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Growth and Differentiation Factor 5 (GDF5) is a secreted growth factor that belongs to the Bone Morphogenetic Protein (BMP) family and plays a pivotal role during limb development. GDF5 is a susceptibility gene for osteoarthritis (OA) and mutations in GDF5 are associated with a wide variety of skeletal malformations ranging from complex syndromes such as acromesomelic chondrodysplasias to isolated forms of brachydactylies or multiple synostoses syndrome 2 (SYNS2). Here, we report on a family with an autosomal dominant inherited combination of SYNS2 and additional brachydactyly type A1 (BDA1) caused by a single point mutation in GDF5 (p.W414R). Functional studies, including chondrogenesis assays with primary mesenchymal cells, luciferase reporter gene assays and Surface Plasmon Resonance analysis, of the GDF5 W-414R variant in comparison to other GDF5 mutations associated with isolated BDA1 (p.R399C) or SYNS2 (p.E491K) revealed a dual pathomechanism characterized by a gain-and loss-of-function at the same time. On the one hand insensitivity to the main GDF5 antagonist NOGGIN (NOG) leads to a GDF5 gain of function and subsequent SYNS2 phenotype. Whereas on the other hand, a reduced signaling activity, specifically via the BMP receptor type IA (BMPR1A), is likely responsible for the BDA1 phenotype. These results demonstrate that one mutation in the overlapping interface of antagonist and receptor binding site in GDF5 can lead to a GDF5 variant with pathophysiological relevance for both, BDA1 and SYNS2 development. Consequently, our study assembles another part of the molecular puzzle of how loss and gain of function mutations in GDF5 affect bone development in hands and feet resulting in specific types of brachydactyly and SYNS2. These novel insights into the biology of GDF5 might also provide further clues on the pathophysiology of OA.
A gene (Imsod) encoding superoxide dismutase (SOD; EC 1.15.1.1) of the facultative intracellular pathogen, Listeria monocytogenes, was cloned by functional complementation of an SOD-deficient Escherichia coli mutant. The nucleotide sequence was determined and the deduced amino acid (aa) sequence (202 aa) showed close similarity to manganese-containing SOD's from other organisms. Subunits of the recombinant L. monocytogenes SOD (re-SOD) and of both E. coli SODs formed enzymatically active hybrid enzymes in vivo. DNA/DNA-hybridization experiments showed that this type of recombinant re-sod gene is conserved within the genus Listeria.
Background: Current imaging methods such as Magnetic Resonance Imaging (MRI), Confocal microscopy, Electron Microscopy (EM) or Selective Plane Illumination Microscopy (SPIM) yield three-dimensional (3D) data sets in need of appropriate computational methods for their analysis. The reconstruction, segmentation and registration are best approached from the 3D representation of the data set. Results: Here we present a platform-independent framework based on Java and Java 3D for accelerated rendering of biological images. Our framework is seamlessly integrated into ImageJ, a free image processing package with a vast collection of community-developed biological image analysis tools. Our framework enriches the ImageJ software libraries with methods that greatly reduce the complexity of developing image analysis tools in an interactive 3D visualization environment. In particular, we provide high-level access to volume rendering, volume editing, surface extraction, and image annotation. The ability to rely on a library that removes the low-level details enables concentrating software development efforts on the algorithm implementation parts. Conclusions: Our framework enables biomedical image software development to be built with 3D visualization capabilities with very little effort. We offer the source code and convenient binary packages along with extensive documentation at http://3dviewer.neurofly.de.
Effects of climate change‐induced events on forest ecosystem dynamics of composition, function and structure call for increased long‐term, interdisciplinary and integrated research on biodiversity indicators, in particular within strictly protected areas with extensive non‐intervention zones. The long‐established concept of forest supersites generally relies on long‐term funds from national agencies and goes beyond the logistic and financial capabilities of state‐ or region‐wide protected area administrations, universities and research institutes.
