Biology subjects

Schmidt, O.

Publications and source records attributed to Schmidt, O..

14 recordsLinked to original sources

bioRxiv2026

Heat tolerance and its seasonal acclimation in Fagus sylvatica compared to Fagus orientalis and Pseudotsuga menziesii

Heat tolerance determines the vitality of tree species under climate change independently of drought tolerance, but has been much less studied than tree water relations. We studied species-specific differences and the capacity for seasonal heat acclimation in Central Europes naturally most important tree species, Fagus sylvatica, in comparison with two exotic tree species (Fagus orientalis, Pseudotsuga menziesii) that are considered for silvicultural climate change adaptation in managed forests. Foliage of mature trees was incubated at temperatures from 35-50 {degrees}C for up to 4 h to simulate daily heat maxima during heat waves. The maximum quantum yield (Fv/Fm) of photosystem II (PS II) of dark-adapted leaves was measured, because the PS II is particularly sensitive to heat and its functionality can decide on plant survival under heat. Fagus sylvatica was much more tolerant to heat than Pseudotsuga menziesii, but weakly (albeit significantly) less tolerant than Fagus orientalis. Within its limits, Pseudotsuga menziesii showed high seasonal heat acclimation with constantly increasing tolerance during the growing season. Fagus orientalis, but practically not Fagus sylvatica, also acclimated to heat. This makes Fagus orientalis slightly superior over Fagus sylvatica in terms of heat tolerance, whereas the suitability of Pseudotsuga menziesii for silvicultural climate change adaptation is questionable. Strong heat acclimation, but also overall low heat tolerance, in Pseudotsuga menziesii might be the result of evergreenness, which requires the generation of both cold and heat tolerance during the year.

ecology↗

bioRxiv2024

Protozoal populations drive system-wide variation in the rumen microbiome

While rapid progress has been made to characterize the bacterial and archaeal populations of the rumen microbiome, insight into how they interact with keystone protozoal species remains elusive. Here, we reveal two distinct rumen community types (RCT-A and RCT-B) that are not strongly associated with host phenotype nor genotype but instead linked to protozoal community patterns. We leveraged a series of multi-omic datasets to show that the dominant Epidinium spp. in animals with RCT-B employ a plethora of fiber-degrading enzymes that present enriched Prevotella spp. a favorable carbon landscape to forage upon. Conversely, animals with RCT-A, dominated by genera Isotricha and Entodinium, harbor a more even distribution of fiber, protein, and amino acid metabolizers, reflected by higher detection of metabolites from both protozoal and bacterial activity. We reveal microbiome variation across key protozoal and bacterial populations is interlinked, which should act as an important consideration for future development of microbiome-based technologies.

microbiology↗

bioRxiv2024

Methanogenesis inhibition remodels microbial fermentation and stimulates acetogenesis in ruminants

Rumen microbiota enable ruminants to grow on fibrous plant materials but also produce methane, driving 5% of global greenhouse gas emissions and leading to a loss of gross energy content. Methanogenesis inhibitors such as 3-nitrooxypropanol (3-NOP) decrease methane emissions in ruminants when supplemented in feed. Yet we lack a system-wide, species-resolved understanding of how the rumen microbiota remodels following inhibition and how this influences animal production. Here, we conducted a large-scale trial with 51 dairy calves to analyse microbiota responses to 3-NOP, pairing host performance, emissions, and nutritional profiles with genome-resolved metagenomic and metatranscriptomic data. 3-NOP supplementation decreased methane emissions by an average of 62%, modulated short-chain fatty acid and H2 levels, and did not affect dietary intake or animal performance. We created a rumen microbial genome catalogue with an unprecedented mapping rate. We observed a strong reduction of methanogens and stimulation of reductive acetogens, primarily novel uncultivated lineages such as Candidatus Faecousia. However, there was a shift in major fermentative communities away from acetate production in response to hydrogen gas accumulation. Thus, the divergent responses of the fermentative and hydrogenotrophic communities limit potential productivity gains from methane reduction. Reporting one of the largest reductions in methane emissions in a field trial to date, this study links ruminant greenhouse gas emissions and productivity to specific microbial species. These findings also emphasise the importance of microbiota-wide analysis for optimising methane mitigation strategies and identify promising strategies to simultaneously reduce emissions while increasing animal production. Significance StatementOne strategy to increase the sustainability and productivity of livestock production is to modulate ruminant microbiota to produce absorbable nutrients rather than the potent greenhouse gas methane. Previous studies show supplementing feed with methanogenesis inhibitors such as 3-nitrooxypropanol reduces methane emissions, but also leads to inconsistent productivity gains. Here we report a definitive field trial, combining animal data, meta-omics, and structural modelling, to resolve the key microbes and pathways controlling nutrient and methane production in ruminants. We show that shifts in composition and gene expression of hydrogen-cycling microbes reduce emissions but limit productivity gains. These findings offer insights at unprecedented resolution, while the data and analytical framework provide valuable resources to develop solutions to enhance livestock productivity and sustainability.

