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Institute
- Biowissenschaften (199) (remove)
Background: Nitric oxide synthase 1 adaptor protein (NOS1AP; previously named CAPON) is linked to the glutamatergic postsynaptic density through interaction with neuronal nitric oxide synthase (nNOS). NOS1AP and its interaction with nNOS have been associated with several mental disorders. Despite the high levels of NOS1AP expression in the hippocampus and the relevance of this brain region in glutamatergic signalling as well as mental disorders, a potential role of hippocampal NOS1AP in the pathophysiology of these disorders has not been investigated yet.
Methods: To uncover the function of NOS1AP in hippocampus, we made use of recombinant adeno-associated viruses to overexpress murine full-length NOS1AP or the NOS1AP carboxyterminus in the hippocampus of mice. We investigated these mice for changes in gene expression, neuronal morphology, and relevant behavioural phenotypes.
Findings: We found that hippocampal overexpression of NOS1AP markedly increased the interaction of nNOS with PSD-95, reduced dendritic spine density, and changed dendritic spine morphology at CA1 synapses. At the behavioural level, we observed an impairment in social memory and decreased spatial working memory capacity.
Interpretation: Our data provide a mechanistic explanation for a highly selective and specific contribution of hippocampal NOS1AP and its interaction with the glutamatergic postsynaptic density to cross-disorder pathophysiology. Our findings allude to therapeutic relevance due to the druggability of this molecule.
Cryo-electron tomography is the only technique that can provide sub-nanometer resolved images of cell regions or even whole cells, without the need of labeling or staining methods. Technological advances over the past decade in electron microscope stability, cameras, stage precision and software have resulted in faster acquisition speeds and considerably improved resolution. In pursuit of even better image resolution, researchers seek to reduce noise – a crucial factor affecting the reliability of the tomogram interpretation and ultimately limiting the achieved resolution. Sub-tomogram averaging is the method of choice for reducing noise in repetitive objects. However, when averaging is not applicable, a trade-off between reducing noise and conserving genuine image details must be achieved. Thus, denoising is an important process that improves the interpretability of the tomogram not only directly but also by facilitating other downstream tasks, such as segmentation and 3D visualization. Here, I review contemporary denoising techniques for cryo-electron tomography by taking into account noise-specific properties of both reconstruction and detector noise. The outcomes of different techniques are compared, in order to help researchers select the most appropriate for each dataset and to achieve better and more reliable interpretation of the tomograms.
Trypanosoma cruzi, the causative agent of Chagas disease (American trypanosomiasis), colonizes the intestinal tract of triatomines. Triatomine bugs act as vectors in the life cycle of the parasite and transmit infective parasite stages to animals and humans. Contact of the vector with T. cruzi alters its intestinal microbial composition, which may also affect the associated metabolic patterns of the insect. Earlier studies suggest that the complexity of the triatomine fecal metabolome may play a role in vector competence for different T. cruzi strains. Using high-resolution mass spectrometry and supervised machine learning, we aimed to detect differences in the intestinal metabolome of the triatomine Rhodnius prolixus and predict whether the insect had been exposed to T. cruzi or not based solely upon their metabolic profile. We were able to predict the exposure status of R. prolixus to T. cruzi with accuracies of 93.6%, 94.2% and 91.8% using logistic regression, a random forest classifier and a gradient boosting machine model, respectively. We extracted the most important features in producing the models and identified the major metabolites which assist in positive classification. This work highlights the complex interactions between triatomine vector and parasite including effects on the metabolic signature of the insect.
Aim: The identification of the mechanisms determining spatial variation in biological diversity along elevational gradients is a central objective in ecology and biogeography. Here, we disentangle the direct and indirect effects of abiotic drivers (climatic conditions, and land use) and biotic drivers (vegetation structure and food resources) on functional diversity and composition of bird and bat assemblages along a tropical elevational gradient. Location: Southern slopes of Mt. Kilimanjaro, Tanzania, East Africa. Methods: We counted birds and recorded bat sonotypes on 58 plots distributed in near-natural and anthropogenically modified habitats from 700 to 4,600 m above sea level. For the recorded taxa, we compiled functional traits related to movement, foraging and body size from museum specimens and databases. Further, we recorded mean annual temperature, precipitation, vegetation complexity as well as the number of fruits, flowers, and insect biomass as measures of resource availability on each study site. Results: Using path analyses, we found similar responses of bird and bat functional diversity to the variation in abiotic and biotic drivers along the elevational gradient. In contrast, the functional composition of both taxa showed distinct responses to abiotic and biotic drivers. For both groups, direct temperature effects were most important, followed by resource availability, precipitation and vegetation complexity. Main Conclusions: Our findings indicate that physiological and metabolic constraints imposed by temperature and resource availability determine the functional diversity of bird and bat assemblages, whereas the composition of individual functional traits is driven by taxon-specific processes. Our study illustrates that distinct filtering mechanisms can result in similar patterns of functional diversity along broad environmental gradients. Such differences need to be taken into account when it comes to conserving the functional diversity of flying vertebrates on tropical mountains.
