Unrecognized controls on microbial functioning in Blue Carbon ecosystems: The role of mineral enzyme stabilization and allochthonous substrate supply

Tidal wetlands are effective carbon sinks, mitigating climate change through the long‐term removal of atmospheric CO2. Studies along surface‐elevation and thus flooding‐frequency gradients in tidal wetlands are often used to understand the effects of accelerated sea‐level rise on carbon sequestration, a process that is primarily determined by the balance of primary production and microbial decomposition. It has often been hypothesized that rates of microbial decomposition would increase with elevation and associated increases in soil oxygen availability; however, previous studies yield a wide range of outcomes and equivocal results. Our mechanistic understanding of the elevation–decomposition relationship is limited because most effort has been devoted to understanding the terminal steps of the decomposition process. A few studies assessed microbial exo‐enzyme activities (EEAs) as initial and rate‐limiting steps that often reveal important insight into microbial energy and nutrient constraints. The present study assessed EEAs and microbial abundance along a coastal ecotone stretching a flooding gradient from tidal flat to high marsh in the European Wadden Sea. We found that stabilization of exo‐enzymes to mineral sediments leads to high specific EEAs at low substrate concentrations in frequently flooded, sediment‐rich zones of the studied ecotone. We argue that the high background activity of a mineral‐associated enzyme pool provides a stable decomposition matrix in highly dynamic, frequently flooded zones. Furthermore, we demonstrate that microbial communities are less nutrient limited in frequently flooded zones, where inputs of nutrient‐rich marine organic matter are higher. This was reflected in both increasing exo‐enzymatic carbon versus nutrient acquisition and decreasing fungal versus bacterial abundance with increasing flooding frequency. Our findings thereby suggest two previously unrecognized mechanisms that may contribute to stimulated microbial activity despite decreasing oxygen availability in response to accelerated sea‐level rise

Top‐down control of carbon sequestration: grazing affects microbial structure and function in salt marsh soils

Tidal wetlands have been increasingly recognized as long‐term carbon sinks in recent years. Work on carbon sequestration and decomposition processes in tidal wetlands focused so far mainly on effects of global‐change factors such as sea‐level rise and increasing temperatures. However, little is known about effects of land use, such as livestock grazing, on organic matter decomposition and ultimately carbon sequestration. The present work aims at understanding the mechanisms by which large herbivores can affect organic matter decomposition in tidal wetlands. This was achieved by studying both direct animal–microbe interactions and indirect animal–plant–microbe interactions in grazed and ungrazed areas of two long‐term experimental field sites at the German North Sea coast. We assessed bacterial and fungal gene abundance using quantitative PCR, as well as the activity of microbial exo‐enzymes by conducting fluorometric assays. We demonstrate that grazing can have a profound impact on the microbial community structure of tidal wetland soils, by consistently increasing the fungi‐to‐bacteria ratio by 38–42%, and therefore potentially exerts important control over carbon turnover and sequestration. The observed shift in the microbial community was primarily driven by organic matter source, with higher contributions of recalcitrant autochthonous (terrestrial) vs. easily degradable allochthonous (marine) sources in grazed areas favoring relative fungal abundance. We propose a novel and indirect form of animal–plant–microbe interaction: top‐down control of aboveground vegetation structure determines the capacity of allochthonous organic matter trapping during flooding and thus the structure of the microbial community. Furthermore, our data provide the first evidence that grazing slows down microbial exo‐enzyme activity and thus decomposition through changes in soil redox chemistry. Activities of enzymes involved in C cycling were reduced by 28–40%, while activities of enzymes involved in N cycling were not consistently affected by grazing. It remains unclear if this is a trampling‐driven direct grazing effect, as hypothesized in earlier studies, or if the effect on redox chemistry is plant mediated and thus indirect. This study improves our process‐level understanding of how grazing can affect the microbial ecology and biogeochemistry of semi‐terrestrial ecosystems that may help explain and predict differences in C turnover and sequestration rates between grazed and ungrazed systems