We introduce the concept of data pools as a smaller‐scale, user‐driven and reasonable alternative to co‐develop remote sensing and forest ecosystem science to validated products, biodiversity indicators and management plans. We demonstrate this concept with the Bohemian Forest Ecosystem Data Pool, which has been established as an interdisciplinary, international data pool within the strictly protected Bavarian Forest and Šumava National Parks and currently comprises 10 active partners. We demonstrate how the structure and impact of the data pool differs from comparable cases.
We assessed the international influence and visibility of the data pool with the help of a systematic literature search and a brief analysis of the results. Results primarily suggest an increase in the impact and visibility of published material during the life span of the data pool, with highest visibilities achieved by research conducted on leaf traits, vegetation phenology and 3D‐based forest inventory.
We conclude that the data pool results in an efficient contribution to the concept of global biodiversity observatory by evolving towards a training platform, functioning as a pool of data and algorithms, directly communicating with management for implementation and providing test fields for feasibility studies on earth observation missions.
Indoor house dust is a blend of organic and inorganic materials, upon which diverse microbial communities such as viruses, bacteria and fungi reside. Adequate moisture in the indoor environment helps microbial communities multiply fast. The outdoor air and materials that are brought into the buildings by airflow, sandstorms, animals pets and house occupants endow the indoor dust particles with extra features that impact human health. Assessment of the health effects of indoor dust particles, the type of indoor microbial inoculants and the secreted enzymes by indoor insects as allergens merit detailed investigation. Here, we discuss the applications of next generation sequencing (NGS) technology which is used to assess microbial diversity and abundance of the indoor dust environments. Likewise, the applications of NGS are discussed to monitor the gene expression profiles of indoor human occupants or their surrogate cellular models when exposed to aqueous solution of collected indoor dust samples. We also highlight the detection methods of dust allergens and analytical procedures that quantify the chemical nature of indoor particulate matter with a potential impact on human health. Our review is thus unique in advocating the applications of interdisciplinary approaches that comprehensively assess the health effects due to bad air quality in built environments.
In recent years, bumblebees have become a prominent insect model organism for a variety of biological disciplines, particularly to investigate learning behaviors as well as visual performance. Understanding these behaviors and their underlying neurobiological principles requires a clear understanding of brain anatomy. Furthermore, to be able to compare neuronal branching patterns across individuals, a common framework is required, which has led to the development of 3D standard brain atlases in most of the neurobiological insect model species. Yet, no bumblebee 3D standard brain atlas has been generated. Here we present a brain atlas for the buff-tailed bumblebee Bombus terrestris using micro-computed tomography (micro-CT) scans as a source for the raw data sets, rather than traditional confocal microscopy, to produce the first ever micro-CT-based insect brain atlas. We illustrate the advantages of the micro-CT technique, namely, identical native resolution in the three cardinal planes and 3D structure being better preserved. Our Bombus terrestris brain atlas consists of 30 neuropils reconstructed from ten individual worker bees, with micro-CT allowing us to segment neuropils completely intact, including the lamina, which is a tissue structure often damaged when dissecting for immunolabeling. Our brain atlas can serve as a platform to facilitate future neuroscience studies in bumblebees and illustrates the advantages of micro-CT for specific applications in insect neuroanatomy.
Stable cell lines are widely used in laboratory research and pharmaceutical industry. They are mainly applied in recombinant protein and antibody productions, gene function studies, drug screens, toxicity assessments, and for cancer therapy investigation. There are two types of cell lines, polyclonal and monoclonal origin, that differ regarding their homogeneity and heterogeneity. Generating a high-quality stable cell line, which can grow continuously and carry a stable genetic modification without alteration is very important for most studies, because polyclonal cell lines of multicellular origin can be highly variable and unstable and lead to inconclusive experimental results. The most commonly used technologies of single cell originate monoclonal stable cell isolation in laboratory are fluorescence-activated cell sorting (FACS) sorting and limiting dilution cloning. Here, we describe a modified limiting dilution method of monoclonal stable cell line selection using the real-time fluorescence imaging system IncuCyte\(^®\)S3.
Background: Teleost fish present a high diversity of sex determination systems, with possible frequent evolutionary turnover of sex chromosomes and sex-determining genes. In order to identify genes involved in male sex determination and differentiation in the platyfish Xiphophorus maculatus, bacterial artificial chromosome contigs from the sex-determining region differentiating the Y from the X chromosome have been assembled and analyzed.