microbiology↗

bioRxiv2024

Microbial aerotrophy enables continuous primary production in diverse cave ecosystems

Most aerated cave ecosystems are assumed to be oligotrophic given they receive minimal inputs of light energy. Diverse microorganisms have nevertheless been detected within caves, though it remains unclear what strategies enable them to meet their energy and carbon needs. Here we determined the processes and mediators of primary production in aerated limestone and basalt caves through paired metagenomic and biogeochemical profiling. Based on 1458 metagenome-assembled genomes, over half of microbial cells in caves encode enzymes to use atmospheric trace gases as energy and carbon sources. The most abundant microbes in these systems are chemosynthetic primary producers, notably the novel gammaproteobacterial methanotrophic order Ca. Methylocavales and two uncultivated actinobacterial genera predicted to grow on atmospheric hydrogen, carbon dioxide, and carbon monoxide. In situ and ex situ biogeochemical and isotopic measurements consistently confirmed that these gases are rapidly consumed at rates sufficient to meet community-wide energy needs and drive continual primary production. Conventional chemolithoautotrophs, which use trace lithic compounds such as ammonium and sulfide, are also enriched and active alongside these trace gas scavengers. These results indicate that caves are unique in both their microbial composition and the biogeochemical processes that sustain them. Based on these findings, we propose caves are the first known ecosystems where atmospheric trace gases primarily sustain growth rather than survival and define this process as aerotrophy. Cave aerotrophy may be a hidden process supporting global biogeochemistry.

microbiology↗

bioRxiv2024

Revealing the Arabidopsis AtGRP7 mRNA binding proteome by specific enhanced RNA interactome capture

BackgroundThe interaction of proteins with RNA in the cell is crucial to orchestrate all steps of RNA processing. RNA interactome capture (RIC) techniques have been implemented to catalogue RNA-binding proteins in the cell. In RIC, RNA-protein complexes are stabilized by UV crosslinking in vivo. Polyadenylated RNAs and associated proteins are pulled down from cell lysates using oligo(dT) beads and the RNA-binding proteome is identified by quantitative mass spectrometry. However, insights into the RNA-binding proteome of a single RNA that would yield mechanistic information on how RNA expression patterns are orchestrated, are scarce. ResultsHere, we explored RIC in Arabidopsis to identify proteins interacting with a single mRNA, using the circadian clock-regulated Arabidopsis thaliana GLYCINE-RICH RNA-BINDING PROTEIN 7 (AtGRP7) transcript, one of the most abundant transcripts in Arabidopsis, as a showcase. Seedlings were treated with UV light to covalently crosslink RNA and proteins. The AtGRP7 transcript was captured from cell lysates with antisense oligonucleotides directed against the 5untranslated region (UTR). The efficiency of RNA capture was greatly enhanced by using locked nucleic acid (LNA)/DNA oligonucleotides, as done in the enhanced RIC protocol. Furthermore, performing a tandem capture with two rounds of pulldown with the 5UTR oligonucleotide increased the yield. In total, we identified 356 proteins enriched relative to a pulldown from atgrp7 mutant plants. These were benchmarked against proteins pulled down from nuclear lysates by AtGRP7 in vitro transcripts immobilized on beads. Among the proteins validated by in vitro interaction we found the family of Acetylation Lowers Binding Affinity (ALBA) proteins. Interaction of ALBA4 with the AtGRP7 RNA was independently validated via individual-nucleotide resolution crosslinking and immunoprecipitation (iCLIP). The expression of the AtGRP7 transcript in an alba loss-of-function mutant was slightly changed compared to wild-type, demonstrating the functional relevance of the interaction. ConclusionWe adapted specific RNA interactome capture with LNA/DNA oligonucleotides for use in plants using AtGRP7 as a showcase. We anticipate that with further optimization and up-scaling the protocol should be applicable for less abundant transcripts.

