre les divergences et convergences en matière de santé mentale, aussi bien dans la manière de désigner les facteurs explicatifs de ces risques de santé – notamment le rôle joué par l’organisation du travail et le management – que de nommer les risques eux-mêmes. Ces clivages ressurgissent toutefois nettement concernant la définition des modalités d’action et, surtout, des finalités poursuivies à travers elles. Les positions syndicales s’ordonnent alors selon un continuum allant du projet de convergence avec les employeurs à la nécessité de construire des rapports de force pour obtenir des avancées sur le sujet

À l’assaut du travail réel. Ce que la santé mentale fait au syndicalisme

International audienceCet article questionne le rapport des syndicats français à la santé mentale au travail dans la période 2000-2022. Il s’appuie sur les principaux résultats d’une enquête conduite entre 2020 et 2022, par le biais d’entretiens auprès de responsables en charge de cette question (aux niveaux confédéral, fédéral et local), par des observations de formations et journées d’études syndicales, et par une analyse documentaire. Il montre que, depuis les années 2000, les syndicats ont largement investi le champ de la santé au travail, s’appropriant, de diverses manières, des savoirs académiques variés. Tout en veillant à conserver une autonomie face aux « expert·e·s » et aux scientifiques, les organisations d’orientation contestataire s’emploient à mobiliser les analyses du travail réel et de l’activité pour transformer leur travail militant dans le sens d’une démarche « ascendante » . Les organisations de tradition réformiste mobilisent davantage ces nouvelles compétences pour mieux influencer les négociations avec le patronat. L’article montre ainsi que la santé mentale au travail est progressivement devenue un objet politique à part entière pour les syndicats français, mais qui demeure vulnérable aux conjonctures politiques, économiques et sociales

Quantitative proteomic analyses reveal the impact of nitrogen starvation on the proteome of the model diatom Phaeodactylum tricornutum

International audienceDiatoms are one of the largest groups in phytoplankton biodiversity. Understanding their response to nitrogen variations, present from micromolar to near-zero levels in oceans and fresh waters, is essential to comprehend their ecological success. Nitrogen starvation is used in biotechnological processes, to trigger the remodeling of carbon metabolism in the direction of fatty acids and triacylglycerol synthesis. We evaluated whole proteome changes in Phaeodactylum tricornutum after 7 days of cultivation with 5.5-mM nitrate (+N) or without any nitrogen source (-N). On a total of 3768 proteins detected in biological replicates, our analysis pointed to 384 differentially abundant proteins (DAP). Analysis of proteins of lower abundance in -N revealed an arrest of amino acid and protein syntheses, a remodeling of nitrogen metabolism, and a decrease of the proteasome abundance suggesting a decline in unselective whole-proteome decay. Analysis of proteins of higher abundance revealed the setting up of a general nitrogen scavenging system dependent on deaminases. The increase of a plastid palmitoyl-ACP desaturase appeared as a hallmark of carbon metabolism rewiring in the direction of fatty acid and triacylglycerol synthesis. This dataset is also valuable to select gene candidates for improved biotechnological properties

Hybrid cluster proteins in a photosynthetic microalga

International audienceHybrid cluster proteins (HCP) are metalloproteins characterized by the presence of an iron-sulfur-oxygen cluster. These proteins occur in all three domains of life. In eukaryotes, HCPs have so far been found only in a few anaerobic parasites and photosynthetic microalgae. With respect to all species harboring an HCP, the green microalga Chlamydomonas reinhardtii stands out by the presence of four HCP genes. The study of the gene and protein structures as well as the phylogenetic analyses strongly support a model in which the HCP family in the alga has emerged from a single gene of alpha proteobacterial origin and then expanded by several rounds of duplications. The spectra and redox properties of HCP1 and HCP3, produced heterologously in Escherichia coli, were analyzed by electron paramagnetic resonance spectroscopy on redox-titrated samples. Both proteins contain a [4Fe-4S]-cluster as well as a [4Fe-2O-2S]-hybrid cluster with paramagnetic properties related to those of HCPs from Desulfovibrio species. Immunoblotting experiments combined with mass spectrometry-based proteomics showed that both nitrate and darkness contribute to the strong up-regulation of the HCP levels in C. reinhardtii growing under oxic conditions. The link to the nitrate metabolism is discussed in the light of recent data on the potential role of HCP in S-nitrosylation in bacteria

Chloroplast biogenesis: towards the role of alternative protein targeting pathways in Arabidopsis

International audienceChloroplasts are a major component of plant cells. Their origin traces back to a cyanobacterial ancestor that was engulfed by an ancient eukaryotic cell and eventually integrated as an organelle during evolution. As a result, more than 95% of the ancestral cyanobacterial genes were transferred to the host cell nucleus. Proteins encoded by these relocated genes need to return to internal chloroplast compartments. This import is mainly achieved by the general TOC/TIC machinery located at the chloroplast surface. Until recently, all proteins destined to chloroplast were believed to possess an N-terminal and cleavable chloroplast targeting peptide, and to engage the TOC/TIC machinery. However, recent studies have revealed the existence of non-canonical preproteins, lacking cleavable transit peptides. Furthermore, few such proteins were demonstrated to use alternative targeting pathways, independent of the TOC/TIC machinery. The aim of our project aims at deciphering the molecular nature of these alternative targeting machineries. For that, we initiated a targeted study combining affinity purification and mass spectrometry aiming to identify alternative receptors at the chloroplast surface. Alternatively, we revisited the envelope proteome composition and initiated a gene candidate approach. We are currently studying the role of putative import receptors using in planta techniques