Results: A novel three-copy gene called teximY (for testis-expressed in Xiphophorus maculatus on the Y) was identified on the Y but not on the X chromosome. A highly related sequence called texim1, probably at the origin of the Y-linked genes, as well as three more divergent texim genes were detected in (pseudo) autosomal regions of the platyfish genome. Texim genes, for which no functional data are available so far in any organism, encode predicted esterases/lipases with a SGNH hydrolase domain. Texim proteins are related to proteins from very different origins, including proteins encoded by animal CR1 retrotransposons, animal platelet-activating factor acetylhydrolases (PAFah) and bacterial hydrolases. Texim gene distribution is patchy in animals. Texim sequences were detected in several fish species including killifish, medaka, pufferfish, sea bass, cod and gar, but not in zebrafish. Texim-like genes are also present in Oikopleura (urochordate), Amphioxus (cephalochordate) and sea urchin (echinoderm) but absent from mammals and other tetrapods. Interestingly, texim genes are associated with a Helitron transposon in different fish species but not in urochordates, cephalochordates and echinoderms, suggesting capture and mobilization of an ancestral texim gene in the bony fish lineage. RT-qPCR analyses showed that Y-linked teximY genes are preferentially expressed in testis, with expression at late stages of spermatogenesis (late spermatids and spermatozeugmata).
Conclusions: These observations suggest either that TeximY proteins play a role in Helitron transposition in the male germ line in fish, or that texim genes are spermatogenesis genes mobilized and spread by transposable elements in fish genomes.
Nasal colonization is a major risk factor for S. aureus infections. The mechanisms responsible for colonization are still not well understood and involve several factors on the host and the bacterial side. One key factor is the cell wall teichoic acid (WTA) of S. aureus, which governs direct interactions with nasal epithelial surfaces. We report here the first receptor for the cell wall glycopolymer WTA on nasal epithelial cells. In several assay systems this type F-scavenger receptor, termed SREC-I, bound WTA in a charge dependent manner and mediated adhesion to nasal epithelial cells in vitro. The impact of WTA and SREC-I interaction on epithelial adhesion was especially pronounced under shear stress, which resembles the conditions found in the nasal cavity. Most importantly, we demonstrate here a key role of the WTA-receptor interaction in a cotton rat model of nasal colonization. When we inhibited WTA mediated adhesion with a SREC-I antibody, nasal colonization in the animal model was strongly reduced at the early onset of colonization. More importantly, colonization stayed low over an extended period of 6 days. Therefore we propose targeting of this glycopolymer-receptor interaction as a novel strategy to prevent or control S. aureus nasal colonization.
Zanthoxylum myriacanthum, a small Rutaceous tree growing mainly in secondary hill forests in SE Asia, is a true myrmecophyte. It possesses stem domatia in the form of hollow branches with slitlike openings. Branch hollows and entrance slits are produced by the plant itself through pith degene~.tion ?u.d growth proceSses. If the entrance is not kept open by ants it closes again by growth ol the surrounding tissue after some time. The domatia are colonized opportunistic ally by different arboreous ants, e.g. Crematogaster and Campono tus. Additionally many small extrafloral nectaries are found on the leaflets of Zanthoxylum myriacanthum. Judging from herbarium studies and literature records at least four more true ant trees are found in the genus Zanthoxylum namely Z. rhetsa in SE Asia, Z. conspersipunctatum, Z. pluviatile and Z. vinkii in New Guinea. We could not confirm ant inhabitation in Drypetes pendula (Euphorbiaceae) on the Malay Peninsula, which has also been recorded to be an anttree.