plant biology↗

bioRxiv2024

The Dsc ubiquitin ligase complex identifies transmembrane degrons to degrade orphaned proteins at the Golgi

The Golgi apparatus is essential for protein sorting, yet its quality control mechanisms are poorly understood. Here we show that the Dsc ubiquitin ligase complex, particularly the rhomboid pseudo-protease subunit, Dsc2, assesses the hydrophobic length of -helical transmembrane domains (TMDs) at the Golgi. Thereby the Dsc complex interacts with orphaned ER and Golgi proteins that have shorter TMDs and ubiquitinates them for targeted degradation. Some Dsc substrates will be K63 polyubiquitinated for ESCRT dependent vacuolar degradation or K48 polyubiquitinated for endosome and Golgi associated proteasomal degradation (EGAD). Other Dsc substrates are exclusively extracted by Cdc48 for EGAD. The accumulation of Dsc substrates entails a specific increase in glycerophospholipids with shorter and asymmetric fatty acyl chains. Hence, the Dsc complex mediates the selective degradation of orphaned proteins at the sorting center of cells, which prevents their spreading across other organelles and thus preserves cellular membrane protein and lipid composition.

cell biology↗

bioRxiv2024

The structure of the Orm2-containing serine palmitoyltransferase complex reveals distinct inhibitory potentials of yeast Orm proteins.

The levels of sphingolipids are crucial determinants of neurodegenerative disorders. Therefore, sphingolipid levels have to be tightly regulated. The Orm protein family and ceramides act as inhibitors in the rate-limiting step of sphingolipid biosynthesis - the condensation of L-serine and palmitoyl-CoA. The two yeast isoforms Orm1 and Orm2 form a complex with the serine palmitoyltransferase (SPT). While the sequences of the Orm proteins are highly similar, however, Orm1 and Orm2 are differentially regulated in yeast cells. The mechanistic details of the differential regulation remain elusive. To elucidate the regulatory mechanism, we determined the cryo-electron microscopy structure of the SPT complex containing Orm2. Through in vitro activity assays and a newly developed plasmid coverage assay combined with targeted lipidomics, we demonstrate that the SPT complex with Orm2 exhibits lower activity in comparison to the complex containing Orm1. This collectively suggests a higher inhibitory potential of Orm2, despite the remarkably similar structures of the Orm1- and Orm2 containing complexes. The high conservation of the SPT from yeast to man implies that the three human ORMDL isoforms could also have different regulatory capacities, which might be essential to understanding their role in sphingolipid-mediated neurodegenerative disorders. HighlightsO_LICryo-EM structures of serine palmitoyltransferase with Orm2 from S. cerevisiae C_LIO_LICeramide-mediated inhibition on SPT activity is conserved amongst SPTs C_LIO_LIThe two yeast Orm isoforms have different regulatory capacity C_LI

biochemistry↗

bioRxiv2023

A metabolically controlled contact site between lipid droplets and vacuoles

The lipid droplet (LD) organization proteins Ldo16 and Ldo45 affect multiple aspects of LD biology in yeast. They are linked to the LD biogenesis machinery seipin, and their loss causes defects in LD positioning, protein targeting, and breakdown. However, their molecular roles remained enigmatic. Here we report that Ldo16/45 form a tether-complex with Vac8 for creation of vacuole lipid droplet (vCLIP) contact sites, which can form in the absence of seipin. The phosphatidylinositol transfer protein Pdr16 is a further vCLIP-resident recruited by Ldo45. While only an LD-subpopulation is engaged in vCLIPs at glucose-replete conditions, nutrient stress results in vCLIP expansion, and vCLIP defects impair lipophagy upon prolonged starvation. In summary, Ldo16/45 are multifunctional proteins that orchestrate formation of a metabolically-regulated contact site. Our studies suggest an unexpected link between LD biogenesis and breakdown, and open the door to a deeper understanding of how lipid homeostasis is maintained during metabolic challenges.

cell biology↗

bioRxiv2023

Utilizing a nanobody recruitment approach for assessing serine palmitoyltransferase activity in ER sub-compartments of yeast.

