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Pan-cancer analyses that examine commonalities and differences among various cancer types have emerged as a powerful way to obtain novel insights into cancer biology. Here we present a comprehensive analysis of genetic alterations in a pan-cancer cohort including 961 tumours from children, adolescents, and young adults, comprising 24 distinct molecular types of cancer. Using a standardized workflow, we identified marked differences in terms of mutation frequency and significantly mutated genes in comparison to previously analysed adult cancers. Genetic alterations in 149 putative cancer driver genes separate the tumours into two classes: small mutation and structural/copy-number variant (correlating with germline variants). Structural variants, hyperdiploidy, and chromothripsis are linked to TP53 mutation status and mutational signatures. Our data suggest that 7–8% of the children in this cohort carry an unambiguous predisposing germline variant and that nearly 50% of paediatric neoplasms harbour a potentially druggable event, which is highly relevant for the design of future clinical trials.
The human body is constantly attacked by pathogens. Various lines of defence have evolved, among which the immune system is principal. In contrast to most pathogens, the African trypanosomes thrive freely in the blood circulation, where they escape immune destruction by antigenic variation and incessant motility. These unicellular parasites are flagellate microswimmers that also withstand the harsh mechanical forces prevailing in the bloodstream. They undergo complex developmental cycles in the bloodstream and organs of the mammalian host, as well as the disease-transmitting tsetse fly. Each life cycle stage has been shaped by evolution for manoeuvring in distinct microenvironments. Here, we introduce trypanosomes as blueprints for nature-inspired design of trypanobots, micromachines that, in the future, could explore the human body without affecting its physiology. We review cell biological and biophysical aspects of trypanosome motion. While this could provide a basis for the engineering of microbots, their actuation and control still appear more like fiction than science. Here, we discuss potentials and challenges of trypanosome-inspired microswimmer robots.
The Drosophila microbiome has a limited influence on sleep, activity, and courtship behaviors
(2018)
In animals, commensal microbes modulate various physiological functions, including behavior. While microbiota exposure is required for normal behavior in mammals, it is not known how widely this dependency is present in other animal species. We proposed the hypothesis that the microbiome has a major influence on the behavior of the vinegar fly (Drosophila melanogaster), a major invertebrate model organism. Several assays were used to test the contribution of the microbiome on some well-characterized behaviors: defensive behavior, sleep, locomotion, and courtship in microbe-bearing, control flies and two generations of germ-free animals. None of the behaviors were largely influenced by the absence of a microbiome, and the small or moderate effects were not generalizable between replicates and/or generations. These results refute the hypothesis, indicating that the Drosophila microbiome does not have a major influence over several behaviors fundamental to the animal’s survival and reproduction. The impact of commensal microbes on animal behaviour may not be broadly conserved.
Background:
Genetic heterogeneity and consanguineous marriages make recessive inherited hearing loss in Iran the second most common genetic disorder. Only two reported pathogenic variants (c.323G>C, p.Arg108Pro and c.419A>G, p.Tyr140Cys) in the S1PR2 gene have previously been linked to autosomal recessive hearing loss (DFNB68) in two Pakistani families. We describe a segregating novel homozygous c.323G>A, p.Arg108Gln pathogenic variant in S1PR2 that was identified in four affected individuals from a consanguineous five generation Iranian family.
Methods:
Whole exome sequencing and bioinformatics analysis of 116 hearing loss-associated genes was performed in an affected individual from a five generation Iranian family. Segregation analysis and 3D protein modeling of the p.Arg108 exchange was performed.
Results:
The two Pakistani families previously identified with S1PR2 pathogenic variants presented profound hearing loss that is also observed in the affected Iranian individuals described in the current study. Interestingly, we confirmed mixed hearing loss in one affected individual. 3D protein modeling suggests that the p.Arg108 position plays a key role in ligand receptor interaction, which is disturbed by the p.Arg108Gln change.
Conclusion:
In summary, we report the third overall mutation in S1PR2 and the first report outside the Pakistani population. Furthermore, we describe a novel variant that causes an amino acid exchange (p.Arg108Gln) in the same amino acid residue as one of the previously reported Pakistani families (p.Arg108Pro). This finding emphasizes the importance of the p.Arg108 amino acid in normal hearing and confirms and consolidates the role of S1PR2 in autosomal recessive hearing loss.