Chronobiological studies of individual activity rhythms in social insects can be constrained by the artificial isolation of individuals from their social context. We present a new experimental set-up that simultaneously measures the temperature rhythm in a queen-less but brood raising mini colony and the walking activity rhythms of singly kept honey bees that have indirect social contact with it. Our approach enables monitoring of individual bees in the social context of a mini colony under controlled laboratory conditions. In a pilot experiment, we show that social contact with the mini colony improves the survival of monitored young individuals and affects locomotor activity patterns of young and old bees. When exposed to conflicting Zeitgebers consisting of a light-dark (LD) cycle that is phase-delayed with respect to the mini colony rhythm, rhythms of young and old bees are socially synchronized with the mini colony rhythm, whereas isolated bees synchronize to the LD cycle. We conclude that the social environment is a stronger Zeitgeber than the LD cycle and that our new experimental set-up is well suited for studying the mechanisms of social entrainment in honey bees.
A new innovative real-time tracking method for flying insects applicable under natural conditions
(2021)
Background
Sixty percent of all species are insects, yet despite global efforts to monitor animal movement patterns, insects are continuously underrepresented. This striking difference between species richness and the number of species monitored is not due to a lack of interest but rather to the lack of technical solutions. Often the accuracy and speed of established tracking methods is not high enough to record behavior and react to it experimentally in real-time, which applies in particular to small flying animals.
Results
Our new method of real-time tracking relates to frequencies of solar radiation which are almost completely absorbed by traveling through the atmosphere. For tracking, photoluminescent tags with a peak emission (1400 nm), which lays in such a region of strong absorption through the atmosphere, were attached to the animals. The photoluminescent properties of passivated lead sulphide quantum dots were responsible for the emission of light by the tags and provide a superb signal-to noise ratio. We developed prototype markers with a weight of 12.5 mg and a diameter of 5 mm. Furthermore, we developed a short wave infrared detection system which can record and determine the position of an animal in a heterogeneous environment with a delay smaller than 10 ms. With this method we were able to track tagged bumblebees as well as hawk moths in a flight arena that was placed outside on a natural meadow.
Conclusion
Our new method eliminates the necessity of a constant or predictable environment for many experimental setups. Furthermore, we postulate that the developed matrix-detector mounted to a multicopter will enable tracking of small flying insects, over medium range distances (>1000m) in the near future because: a) the matrix-detector equipped with an 70 mm interchangeable lens weighs less than 380 g, b) it evaluates the position of an animal in real-time and c) it can directly control and communicate with electronic devices.
Xiphophorus meyeri n. sp. is described as an endemic to Muzquiz, Coahuila, Mexico. It appears to be the northernmost species of the genus. The new species is related to X. couchianus and X. gordoni, but differs morphologically from those by dorsal fin ray number, by the expression of some gonopodial features and most markedly by the appearance of macromelanophores or tr-melanophores.
The synaptonemal complex (SC) is a proteinaceous, meiosis-specific structure that is highly conserved in evolution. During meiosis, the SC mediates synapsis of homologous chromosomes. It is essential for proper recombination and segregation of homologous chromosomes, and therefore for genome haploidization. Mutations in human SC genes can cause infertility. In order to gain a better understanding of the process of SC assembly in a model system that would be relevant for humans, we are investigating meiosis in mice. Here, we report on a newly identified component of the murine SC, which we named SYCE3. SYCE3 is strongly conserved among mammals and localizes to the central element (CE) of the SC. By generating a Syce3 knockout mouse, we found that SYCE3 is required for fertility in both sexes. Loss of SYCE3 blocks synapsis initiation and results in meiotic arrest. In the absence of SYCE3, initiation of meiotic recombination appears to be normal, but its progression is severely impaired resulting in complete absence of MLH1 foci, which are presumed markers of crossovers in wild-type meiocytes. In the process of SC assembly, SYCE3 is required downstream of transverse filament protein SYCP1, but upstream of the other previously described CE–specific proteins. We conclude that SYCE3 enables chromosome loading of the other CE–specific proteins, which in turn would promote synapsis between homologous chromosomes.
