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Institute
- Institut für Biochemie und Biologie (193) (remove)
Human induced pluripotent stem cells (hiPSCs) are a promising cell source to generate the patient-specific lung organoid given their superior differentiation potential. However, the current 3D cell culture approach is tedious and time-consuming with a low success rate and high batch-to-batch variability.
Here, we explored the establishment of lung bud organoids by systematically adjusting the initial confluence levels and homogeneity of cell distribution.
The efficiency of single cell seeding and clump seeding was compared. Instead of the traditional 3D culture, we established a 2.5D organoid culture to enable the direct monitoring of the internal structure via microscopy.
It was found that the cell confluence and distribution prior to induction were two key parameters, which strongly affected hiPSC differentiation trajectories. Lung bud organoids with positive expression of NKX 2.1, in a single-cell seeding group with homogeneously distributed hiPSCs at 70% confluence (SC 70% hom) or a clump seeding group with heterogeneously distributed cells at 90% confluence (CL 90% het), can be observed as early as 9 days post induction.
These results suggest that a successful lung bud organoid formation with single-cell seeding of hiPSCs requires a moderate confluence and homogeneous distribution of cells, while high confluence would be a prominent factor to promote the lung organoid formation when seeding hiPSCs as clumps. 2.5D organoids generated with defined culture conditions could become a simple, efficient, and valuable tool facilitating drug screening, disease modeling and personalized medicine.
Three mitochondrial genomes of early-winged insects (Ephemeroptera: Baetidae and Leptophlebiidae)
(2021)
Mayflies (Ephemeroptera) are a semi-aquatic insect order with comparatively few genomic data available despite their phylogenetic position at the root of the winged-insects and possession of ancestral traits.
Here, we provide three mitochondrial genomes (mtgenomes) from representatives of the two most species-rich families, Baetis rutilocylindratus and Cloeon dipterum (Baetidae), and Habrophlebiodes zijinensis (Leptophlebiidae).
All mtgenomes had a complete set of 13 protein-coding genes and a conserved orientation except for two inverted tRNAs in H. zijinensis.
Phylogenetic reconstructions using 21 mayfly mtgenomes and representatives of seven additional orders recovered both Baetidae and Leptophlebiidae as well supported monophyletic clades, with Ephemeroptera as the sister-taxon to all other winged insects (i.e. Odonata and Neoptera).
PC2P
(2021)
Motivation:
Prediction of protein complexes from protein-protein interaction (PPI) networks is an important problem in systems biology, as they control different cellular functions. The existing solutions employ algorithms for network community detection that identify dense subgraphs in PPI networks. However, gold standards in yeast and human indicate that protein complexes can also induce sparse subgraphs, introducing further challenges in protein complex prediction.
Results:
To address this issue, we formalize protein complexes as biclique spanned subgraphs, which include both sparse and dense subgraphs. We then cast the problem of protein complex prediction as a network partitioning into biclique spanned subgraphs with removal of minimum number of edges, called coherent partition. Since finding a coherent partition is a computationally intractable problem, we devise a parameter-free greedy approximation algorithm, termed Protein Complexes from Coherent Partition (PC2P), based on key properties of biclique spanned subgraphs. Through comparison with nine contenders, we demonstrate that PC2P: (i) successfully identifies modular structure in networks, as a prerequisite for protein complex prediction, (ii) outperforms the existing solutions with respect to a composite score of five performance measures on 75% and 100% of the analyzed PPI networks and gold standards in yeast and human, respectively, and (iii,iv) does not compromise GO semantic similarity and enrichment score of the predicted protein complexes. Therefore, our study demonstrates that clustering of networks in terms of biclique spanned subgraphs is a promising framework for detection of complexes in PPI networks.
Sedimentary ancient DNA-based studies have been used to probe centuries of climate and environmental changes and how they affected cyanobacterial assemblages in temperate lakes. Due to cyanobacteria containing potential bloom-forming and toxin-producing taxa, their approximate reconstruction from sediments is crucial, especially in lakes lacking long-term monitoring data. To extend the resolution of sediment record interpretation, we used high-throughput sequencing, amplicon sequence variant (ASV) analysis, and quantitative PCR to compare pelagic cyanobacterial composition to that in sediment traps (collected monthly) and surface sediments in Lake Tiefer See. Cyanobacterial composition, species richness, and evenness was not significantly different among the pelagic depths, sediment traps and surface sediments (p > 0.05), indicating that the cyanobacteria in the sediments reflected the cyanobacterial assemblage in the water column. However, total cyanobacterial abundances (qPCR) decreased from the metalimnion down the water column. The aggregate-forming (Aphanizomenon) and colony-forming taxa (Snowella) showed pronounced sedimentation. In contrast, Planktothrix was only very poorly represented in sediment traps (meta- and hypolimnion) and surface sediments, despite its highest relative abundance at the thermocline (10 m water depth) during periods of lake stratification (May-October). We conclude that this skewed representation in taxonomic abundances reflects taphonomic processes, which should be considered in future DNA-based paleolimnological investigations.