Teleost fish provide some of the most intriguing examples of sexually dimorphic coloration, which is often advantageous for only one of the sexes. Mapping studies demonstrated that the genetic loci underlying such color patterns are frequently in tight linkage to the sex-determining locus of a species, ensuring sex-specific expression of the corresponding trait. Several genes affecting color synthesis and pigment cell development have been previously described, but the color loci on the sex chromosomes have mostly remained elusive as yet. Here, we summarize the current knowledge about the genetics of such color loci in teleosts, mainly from studies on poeciliids and cichlids. Further studies on these color loci will certainly provide important insights into the evolution of sex chromosomes.
The fruit fly Drosophila melanogaster possesses approximately 150 brain clock neurons that control circadian behavioral rhythms. Even though individual clock neurons have self-sustaining oscillators, they interact and synchronize with each other through a network. However, little is known regarding the factors responsible for these network interactions. In this study, we investigated the role of CCHamide1 (CCHa1), a neuropeptide expressed in the anterior dorsal neuron 1 (DN1a), in intercellular communication of the clock neurons. We observed that CCHa1 connects the DN1a clock neurons to the ventral lateral clock neurons (LNv) via the CCHa1 receptor, which is a homolog of the gastrin-releasing peptide receptor playing a role in circadian intercellular communications in mammals. CCHa1 knockout or knockdown flies have a generally low activity level with a special reduction of morning activity. In addition, they exhibit advanced morning activity under light-dark cycles and delayed activity under constant dark conditions, which correlates with an advance/delay of PAR domain Protein 1 (PDP1) oscillations in the small-LNv (s-LNv) neurons that control morning activity. The terminals of the s-LNv neurons show rather high levels of Pigment-dispersing factor (PDF) in the evening, when PDF is low in control flies, suggesting that the knockdown of CCHa1 leads to increased PDF release; PDF signals the other clock neurons and evidently increases the amplitude of their PDP1 cycling. A previous study showed that high-amplitude PDP1 cycling increases the siesta of the flies, and indeed, CCHa1 knockout or knockdown flies exhibit a longer siesta than control flies. The DN1a neurons are known to be receptive to PDF signaling from the s-LNv neurons; thus, our results suggest that the DN1a and s-LNv clock neurons are reciprocally coupled via the neuropeptides CCHa1 and PDF, and this interaction fine-tunes the timing of activity and sleep.
TelAP1 links telomere complexes with developmental expression site silencing in African trypanosomes
(2018)
During its life cycle, Trypanosoma brucei shuttles between a mammalian host and the tsetse fly vector. In the mammalian host, immune evasion of T. brucei bloodstream form (BSF) cells relies on antigenic variation, which includes monoallelic expression and periodic switching of variant surface glycoprotein (VSG) genes. The active VSG is transcribed from only 1 of the 15 subtelomeric expression sites (ESs). During differentiation from BSF to the insect-resident procyclic form (PCF), the active ES is transcriptionally silenced. We used mass spectrometry-based interactomics to determine the composition of telomere protein complexes in T. brucei BSF and PCF stages to learn more about the structure and functions of telomeres in trypanosomes. Our data suggest a different telomere complex composition in the two forms of the parasite. One of the novel telomere-associated proteins, TelAP1, forms a complex with telomeric proteins TbTRF, TbRAP1 and TbTIF2 and influences ES silencing kinetics during developmental differentiation.
Optogenetic manipulation of cells or living organisms became widely used in neuroscience following the introduction of the light-gated ion channel channelrhodopsin-2 (ChR2). ChR2 is a non-selective cation channel, ideally suited to depolarize and evoke action potentials in neurons. However, its calcium (Ca2\(^{2+}\)) permeability and single channel conductance are low and for some applications longer-lasting increases in intracellular Ca\(^{2+}\) might be desirable. Moreover, there is need for an efficient light-gated potassium (K\(^{+}\)) channel that can rapidly inhibit spiking in targeted neurons. Considering the importance of Ca\(^{2+}\) and K\(^{+}\) in cell physiology, light-activated Ca\(^{2+}\)-permeant and K\(^{+}\)-specific channels would be welcome additions to the optogenetic toolbox. Here we describe the engineering of novel light-gated Ca\(^{2+}\)-permeant and K\(^{+}\)-specific channels by fusing a bacterial photoactivated adenylyl cyclase to cyclic nucleotide-gated channels with high permeability for Ca\(^{2+}\) or for K\(^{+}\), respectively. Optimized fusion constructs showed strong light-gated conductance in Xenopus laevis oocytes and in rat hippocampal neurons. These constructs could also be used to control the motility of Drosophila melanogaster larvae, when expressed in motoneurons. Illumination led to body contraction when motoneurons expressed the light-sensitive Ca\(^{2+}\)-permeant channel, and to body extension when expressing the light-sensitive K\(^{+}\) channel, both effectively and reversibly paralyzing the larvae. Further optimization of these constructs will be required for application in adult flies since both constructs led to eclosion failure when expressed in motoneurons.