A cascade of histone acetylation events with subsequent incorporation of a histone H2A variant plays an essential part in transcription regulation in various model organisms. A key player in this cascade is the chromatin remodelling complex SWR1, which replaces the canonical histone H2A with its variant H2A.Z. Transcriptional regulation of polycistronic transcription units in the unicellular parasite Trypanosoma brucei has been shown to be highly dependent on acetylation of H2A.Z, which is mediated by the histone-acetyltransferase HAT2. The chromatin remodelling complex which mediates H2A.Z incorporation is not known and an SWR1 orthologue in trypanosomes has not yet been reported. In this study, we identified and characterised an SWR1-like remodeller complex in T. brucei that is responsible for Pol II-dependent transcriptional regulation. Bioinformatic analysis of potential SNF2 DEAD/Box helicases, the key component of SWR1 complexes, identified a 1211 amino acids-long protein that exhibits key structural characteristics of the SWR1 subfamily. Systematic protein-protein interaction analysis revealed the existence of a novel complex exhibiting key features of an SWR1-like chromatin remodeller. RNAi-mediated depletion of the ATPase subunit of this complex resulted in a significant reduction of H2A.Z incorporation at transcription start sites and a subsequent decrease of steady-state mRNA levels. Furthermore, depletion of SWR1 and RNA-polymerase II (Pol II) caused massive chromatin condensation. The potential function of several proteins associated with the SWR1-like complex and with HAT2, the key factor of H2A.Z incorporation, is discussed.
A novel chromatin configuration is described in lampbrush chromosomes of Pleurodeles waltlii oocytes which is different from transcriptionally inactive chromatin as weil as from the various forms of transcribed chromatin hitherto described. This novel type of chromatin is not arranged in Christmas tree-Iike configurations of densely packed lateral ribonucleoprotein (RNP) fibriIs but is characterized by a periodic alternating pattern of thick and thin regions which occur in clusters 01 some 10,000 repeats. Each thickened unit with an average length of 45 nm contains two c10sely spaced particles, the putative RNA polymerases, and each thickened unit is separated from the next one by a beaded chromatin spacer with a length of about 80 nm. This chromatin spacer contains on average two particles of approximately 14 nm in diameter, assumed to be nucleosomes. The thickened regions are interpreted to represent short transcriptional units containing approximately 130 base pairs of DNA which are separated from each other by nontranscribed spacers of 240-400 base pairs of DNA. The possibility is discussed that these transcriptional units represent 5S rRNA or tRNA genes.
Many plants combat herbivore and pathogen attack indirectly by attracting predators of their herbivores. Here we describe a novel type of insect-plant interaction where a carnivorous plant uses such an indirect defence to prevent nutrient loss to kleptoparasites. The ant Camponotus schmitzi is an obligate inhabitant of the carnivorous pitcher plant Nepenthes bicalcarata in Borneo. It has recently been suggested that this ant-plant interaction is a nutritional mutualism, but the detailed mechanisms and the origin of the ant-derived nutrient supply have remained unexplained. We confirm that N. bicalcarata host plant leaves naturally have an elevated \(^{15}N/^{14}N\) stable isotope abundance ratio (\(\delta ^{15}N\)) when colonised by C. schmitzi. This indicates that a higher proportion of the plants' nitrogen is insect-derived when C. schmitzi ants are present (ca. 100%, vs. 77% in uncolonised plants) and that more nitrogen is available to them. We demonstrated direct flux of nutrients from the ants to the host plant in a \(^{15}N\) pulse-chase experiment. As C. schmitzi ants only feed on nectar and pitcher contents of their host, the elevated foliar \(\delta ^{15}N\) cannot be explained by classic ant-feeding (myrmecotrophy) but must originate from a higher efficiency of the pitcher traps. We discovered that C. schmitzi ants not only increase the pitchers' capture efficiency by keeping the pitchers' trapping surfaces clean, but they also reduce nutrient loss from the pitchers by predating dipteran pitcher inhabitants (infauna). Consequently, nutrients the pitchers would have otherwise lost via emerging flies become available as ant colony waste. The plants' prey is therefore conserved by the ants. The interaction between C. schmitzi, N. bicalcarata and dipteran pitcher infauna represents a new type of mutualism where animals mitigate the damage by nutrient thieves to a plant.