Exchange of metabolites and metabolic signals between plastids and cytosol and novel pathways of protein sorting towards plastids using the QORH protein as model

International audienceChloroplasts are a major component of plant cells. Their origin traces back to a cyanobacterial ancestor that was engulfed by an ancient eukaryotic cell and eventually integrated as an organelle during evolution. As a result, more than 95% of the ancestral cyanobacterial genes were transferred to the host cell nucleus. Proteins encoded by these relocated genes need to return to internal chloroplast compartments. This import is mainly achieved by the general TOC/TIC machinery located at the chloroplast surface. Until recently, all proteins destined to chloroplast were believed to possess an N-terminal and cleavable chloroplast targeting peptide, and to engage the TOC/TIC machinery. However, recent studies have revealed the existence of non-canonical preproteins, lacking cleavable transit peptides. Furthermore, few such proteins were demonstrated to use alternative targeting pathways, independent of the TOC/TIC machinery. The aim of our project aims at deciphering the molecular nature of these alternative targeting machineries. For that, we initiated a targeted study combining affinity purification and mass spectrometry aiming to identify alternative receptors at the chloroplast surface. Alternatively, we revisited the envelope proteome composition and initiated a gene candidate approach. We are currently studying the role of putative import receptors using in planta techniques

Proteomics unveil a central role for peroxisomes in butyrate assimilation of the heterotrophic Chlorophyte alga Polytomella sp.

Volatile fatty acids found in effluents of the dark fermentation of biowastes can be used for mixotrophic growth of microalgae, improving productivity and reducing the cost of the feedstock. Microalgae can use the acetate in the effluents very well, but butyrate is poorly assimilated and can inhibit growth above 1 gC.L−1. The non-photosynthetic chlorophyte alga Polytomella sp. SAG 198.80 was found to be able to assimilate butyrate fast. To decipher the metabolic pathways implicated in butyrate assimilation, quantitative proteomics study was developed comparing Polytomella sp. cells grown on acetate and butyrate at 1 gC.L−1. After statistical analysis, a total of 1772 proteins were retained, of which 119 proteins were found to be overaccumulated on butyrate vs. only 46 on acetate, indicating that butyrate assimilation necessitates additional metabolic steps. The data show that butyrate assimilation occurs in the peroxisome via the β-oxidation pathway to produce acetyl-CoA and further tri/dicarboxylic acids in the glyoxylate cycle. Concomitantly, reactive oxygen species defense enzymes as well as the branched amino acid degradation pathway were strongly induced. Although no clear dedicated butyrate transport mechanism could be inferred, several membrane transporters induced on butyrate are identified as potential condidates. Metabolic responses correspond globally to the increased needs for central cofactors NAD, ATP and CoA, especially in the peroxisome and the cytosol

Unravelling hidden components of the chloroplast envelope proteome: opportunities and limits of better MS sensitivity

International audienceThe chloroplast is a major plant cell organelle that fulfills essential metabolic and biosynthetic functions. Located at the interface between the chloroplast and other cell compartments, the chloroplast envelope system is a strategic barrier controlling the exchange of ions, metabolites and proteins, thus regulating essential metabolic functions (synthesis of hormones precursors, amino acids, pigments, sugars, vitamins, lipids, nucleotides…) of the plant cell. However, unraveling the contents of the chloroplast envelope proteome remains a difficult challenge; many proteins constituting this functional double membrane system remain to be identified. Indeed, the envelope contains only 1% of the chloroplast proteins (i.e. 0.4% of the whole cell proteome). In other words, most envelope proteins are so rare at the cell, chloroplast, or even envelope level, that they remained undetectable using targeted MS studies. Cross-contamination of chloroplast sub-compartments by each other and by other cell compartments during cell fractionation, impedes accurate localization of many envelope proteins. The aim of the present study was to take advantage of technologically improved MS sensitivity to better define the proteome of the chloroplast envelope (differentiate genuine envelope proteins from contaminants). This MS-based analysis relied on an enrichment factor that was calculated for each protein identified in purified envelope fractions as compared to the value obtained for the same protein in crude cell extracts. Using this approach, a total of 1269 proteins were detected in purified envelope fractions, of which, 462 could be assigned an envelope localization by combining MS-based spectral count analyses with manual annotation using data from the literature and prediction tools. Many of such proteins being previously unknown envelope components, these data constitute a new resource of significant value to the broader plant science community aiming to define principles and molecular mechanisms controlling fundamental aspects of plastid biogenesis and functions