Animal-microbe mutualisms are typically maintained by vertical symbiont transmission or partner choice. A third mechanism, screening of high-quality symbionts, has been predicted in theory, but empirical examples are rare. Here we demonstrate that ambrosia beetles rely on ethanol within host trees for promoting gardens of their fungal symbiont and producing offspring. Ethanol has long been known as the main attractant for many of these fungus-farming beetles as they select host trees in which they excavate tunnels and cultivate fungal gardens. More than 300 attacks by Xylosandrus germanus and other species were triggered by baiting trees with ethanol lures, but none of the foundresses established fungal gardens or produced broods unless tree tissues contained in vivo ethanol resulting from irrigation with ethanol solutions. More X. germanus brood were also produced in a rearing substrate containing ethanol. These benefits are a result of increased food supply via the positive effects of ethanol on food-fungus biomass. Selected Ambrosiella and Raffaelea fungal isolates from ethanol-responsive ambrosia beetles profited directly and indirectly by (i) a higher biomass on medium containing ethanol, (ii) strong alcohol dehydrogenase enzymatic activity, and (iii) a competitive advantage over weedy fungal garden competitors (Aspergillus, Penicillium) that are inhibited by ethanol. As ambrosia fungi both detoxify and produce ethanol, they may maintain the selectivity of their alcohol-rich habitat for their own purpose and that of other ethanol-resistant/producing microbes. This resembles biological screening of beneficial symbionts and a potentially widespread, unstudied benefit of alcohol-producing symbionts (e.g., yeasts) in other microbial symbioses.
Meniscal pathologies are among the most common injuries of the femorotibial joint in both human and equine patients. Pathological forces and ensuing injuries of the cranial horn of the equine medial meniscus are considered analogous to those observed in the human posterior medial horn. Biomechanical properties of human menisci are site-and depth-specific. However, the influence of equine meniscus topography and composition on its biomechanical properties is yet unknown. A better understanding of equine meniscus composition and biomechanics could advance not only veterinary therapies for meniscus degeneration or injuries, but also further substantiate the horse as suitable translational animal model for (human) meniscus tissue engineering. Therefore, the aim of this study was to investigate the composition and structure of the equine knee meniscus in a site-and age-specific manner and their relationship with potential site-specific biomechanical properties. The meniscus architecture was investigated histologically. Biomechanical testing included evaluation of the shore hardness (SH), stiffness and energy loss of the menisci. The SH was found to be subjected to both age and site-specific changes, with an overall higher SH of the tibial meniscus surface and increase in SH with age. Stiffness and energy loss showed neither site nor age related significant differences. The macroscopic and histologic similarities between equine and human menisci described in this study, support continued research in this field.
Meiotic chromosomes undergo rapid prophase movements, which are thought to facilitate the formation of inter-homologue recombination intermediates that underlie synapsis, crossing over and segregation. The meiotic telomere complex (MAJIN, TERB1, TERB2) tethers telomere ends to the nuclear envelope and transmits cytoskeletal forces via the LINC complex to drive these rapid movements. Here, we report the molecular architecture of the meiotic telomere complex through the crystal structure of MAJIN-TERB2, together with light and X-ray scattering studies of wider complexes. The MAJIN-TERB2 2:2 hetero-tetramer binds strongly to DNA and is tethered through long flexible linkers to the inner nuclear membrane and two TRF1-binding 1:1 TERB2-TERB1 complexes. Our complementary structured illumination microscopy studies and biochemical findings reveal a telomere attachment mechanism in which MAJIN-TERB2-TERB1 recruits telomere-bound TRF1, which is then displaced during pachytene, allowing MAJIN-TERB2-TERB1 to bind telomeric DNA and form a mature attachment plate.