Butyrate production in the acetogen Eubacterium limosum is dependent on the carbon and energy source
(2021)
Eubacterium limosum KIST612 is one of the few acetogenic bacteria that has the genes encoding for butyrate synthesis from acetyl-CoA, and indeed, E. limosum KIST612 is known to produce butyrate from CO but not from H2 + CO2. Butyrate production from CO was only seen in bioreactors with cell recycling or in batch cultures with addition of acetate. Here, we present detailed study on growth of E. limosum KIST612 on different carbon and energy sources with the goal, to find other substrates that lead to butyrate formation. Batch fermentations in serum bottles revealed that acetate was the major product under all conditions investigated. Butyrate formation from the C1 compounds carbon dioxide and hydrogen, carbon monoxide or formate was not observed. However, growth on glucose led to butyrate formation, but only in the stationary growth phase. A maximum of 4.3 mM butyrate was observed, corresponding to a butyrate:glucose ratio of 0.21:1 and a butyrate:acetate ratio of 0.14:1. Interestingly, growth on the C1 substrate methanol also led to butyrate formation in the stationary growth phase with a butyrate:methanol ratio of 0.17:1 and a butyrate:acetate ratio of 0.33:1. Since methanol can be produced chemically from carbon dioxide, this offers the possibility for a combined chemical-biochemical production of butyrate from H2 + CO2 using this acetogenic biocatalyst. With the advent of genetic methods in acetogens, butanol production from methanol maybe possible as well.
The pyruvate:ferredoxin oxidoreductase of the thermophilic acetogen, Thermoanaerobacter kivui
(2021)
Pyruvate:ferredoxin oxidoreductase (PFOR) is a key enzyme in bacterial anaerobic metabolism. Since a low-potential ferredoxin (Fd2−) is used as electron carrier, PFOR allows for hydrogen evolution during heterotrophic growth as well as pyruvate synthesis during lithoautotrophic growth. The thermophilic acetogenic model bacterium Thermoanaerobacter kivui can use both modes of lifestyle, but the nature of the PFOR in this organism was previously unestablished. Here, we have isolated PFOR to apparent homogeneity from cells grown on glucose. Peptide mass fingerprinting revealed that it is encoded by pfor1. PFOR uses pyruvate as an electron donor and methylene blue (1.8 U·mg−1) and ferredoxin (Fd; 27.2 U·mg−1) as electron acceptors, and the reaction is dependent on thiamine pyrophosphate, pyruvate, coenzyme A, and Fd. The pH and temperature optima were 7.5 and 66 °C, respectively. We detected 13.6 mol of iron·mol of protein−1, consistent with the presence of three predicted [4Fe–4S] clusters. The ability to provide reduced Fd makes PFOR an interesting auxiliary enzyme for enzyme assays. To simplify and speed up the purification procedure, we established a protocol for homologous protein production in T. kivui. Therefore, pfor1 was cloned and expressed in T. kivui and the encoded protein containing a genetically engineered His-tag was purified in only two steps to apparent homogeneity. The homologously produced PFOR1 had the same properties as the enzyme from T. kivui. The enzyme can be used as auxiliary enzyme in enzymatic assays that require reduced Fd as electron donor, such as electron-bifurcating enzymes, to keep a constant level of reduced Fd.