Human induced pluripotent stem cells (hiPSCs) are highly sensitive to extrinsic physical and biochemical signals from their extracellular microenvironments. In this study, we analyzed the effect of cyclic temperature changes on hiPSCs behaviors, especially by means of scanning force microscopy (BIO-AFM). The alternation in cellular mechanics, as well as the secretion and pattern of deposition of extracellular matrix (ECM) protein in hiPSCs were evaluated. The arrangement of the actin cytoskeleton changed with the variation of the temperature. The rearranged cytoskeleton architecture led to the subsequent changes in cell mechanics (Young's modulus of hiPSCs). With the exposure to the cyclic cold stimuli, an increase in the average surface roughness (Ra) and roughness mean square (RMS) was detected. This observation might be at least in part due to the upregulated secretion of Laminin alpha 5 during repeated temporary cooling. The expression of pluripotent markers, NANOG and SOX2, was not impaired in hiPSCs, when exposed to the cyclic cold stimuli for 24 h. Our findings provide an insight into the effect of temperature on the hiPSC behaviors, which may contribute to a better understanding of the application of locally controlled therapeutic hypothermia.
1. Microplastics in soils have become an important threat for terrestrial systems as they may potentially alter the geochemical/biophysical soil environment and can interact with drought. As microplastics may affect soil water content, this could exacerbate the well-known negative effects of drought on ecosystem functionality. Thus, functions including litter decomposition, soil aggregation or those related with nutrient cycling can be altered. Despite this potential interaction, we know relatively little about how microplastics, under different soil water conditions, affect ecosystem functions and multifunctionality.
2. To address this gap, we performed an experiment using grassland plant communities growing in microcosms. Microplastic fibres (absent, present) and soil water conditions (well-watered, drought) were applied in a fully factorial design. At harvest, we measured soil ecosystem functions related to nutrient cycling (beta-glucosaminidase, beta-D-cellobiosidase, phosphatase, beta-glucosidase enzymes), respiration, nutrient retention, pH, litter decomposition and soil aggregation (water stable aggregates). As terrestrial systems provide these functions simultaneously, we also assessed ecosystem multifunctionality, an index that encompasses the array of ecosystem functions measured here.
3. We found that the interaction between microplastic fibres and drought affected ecosystem functions and multifunctionality. Drought had negatively affected nutrient cycling by decreasing enzymatic activities by up to similar to 39%, while microplastics increased soil aggregation by similar to 18%, soil pH by similar to 4% and nutrient retention by up to similar to 70% by diminishing nutrient leaching. Microplastic fibres also impacted soil enzymes, respiration and ecosystem multifunctionality, but importantly, the direction of these effects depended on soil water status. That is, under well-watered conditions, these functions decreased with microplastic fibres by up to similar to 34% while under drought they had similar values irrespective of the microplastic presence, or tended to increase with microplastics. Litter decomposition had a contrary pattern increasing with microplastics by similar to 6% under well-watered conditions while decreasing to a similar percentage under drought.
4. Synthesis and applications. Single ecosystem functions can be positively or negatively affected by microplastics fibres depending on soil water status. However, our results suggest that microplastic fibres may cause negative effects on ecosystem soil multifunctionality of a similar magnitude as drought. Thus, strategies to counteract this new global change factor are necessary.
Full-length transcriptome
(2021)
Fish is considered as a supreme model for clarifying the evolution and regulatory mechanism of vertebrate immunity. However, the knowledge of distinct immune cell populations in fish is still limited, and further development of techniques advancing the identification of fish immune cell populations and their functions are required. Single cell RNA-seq (scRNA-seq) has provided a new approach for effective in-depth identification and characterization of cell subpopulations. Current approaches for scRNA-seq data analysis usually rely on comparison with a reference genome and hence are not suited for samples without any reference genome, which is currently very common in fish research. Here, we present an alternative, i.e. scRNA-seq data analysis with a full-length transcriptome as a reference, and evaluate this approach on samples from Epinephelus coioides-a teleost without any published genome. We show that it reconstructs well most of the present transcripts in the scRNA-seq data achieving a sensitivity equivalent to approaches relying on genome alignments of related species. Based on cell heterogeneity and known markers, we characterized four cell types: T cells, B cells, monocytes/macrophages (Mo/M phi) and NCC (non-specific cytotoxic cells). Further analysis indicated the presence of two subsets of Mo/M phi including M1 and M2 type, as well as four subsets in B cells, i.e. mature B cells, immature B cells, pre B cells and early-pre B cells. Our research will provide new clues for understanding biological characteristics, development and function of immune cell populations of teleost. Furthermore, our approach provides a reliable alternative for scRNA-seq data analysis in teleost for which no reference genome is currently available.