A nucleolar skeleton of protein filaments demonstrated in amplified nucleoli of Xenopus laevis
(1981)
The amplified, extrachromosomal nucleoli of Xenopus oocytes contain a meshwork of -4-nm-thick filaments, which are densely coiled into higher-order fibrils of diameter 30-40 nm and are resistant to treatment with high- and low-salt concentrations, nucleases (DNase I, pancreatic RNase, micrococcal nuclease), sulfhydryl agents, and various nonionic detergents. This filamentous "skeleton" has been prepared from manually isolated nuclear contents and nucleoli as weil as from nucleoli isolated by fluorescence-activated particle sorting. The nucleolar skeletons are observed in light and electron microscopy and are characterized by ravels of filaments that are especially densely packed in the nucleolar cortex. DNA as weil as RNA are not constituents of this structure, and precursors to ribosomal RNAs are completely removed from the extraction-resistant filaments by treatment with high-salt buffer or RN ase. Fractions of isolated nucleolar skeletons show specific enrichment of an acidic major protein of 145,000 mol wt and an apparent pi value of -6.15, accompanied in some preparations by various amounts of minor proteins. The demonstration of this skeletal structure in "free" extrachromosomal nucleoli excludes the problem of contaminations by nonnucleolar material such as perinucleolar heterochromatin normally encountered in studies of nucleoli from somatic cells. It is suggested that this insoluble protein filament complex forms a skeleton specific to the nucleolus proper that is different from other extraction-resistant components of the nucleus such as matrix and lamina and is involved in the spatial organization of the nucleolar chromatin and its transcriptional products. In studies of the organization of the interphase nucleus, considerable progress has been made in the elucidation of the arrangement of chromatin components and transcriptional products. However, relatively little is known about the composition and function of another category of nuclear structures, the nonnucleoproteinaceous architectural components that are insoluble in solutions of low and high ionic strength, despite numerous studies dedicated to this problem. Such structures include (a) the nuclear envelope and its pore complexes (I, 15, 18, 23, 37, 41), (b) a peripheral layer of insoluble protein ("lamina"; I, 15, 22, 23, 59), (e) certain skeletal proteins related to the chromosome "scaffold" described by Laemmli and coworkers (see references 2 and 3), and (d) ill-defined tangles of fibrillar structures of the nuclear interior that are collectively described as residual "matrix" (6, 21 ; for reviews, see references THE JOURNAL OF CEll BrOlOGY . VOlUME 90 AUGUST 1981 289-299 © The RockefeIler University Press · 0021 -9525/ 81 / 08/ 0289/ 11 $1 .00 4 and 12). The latter, preparatively
A long-range restriction map of part of the short arm of ehromosome 11 including the WAGR region has been constructed using pulsed-field gel electrophoresis and a number of infrequently cutting restriction enzymes. A total of 15.4 Mbp has been mapped in detall, extending from proximal 11p14 to the distal part of 11p12. The map localizes 35 different DNA probes and reveals at least nine areas with features eharaeteristle of BTF islands, some of which may be candidates for the different loci underlying the phenotype of the WAGR syndrome. This map will furthermore allow screening of DNA from individuals with WAGR-related phenotypes and from Wilms tumors for associated chromosomal rearrangements.
Infection research largely relies on classical cell culture or mouse models. Despite having delivered invaluable insights into host-pathogen interactions, both have limitations in translating mechanistic principles to human pathologies. Alternatives can be derived from modern Tissue Engineering approaches, allowing the reconstruction of functional tissue models in vitro. Here, we combined a biological extracellular matrix with primary tissue-derived enteroids to establish an in vitro model of the human small intestinal epithelium exhibiting in vivo-like characteristics. Using the foodborne pathogen Salmonella enterica serovar Typhimurium, we demonstrated the applicability of our model to enteric infection research in the human context. Infection assays coupled to spatio-temporal readouts recapitulated the established key steps of epithelial infection by this pathogen in our model. Besides, we detected the upregulation of olfactomedin 4 in infected cells, a hitherto unrecognized aspect of the host response to Salmonella infection. Together, this primary human small intestinal tissue model fills the gap between simplistic cell culture and animal models of infection, and shall prove valuable in uncovering human-specific features of host-pathogen interplay.