MDF is a conserved splicing factor and modulates cell division and stress response in Arabidopsis

21 p.-8 fig.-1 tab.The coordination of cell division with stress response is essential for maintaining genome stability in plant meristems. Proteins involved in pre-mRNA splicing are important for these processes in animal and human cells. Based on its homology to the splicing factor SART1, which is implicated in the control of cell division and genome stability in human cells, we analyzed if MDF has similar functions in plants. We found that MDF associates with U4/U6.U5 tri-snRNP proteins and is essential for correct splicing of 2,037 transcripts. Loss of MDF function leads to cell division defects and cell death in meristems and was associated with up-regulation of stress-induced genes and down-regulation of mitotic regulators. In addition, the mdf-1 mutant is hypersensitive to DNA damage treatment supporting its role in coordinating stress response with cell division. Our analysis of a dephosphomutant of MDF suggested how its protein activity might be controlled. Our work uncovers the conserved function of a plant splicing factor and provides novel insight into the interplay of pre-mRNA processing and genome stability in plants.This work was funded by the Deutsche Forschungsgemeinschaft (DFG) grant to M Weingartner (WE4506/6-1). C de Luxán-Hernández was recipient of a PhD fellowship from the University of Hamburg. In addition, funding was received from the Spanish Ministry of Science and Innovation to J Salinas (PID2019-106987RB-I00/AEI/10.13039/5011033), from the Spanish Ministry of Science and Innovation to E Tranque (FPI contract), from the Czech Academy of Science, Deutsche Akademische Austauschdienst, and Bundesministeriums für Bildung und Forschung (BMBF) (CAS Mobility DAAD-19-04 and CAS Mobility DAAD-21-07) to M Weingartner and P Binarova.Peer reviewe

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Control of the AtMAP65-1 interaction with microtubules through the cell cycle

Cell division depends on the fine control of both microtubule dynamics and microtubule organisation. The microtubule bundling protein MAP65 is a ;midzone MAP' essential for the integrity of the anaphase spindle and cell division. Arabidopsis thaliana MAP65-1 (AtMAP65-1) binds and bundles microtubules by forming 25 nm cross-bridges. Moreover, as AtMAP65-1 bundles microtubules in interphase, anaphase and telophase but does not bind microtubules in prophase or metaphase, its activity through the cell cycle must be under tight control. Here we show that AtMAP65-1 is hyperphosphorylated during prometaphase and metaphase and that CDK and MAPK are involved in this phosphorylation. This phosphorylation inhibits AtMAP65-1 activity. Expression of non-phosphorylatable AtMAP65-1 has a negative effect on mitotic progression resulting in excessive accumulation of microtubules in the metaphase spindle midzone causing a delay in mitosis. We conclude that normal metaphase spindle organisation and the transition to anaphase is dependent on inactivation of AtMAP65-1

Dynamic Recruitment of Cdc2 to Specific Microtubule Structures during Mitosis

A-type cyclin-dependent kinases (CDKs), also known as cdc2, are central to the orderly progression of the cell cycle. We made a functional Green Fluorescent Protein (GFP) fusion with CDK-A (Cdc2-GFP) and followed its subcellular localization during the cell cycle in tobacco cells. During interphase, the Cdc2-GFP fusion protein was found in both the cytoplasm and the nucleus, where it was highly resistant to extraction. In premitotic cells, a bright and narrow equatorial band appeared on the cell surface, resembling the late preprophase band, which disintegrated within 10 min as followed by time-lapse images. Cdc2-GFP was not found on prophase spindles but left the chromatin soon after this stage and associated progressively with the metaphase spindle in a microtubule-dependent manner. Arresting cells in mitosis through the stabilization of microtubules by taxol further enhanced the spindle-localized pool of Cdc2-GFP. Toward the end of mitosis, Cdc2-GFP was found at the midzone of the anaphase spindle and phragmoplast; eventually, it became focused at the midline of these microtubule structures. In detergent-extracted cells, the Cdc2-GFP remained associated with mitotic structures. Retention on spindles was prevented by pretreatment with the CDK-specific inhibitor roscovitine and was enhanced by the protein phosphatase inhibitor okadaic acid. Furthermore, we demonstrate that both the endogenous CDK-A and Cdc2-GFP were cosedimented with taxol-stabilized plant microtubules from cell extracts and that Cdc2 activity was detected together with a fraction of polymerized tubulin. These data provide evidence that the A-type CDKs associate physically with mitotic structures in a microtubule-dependent manner and may be involved in regulating the behavior of specific microtubule arrays throughout mitosis