Since the beginning of the Anthropocene, lacustrine biodiversity has been influenced by climate change and human activities. These factors advance the spread of harmful cyanobacteria in lakes around the world, which affects water quality and impairs the aquatic food chain. In this study, we assessed changes in cyanobacterial community dynamics via sedimentary DNA (sedaDNA) from well-dated lake sediments of Lake Tiefer See, which is part of the Klocksin Lake Chain spanning the last 350 years. Our diversity and community analysis revealed that cyanobacterial communities form clusters according to the presence or absence of varves. Based on distance-based redundancy and variation partitioning analyses (dbRDA and VPA) we identified that intensified lake circulation inferred from vegetation openness reconstructions, delta C-13 data (a proxy for varve preservation) and total nitrogen content were abiotic factors that significantly explained the variation in the reconstructed cyanobacterial community from Lake Tiefer See sediments. Operational taxonomic units (OTUs) assigned to Microcystis sp. and Aphanizomenon sp. were identified as potential eutrophication-driven taxa of growing importance since circa common era (ca. CE) 1920 till present. This result is corroborated by a cyanobacteria lipid biomarker analysis. Furthermore, we suggest that stronger lake circulation as indicated by non-varved sediments favoured the deposition of the non-photosynthetic cyanobacteria sister clade Sericytochromatia, whereas lake bottom anoxia as indicated by subrecent- and recent varves favoured the Melainabacteria in sediments. Our findings highlight the potential of high-resolution amplicon sequencing in investigating the dynamics of past cyanobacterial communities in lake sediments and show that lake circulation, anoxic conditions, and human-induced eutrophication are main factors explaining variations in the cyanobacteria community in Lake Tiefer See during the last 350 years.
Wildfires, as a key disturbance in forest ecosystems, are shaping the world's boreal landscapes. Changes in fire regimes are closely linked to a wide array of environmental factors, such as vegetation composition, climate change, and human activity. Arctic and boreal regions and, in particular, Siberian boreal forests are experiencing rising air and ground temperatures with the subsequent degradation of permafrost soils leading to shifts in tree cover and species composition. Compared to the boreal zones of North America or Europe, little is known about how such environmental changes might influence long-term fire regimes in Russia. The larch-dominated eastern Siberian deciduous boreal forests differ markedly from the composition of other boreal forests, yet data about past fire regimes remain sparse. Here, we present a high-resolution macroscopic charcoal record from lacustrine sediments of Lake Khamra (southwest Yakutia, Siberia) spanning the last ca. 2200 years, including information about charcoal particle sizes and morphotypes. Our results reveal a phase of increased charcoal accumulation between 600 and 900 CE, indicative of relatively high amounts of burnt biomass and high fire frequencies. This is followed by an almost 900-year-long period of low charcoal accumulation without significant peaks likely corresponding to cooler climate conditions. After 1750 CE fire frequencies and the relative amount of biomass burnt start to increase again, coinciding with a warming climate and increased anthropogenic land development after Russian colonization. In the 20th century, total charcoal accumulation decreases again to very low levels despite higher fire frequency, potentially reflecting a change in fire management strategies and/or a shift of the fire regime towards more frequent but smaller fires. A similar pattern for different charcoal morphotypes and comparison to a pollen and non-pollen palynomorph (NPP) record from the same sediment core indicate that broad-scale changes in vegetation composition were probably not a major driver of recorded fire regime changes. Instead, the fire regime of the last two millennia at Lake Khamra seems to be controlled mainly by a combination of short-term climate variability and anthropogenic fire ignition and suppression.
Relationships between climate, species composition, and species richness are of particular importance for understanding how boreal ecosystems will respond to ongoing climate change. This study aims to reconstruct changes in terrestrial vegetation composition and taxa richness during the glacial Late Pleistocene and the interglacial Holocene in the sparsely studied southeastern Yakutia (Siberia) by using pollen and sedimentary ancient DNA (sedaDNA) records. Pollen and sedaDNA metabarcoding data using the trnL g and h markers were obtained from a sediment core from Lake Bolshoe Toko. Both proxies were used to reconstruct the vegetation composition, while metabarcoding data were also used to investigate changes in plant taxa richness. The combination of pollen and sedaDNA approaches allows a robust estimation of regional and local past terrestrial vegetation composition around Bolshoe Toko during the last similar to 35,000 years. Both proxies suggest that during the Late Pleistocene, southeastern Siberia was covered by open steppe-tundra dominated by graminoids and forbs with patches of shrubs, confirming that steppe-tundra extended far south in Siberia. Both proxies show disturbance at the transition between the Late Pleistocene and the Holocene suggesting a period with scarce vegetation, changes in the hydrochemical conditions in the lake, and in sedimentation rates. Both proxies document drastic changes in vegetation composition in the early Holocene with an increased number of trees and shrubs and the appearance of new tree taxa in the lake's vicinity. The sedaDNA method suggests that the Late Pleistocene steppe-tundra vegetation supported a higher number of terrestrial plant taxa than the forested Holocene. This could be explained, for example, by the "keystone herbivore" hypothesis, which suggests that Late Pleistocene megaherbivores were able to maintain a high plant diversity. This is discussed in the light of the data with the broadly accepted species-area hypothesis as steppe-tundra covered such an extensive area during the Late Pleistocene.