Sphingolipids (SP) are one of the three major lipid classes in eukaryotic cells and serve as structural components of the plasma membrane. The rate-limiting step in SP biosynthesis is catalyzed by serine palmitoyltransferase (SPT). In yeast, SPT consists of two catalytic subunits (Lcb1 and Lcb2), a regulatory subunit (Tsc3), negative regulators (Orm1 and Orm2), and the phosphatidylinositol-4-phosphate (PI4P) phosphatase Sac1, collectively known as the SPOTS complex. Regulating SPT activity enables cells to adapt SP metabolism to changing environmental conditions. Therefore, the Orm proteins are phosphorylated by two signaling pathways originating from either the plasma membrane localized target of rapamycin (TOR) complex 2 or the lysosomal/vacuolar TOR complex 1. Moreover, uptake of exogenous serine is necessary for the regulation of SP biosynthesis, which suggests the existence of differentially regulated SPT pools based on their intracellular localization. However, tools for measuring lipid metabolic enzyme activity in different cellular compartments are currently not available. We have developed a nanobody recruitment system that enables the re-localization of the SPOTS complex to the nuclear or peripheral ER. By combining this system with sphingolipid flux analysis, we have identified two distinct active SPT pools in cells. Our method thus serves as a new and versatile tool to measure lipid metabolism with sub-cellular resolution.

biochemistry↗

bioRxiv2022

Global monitoring of soil animal communities using a common methodology

Here we introduce the Soil BON Foodweb Team, a cross-continental collaborative network that aims to monitor soil animal communities and food webs using consistent methodology at a global scale. Soil animals support vital soil processes via soil structure modification, direct consumption of dead organic matter, and interactions with microbial and plant communities. Soil animal effects on ecosystem functions have been demonstrated by correlative analyses as well as in laboratory and field experiments, but these studies typically focus on selected animal groups or species at one or few sites with limited variation in environmental conditions. The lack of comprehensive harmonised large-scale soil animal community data including microfauna, mesofauna, and macrofauna, in conjunction with related soil functions, limits our understanding of biological interactions in soil communities and how these interactions affect ecosystem functioning. To provide such data, the Soil BON Foodweb Team invites researchers worldwide to use a common methodology to address six long-term goals: (1) to collect globally representative harmonised data on soil micro-, meso-, and macrofauna communities; (2) to describe key environmental drivers of soil animal communities and food webs; (3) to assess the efficiency of conservation approaches for the protection of soil animal communities; (4) to describe soil food webs and their association with soil functioning globally; (5) to establish a global research network for soil biodiversity monitoring and collaborative projects in related topics; (6) to reinforce local collaboration networks and expertise and support capacity building for soil animal research around the world. In this paper, we describe the vision of the global research network and the common sampling protocol to assess soil animal communities and advocate for the use of standard methodologies across observational and experimental soil animal studies. We will use this protocol to conduct soil animal assessments and reconstruct soil food webs on the sites included in the global soil biodiversity monitoring network, Soil BON, allowing us to assess linkages among soil biodiversity, vegetation, soil physico-chemical properties, and ecosystem functions. In the present paper, we call for researchers especially from countries and ecoregions that remain underrepresented in the majority of soil biodiversity assessments to join us. Together we will be able to provide science-based evidence to support soil biodiversity conservation and functioning of terrestrial ecosystems.

ecology↗

bioRxiv2020

In vivo imaging of swimming micromotors using hybrid high-frequency ultrasound and photoacoustic imaging

