Physico-chemical and high frequency monitoring dataset from mesocosm experiments simulating extreme climate events in lakes.

We present two datasets composed of high frequency sensors data, vertical in situ profiles and laboratory chemical analysis data, acquired during two different aquatic mesocosm experiments performed at the OLA ("Long-term observation and experimentation for lake ecosystems") facility at the UMR CARRTEL in Thonon les Bains, on the French shore of Lake Geneva. The DOMLAC experiment lasted 3 weeks (4-21 October 2021) and aimed to simulate predicted climate scenarios (i.e. extreme events such as storms and floods) by reproducing changes in quality and composition of lake subsidies and runoff by increased inputs of terrestrial organic matter. The PARLAC experiment lasted 3 weeks (5-23 September 2022) and aimed to simulate turbid storms by light reduction. The experimental setup consisted of nine inland polyester laminated tanks (2.1 m length, 2.1 m width and 1.1 m depth) with a total volume of approximately 4000 L and filled with water directly supplied from the lake at 4m depth. Both experimental design included three treatments each replicated three times. The DOMLAC experiment involved a control treatment (no treatment applied) and two treatments simulating allochthonous inputs from two different dissolved organic matter (DOM) extract from peat moss Sphagnum sp. (Peat-Moss treatment) and Phragmites australis (Phragmite treatment). The PARLAC experiment involved a control treatment (no treatment applied) and two treatments simulating two different intensity of light reduction. In the Medium treatment transmitted light was reduced to 70% and in the High treatment transmitted light was reduced to 15%. The datasets are composed of: 1. In situ measures from automated data loggers of temperature, conductivity, dissolved oxygen and CO2 acquired every 5 minutes at 0.1, 0.5 and 1 m depth (DOMLAC) and 0.5m (PARLAC) for the entire period of the experiment. 2. In situ profiles (0-1 m) of temperature, conductivity, pH, dissolved oxygen (concentration and saturation) acquired twice a week during the experiment. 3. In situ measures of light spectral UV/VIS/IR irradiance (300-950 nm wavelength range) taken in the air and at 0, 0.5 and 1 m twice a week on the same day of the profiles at point 2. 4. Laboratory chemical analysis of integrated samples taken twice a week on the same day of the in situ profiles at point 2 and 3 of conductivity, pH, total alkalinity, NO3, total and particulate nitrogen (Ntot, Npart), PO4, total and particulate phosphorus (Ptot, Ppart), total and particulate organic carbon (TOC, POC), Ca, K, Mg, Na, Cl, SO4 and SiO2. Only for DOMLAC also analyses of NH4, NO2 and dissolved organic carbon (DOC). 5. Laboratory analysis of pigments (Chla, Chlc, carotenoids, phaeopigments) extracted from samples collected at point 4. 6. Only for DOMLAC, specific absorbance on the range 600-200nm of DOM (i.e. <0.7 µm) measured on samples collected at point 4. This dataset aims to contribute our understanding of how extreme climate events can alter lake subsidies and affect the regulation of ecosystem processes such as production, respiration, nutrient uptake and pigment composition. The data can be used for a wide range of applications as being included in meta-analysis aiming at generalising the effect of climate change on large lakes including simulating future scenarios in a broad range of geographical areas as we used different inputs of DOM leached from litters reproducing catchments characteristics typical of different latitudes, such as mostly dominated by large leaf forests and phragmites at middle latitude, and coniferous forests rich of peat mosses that spread along the water surface typical of Northern regions.

Quantitative phosphoproteome analysis of Bacillus subtilis reveals novel substrates of the kinase PrkC and phosphatase PrpC.