In the past two decades, an increasing body of studies has been published on the intersex phenomenon in separate-sexed crustaceans from marine and freshwater ecosystems. Various causes are being considered that could have an influence on the occurrence of intersex. Besides genetic factors, environmental conditions such as photoperiodicity, temperature, salinity and parasitism, but also environmental pollution with endocrine disrupting chemicals (EDCs) are discussed. As part of a long-term monitoring (2012 – 2020) in north-west Brittany, we recorded the occurrence of intersex in the marine amphipod Echinogammarus marinus. We quantified the intersex incidence at marine and estuarine sites and analyzed the incidence in relation to the endocrine potential of the sediments. Intersex occurred with mean frequencies between 0.87% and 12%. It was striking that the incidence of intersex increased with increasing distance from the sea. Since the highest incidence was observed at the range boundary of this stenohaline species, we assume that intersex is triggered by endocrine potential and increasing stress due to increasing freshwater content − and thus an interplay of different environmental factors.
Background: Genome sequencing of all known eukaryotes on Earth promises unprecedented advances in biological sciences and in biodiversity-related applied fields such as environmental management and natural product research. Advances in long-read DNA sequencing make it feasible to generate high-quality genomes for many non–genetic model species. However, long-read sequencing today relies on sizable quantities of high-quality, high molecular weight DNA, which is mostly obtained from fresh tissues. This is a challenge for biodiversity genomics of most metazoan species, which are tiny and need to be preserved immediately after collection. Here we present de novo genomes of 2 species of submillimeter Collembola. For each, we prepared the sequencing library from high molecular weight DNA extracted from a single specimen and using a novel ultra-low input protocol from Pacific Biosciences. This protocol requires a DNA input of only 5 ng, permitted by a whole-genome amplification step.
Results: The 2 assembled genomes have N50 values >5.5 and 8.5 Mb, respectively, and both contain ∼96% of BUSCO genes. Thus, they are highly contiguous and complete. The genomes are supported by an integrative taxonomy approach including placement in a genome-based phylogeny of Collembola and designation of a neotype for 1 of the species. Higher heterozygosity values are recorded in the more mobile species. Both species are devoid of the biosynthetic pathway for β-lactam antibiotics known in several Collembola, confirming the tight correlation of antibiotic synthesis with the species way of life.
Conclusions: It is now possible to generate high-quality genomes from single specimens of minute, field-preserved metazoans, exceeding the minimum contig N50 (1 Mb) required by the Earth BioGenome Project.
Dealing with potential stress in species that have high husbandry requirements, such as elephants, is a challenge for zoos. The objective of the present study was to determine whether positive reinforcement training (PRT) and exposure to a novel object (NOV) for enrichment induced a salivary cortisol response indicative of activation of the hypothalamic–pituitary–adrenal (HPA) axis and which factors determine individual variation in this regard in captive African elephants. We repeatedly sampled the saliva of ten animals (three zoos) for the analysis of cortisol (SACort) before and up to 60 min (in 10–15 min intervals) after the onset of PRT (three repeats) or NOV (nine repeats), which lasted 10 min. There was considerable individual variation in SACort in response to PRT or NOV. Using mixed models, we were able to control these and to reveal that PRT was associated with high SACort before and relatively low SACort after PRT, while NOV induced a moderate SACort increase. The individual differences in SACort were related to age and sex (NOV), while the effects of zoo, handling method (free vs. protected contact) and reproductive and social status were variable. We conclude that positive affective states, such as anticipation or arousal, should be taken into account when interpreting the differences in the SACort responses between PRT and NOV. In addition, understanding the individuality of stress will support management decisions aimed at promoting captive elephant welfare.
The main aim of this thesis work was to elucidate the catalytic mechanism of several enzyme complexes on the basis of their three-dimensional structure. All investigated enzyme complexes occur in the anaerobic energy metabolism and have an essential function by the challenging degradation of aromatic compounds and the flavin-based electron bifurcation (FBEB)/confurcation, an energy-coupling mechanism. More specifically, I studied the phthaloyl-CoA decarboxylase of Thauera chlorobenzoica (Pcd) involved in phthalate ester decomposition, the FBEB protein complexes lactate dehydrogenase/electron-transfer flavoprotein (Ldh/EtfAB) of Acetobacterium woodii, the heterodisulfide-related subunit HdrA of the sulfur- oxidizing bacteria Hyphomicrobium denitrificans (sHdrA). In addition, I contributed to the structure determination of the caffeyl-CoA reductase- EtfAB complex of A. woodii and the naphthoyl-CoA reductase of the sulfate-respiring enrichment culture N47 (mentioned in the Appendix E and F).