Structural and functional analysis of a plant nucleolar RNA chaperone-like protein

Ribosome biogenesis is a key process in all eukaryotic cells that requires hundreds of ribosome biogenesis factors (RBFs), which are essential to build the mature ribosomes consisting of proteins and rRNAs. The processing of the required rRNAs has been studied extensively in yeast and mammals, but in plants much is still unknown. In this study, we focused on a RBF from A. thaliana that we named NUCLEOLAR RNA CHAPERONE-LIKE 1 (NURC1). NURC1 was localized in the nucleolus of plant cell nuclei, and other plant RBF candidates shared the same localization. SEC-SAXS experiments revealed that NURC1 has an elongated and flexible structure. In addition, SEC-MALLS experiments confirmed that NURC1 was present in its monomeric form with a molecular weight of around 28 kDa. RNA binding was assessed by performing microscale thermophoresis with the Arabidopsis internal transcribed spacer 2 (ITS2) of the polycistronic pre-rRNA precursor, which contains the 5.8S, 18S, and 25S rRNA. NURC1 showed binding activity to the ITS2 with a dissociation constant of 228 nM and exhibited RNA chaperone-like activity. Our data suggested that NURC1 may have a function in pre-rRNA processing and thus ribosome biogenesis

A plant cyclin B2 is degraded early in mitosis and its ectopic expression shortens G2-phase and alleviates the DNA-damage checkpoint

Mitotic progression is timely regulated by the accumulation and degradation of A- and B-type cyclins. In plants, there are three classes of A-, and two classes of B-type cyclins, but their specific roles are not known. We have generated transgenic tobacco plants in which the ectopic expression of a plant cyclin B2 gene is under the control of a tetracycline-inducible promoter. We show that the induction of cyclin B2 expression in cultured cells during G2 phase accelerates the entry into mitosis and allows cells to override the replication checkpoint induced by hydroxyurea in the simultaneous presence of caffeine or okadaic acid, drugs that are known to alleviate checkpoint control. These results indicate that in plants, a B2-type cyclin is a rate-limiting regulator for the entry into mitosis and a cyclin B2-CDK complex might be a target for checkpoint control pathways. The cyclin B2 localization and the timing of its degradation during mitosis corroborate these conclusions: cyclin B2 protein is confined to the nucleus and during mitosis it is only present during a short time window until mid prophase, but it is effectively degraded from this timepoint onwards. Although cyclin B2 is not present in cells arrested by the spindle checkpoint in metaphase, cyclin B1 is accumulating in these cells. Ectopic expression of cyclin B2 in developing plants interferes with differentiation events and specifically blocks root regeneration, indicating the importance of control mechanisms at the G2- to M-phase transition during plant developmental processes.M.W. was supported by a PhD fellowship from the Austrian Academy of Sciences. This work was supported by an Austrian Förderung der Wissenschaftlichen Forschung grant to E.H.-B.; by European Union Framework V project ECCO Grant QLRT-1999-00454 to E.H.-B., L.B. and P.B.; by a Biotechnology and Biological Science Research Council grant (111/P133340) to L.B.; by grant LN00A081 from the Czech Ministry of Education; by a collaborative Wellcome Trust grant to P.B. and L.B.; and by a Spanish Ministerio de Ciencia y Tecnologia grant (BMC2001-2195) to C.T. Moreover, we thank Gireg Weingartner for his patience and Giovanna Vinti, among others, for taking care of him

Data_Mueller_etal2017_EcolAppl

Data_Mueller_etal2017_EcolApp