The fast evolution of medical micro- and nanorobots in the endeavor to perform non-invasive medical operations in living organisms boosted the use of diverse medical imaging techniques in the last years. Among those techniques, photoacoustic (PA) tomography has shown to be promising for the imaging of microrobots in deep-tissue (ex vivo and in vivo), as it possesses the molecular specificity of optical techniques and the penetration depth of ultrasound imaging. However, the precise maneuvering and function control of microrobots, in particular in living organisms, demand the combination of both anatomical and functional imaging methods. Therefore, herein, we report the use of a hybrid High-Frequency Ultrasound (HFUS) and PA imaging system for the real-time tracking of magnetically driven micromotors (single and swarms) in phantoms, ex vivo, and in vivo (in mice bladder and uterus), envisioning their application for targeted drug-delivery.

bioengineering↗

bioRxiv2020

Complementary α-arrestin - Rsp5 ubiquitin ligase complexes control selective nutrient transporter endocytosis in response to amino acid availability

How cells adjust transport across their membranes is incompletely understood. Previously, we have shown that S.cerevisiae broadly re-configures the nutrient transporters at the plasma membrane in response to amino acid availability, through selective endocytosis of sugar- and amino acid transporters (AATs) (Muller et al., 2015). A genome-wide screen now revealed that Art2/Ecm21, a member of the -arrestin family of Rsp5 ubiquitin ligase adaptors, is required for the simultaneous endocytosis of four AATs and induced during starvation by the general amino acid control pathway. Art2 uses a basic patch to recognize C-terminal acidic sorting motifs in these AATs and instructs Rsp5 to ubiquitinate proximal lysine residues. In response to amino acid excess, Rsp5 instead uses TORC1-activated Art1 to detect N-terminal acidic sorting motifs within the same AATs, which initiates exclusive substrate-induced endocytosis of individual AATs. Thus, amino acid availability activates complementary -arrestin-Rsp5-complexes to control selective endocytosis for nutrient acquisition.

cell biology↗

bioRxiv2020

Multiple roles for the ESCRT machinery in maintaining plasma membrane homeostasis

The endosomal sorting complexes required for transport (ESCRT) execute evolutionary conserved membrane remodeling processes. Here we used budding yeast to explore how the ESCRT machinery contributes to plasma membrane (PM) homeostasis. In response to reduced membrane tension and inhibition of the target of rapamycin complex 2 (TORC2), ESCRT-III/Vps4 assemblies form at the PM and help to maintain membrane integrity. Conversely, the growth of ESCRT mutants strongly depends on TORC2-mediated homeostatic regulation of sphingolipid (SL) metabolism. This is caused by calcineurin phosphatase activity which causes Orm2 to accumulate at the endoplasmic reticulum (ER) in ESCRT mutants. Orm2 is a repressor of SL biosynthesis and its accumulation provokes increased membrane stress. This necessitates TORC2 signaling through its downstream kinase Ypk1 to control Orm2 protein levels and prevent a detrimental imbalance of SL metabolism. Our findings reveal new aspects of antagonistic calcineurin/TORC2 signaling for the regulation of SL biosynthesis and the maintenance of PM homeostasis, and suggest that the ESCRT machinery contributes directly and indirectly to these processes.

cell biology↗

bioRxiv2020

Multiplex genetic engineering exploiting pyrimidine salvage pathway-based self-encoded selectable markers

Selectable markers are indispensable for genetic engineering, yet their number and variety is limited. Most selection procedures for prototrophic cells rely on the introduction of antibiotic resistance genes. New minimally invasive tools are needed to facilitate sophisticated genetic manipulations. Here, we characterized three endogenous genes in the human fungal pathogen Aspergillus fumigatus for their potential as markers for targeted genomic insertions of DNAs-of-interest (DOIs). Since these genes are involved in uptake and metabolization of pyrimidines, resistance to the toxic effects of prodrugs 5-fluorocytosine and 5-fluorouracil can be used to select successfully integrated DOIs. We show that DOI integration, resulting in the inactivation of these genes, caused no adverse effects with respect to nutrient requirements, growth or virulence. Beside the individual use of markers for site-directed integration of reporter cassettes including the 17-kb penicillin biosynthetic cluster, we demonstrate their sequential use inserting three fluorescent protein encoding genes into a single strain for simultaneous multicolor localization microscopy. In addition to A. fumigatus, we validated the applicability of this novel toolbox in Penicillium chrysogenum and Fusarium oxysporum. Enabling multiple targeted insertions of DOIs without the necessity for exogenous markers, this technology has the potential to significantly advance genetic engineering.

genetics↗