Reversible protein phosphorylation on serine, threonine, and tyrosine (Ser/Thr/Tyr) residues plays a critical role in regulation of vital processes in the cell. Despite of considerable progress in our understanding of the role of this modification in bacterial physiology, the dynamics of protein phosphorylation during bacterial growth has rarely been systematically addressed. In addition, little is known about in vivo substrates of bacterial Ser/Thr/Tyr kinases and phosphatases. An excellent candidate to study these questions is the Gram-positive bacterium Bacillus subtilis, one of the most intensively investigated bacterial model organism with both research and industrial applications. Here we employed gel-free phosphoproteomics combined with SILAC labeling and high resolution mass spectrometry to study the proteome and phosphoproteome dynamics during the batch growth of B. subtilis. We measured the dynamics of 1666 proteins and 64 phosphorylation sites in five distinct phases of growth. Enzymes of the central carbon metabolism and components of the translation machinery appear to be highly phosphorylated in the stationary phase, coinciding with stronger expression of Ser/Thr kinases. We further used the SILAC workflow to identify novel putative substrates of the Ser/Thr kinase PrkC and the phosphatase PrpC during stationary phase. The overall number of putative substrates was low, pointing to a high kinase and phosphatase specificity. One of the phosphorylation sites affected by both, PrkC and PrpC, was the Ser281 on the oxidoreductase YkwC. We showed that PrkC phosphorylates and PrpC dephosphorylates YkwC in vitro and that phosphorylation at Ser281 abolishes the oxidoreductase activity of YkwC in vitro and in vivo. Our results present the most detailed phosphoproteomic analysis of B. subtilis growth to date and provide the first global in vivo screen of PrkC and PrpC substrates.

A forward genetic approach in Chlamydomonas reinhardtii as a strategy for exploring starch catabolism.

A screen was recently developed to study the mobilization of starch in the unicellular green alga Chlamydomonas reinhardtii. This screen relies on starch synthesis accumulation during nitrogen starvation followed by the supply of nitrogen and the switch to darkness. Hence multiple regulatory networks including those of nutrient starvation, cell cycle control and light to dark transitions are likely to impact the recovery of mutant candidates. In this paper we monitor the specificity of this mutant screen by characterizing the nature of the genes disrupted in the selected mutants. We show that one third of the mutants consisted of strains mutated in genes previously reported to be of paramount importance in starch catabolism such as those encoding ß-amylases, the maltose export protein, and branching enzyme I. The other mutants were defective for previously uncharacterized functions some of which are likely to define novel proteins affecting starch mobilization in green algae.

Protein-serine/threonine/tyrosine kinases in bacterial signaling and regulation.

In this review, we address some recent developments in the field of bacterial protein phosphorylation, focusing specifically on serine/threonine and tyrosine kinases. We present an overview of recent studies outlining the scope of physiological processes that are regulated by phosphorylation, ranging from cell cycle, growth, cell morphology, to metabolism, developmental phenomena, and virulence. Specific emphasis is placed on Mycobacterium tuberculosis as a showcase organism for serine/threonine kinases, and Bacillus subtilis to illustrate the importance of protein phosphorylation in developmental processes. We argue that bacterial serine/threonine and tyrosine kinases have a distinctive feature of phosphorylating multiple substrates and might thus represent integration nodes in the signaling network. Some open questions regarding the evolutionary benefits of relaxed substrate selectivity of these kinases are treated, as well as the notion of nonfunctional 'background' phosphorylation of cellular proteins. We also argue that phosphorylation events for which an immediate regulatory effect is not clearly established should not be dismissed as unimportant, as they may have a role in cross-talk with other post-translational modifications. Finally, recently developed methods for studying protein phosphorylation networks in bacteria are briefly discussed.

Protein phosphorylation from the perspective of systems biology.

Protein phosphorylation pathways emerge as large and interconnected networks, involving mutually activating protein kinases, kinases acting as network nodes by phosphorylating different substrates, and cross-talk of phosphorylation with other post-translational modifications. The complexity of these networks clearly necessitates the use of systems biology approaches. Phosphoproteomics represents the basis for detection of phosphoproteins and phosphorylation sites, but it must be combined with transcriptomics and interactomics in attempts to build in silico phosphorylation networks. This review highlights the implication of phosphorylation in cellular physiology across all domains of life. It focuses particularly on reports of human disease correlated to defects in phosphorylation networks. Brief outline of developments in quantitative mass spectrometry-based proteomics and bioinformatic tools specific for phosphoproteome studies is provided.