Background:
Sex determination relies on a hierarchically structured network of genes, and is one of the most plastic processes in evolution. The evolution of sex-determining genes within a network, by neo- or sub-functionalization, also requires the regulatory landscape to be rewired to accommodate these novel gene functions. We previously showed that in medaka fish, the regulatory landscape of the master male-determining gene dmrt1bY underwent a profound rearrangement, concomitantly with acquiring a dominant position within the sex-determining network. This rewiring was brought about by the exaptation of a transposable element (TE) called Izanagi, which is co-opted to act as a silencer to turn off the dmrt1bY gene after it performed its function in sex determination.
Results:
We now show that a second TE, Rex1, has been incorporated into Izanagi. The insertion of Rex1 brought in a preformed regulatory element for the transcription factor Sox5, which here functions in establishing the temporal and cell-type-specific expression pattern of dmrt1bY. Mutant analysis demonstrates the importance of Sox5 in the gonadal development of medaka, and possibly in mice, in a dmrt1bY-independent manner. Moreover, Sox5 medaka mutants have complete female-to-male sex reversal.
Conclusions:
Our work reveals an unexpected complexity in TE-mediated transcriptional rewiring, with the exaptation of a second TE into a network already rewired by a TE. We also show a dual role for Sox5 during sex determination: first, as an evolutionarily conserved regulator of germ-cell number in medaka, and second, by de novo regulation of dmrt1 transcriptional activity during primary sex determination due to exaptation of the Rex1 transposable element.
Bee population declines are often linked to human impacts, especially habitat and biodiversity loss, but empirical evidence is lacking. To clarify the link between biodiversity loss and bee decline, we examined how floral diversity affects (reproductive) fitness and population growth of a social stingless bee. For the first time, we related available resource diversity and abundance to resource (quality and quantity) intake and colony reproduction, over more than two years. Our results reveal plant diversity as key driver of bee fitness. Social bee colonies were fitter and their populations grew faster in more florally diverse environments due to a continuous supply of food resources. Colonies responded to high plant diversity with increased resource intake and colony food stores. Our findings thus point to biodiversity loss as main reason for the observed bee decline.
Body size is a fundamental trait known to allometrically scale with metabolic rate and therefore a key determinant of individual development, life history, and consequently fitness. In spatially structured environments, movement is an equally important driver of fitness. Because movement is tightly coupled with body size, we expect habitat fragmentation to induce a strong selection pressure on size variation across and within species. Changes in body size distributions are then, in turn, expected to alter food web dynamics. However, no consensus has been reached on how spatial isolation and resource growth affect consumer body size distributions. Our aim was to investigate how these two factors shape the body size distribution of consumers under scenarios of size-dependent and size-independent consumer movement by applying a mechanistic, individual-based resource–consumer model. We also assessed the consequences of altered body size distributions for important ecosystem traits such as resource abundance and consumer stability. Finally, we determined those factors that explain most variation in size distributions. We demonstrate that decreasing connectivity and resource growth select for communities (or populations) consisting of larger species (or individuals) due to strong selection for the ability to move over longer distances if the movement is size-dependent. When including size-dependent movement, intermediate levels of connectivity result in increases in local size diversity. Due to this elevated functional diversity, resource uptake is maximized at the metapopulation or metacommunity level. At these intermediate levels of connectivity, size-dependent movement explains most of the observed variation in size distributions. Interestingly, local and spatial stability of consumer biomass is lowest when isolation and resource growth are high. Finally, we highlight that size-dependent movement is of vital importance for the survival of populations or communities within highly fragmented landscapes. Our results demonstrate that considering size-dependent movement is essential to understand how habitat fragmentation and resource growth shape body size distributions—and the resulting metapopulation or metacommunity dynamics—of consumers.
Background
Gut microbes influence their hosts in many ways, in particular by modulating the impact of diet. These effects have been studied most extensively in humans and mice. In this work, we used whole genome metagenomics to investigate the relationship between the gut metagenomes of dogs, humans, mice, and pigs.
Results
We present a dog gut microbiome gene catalog containing 1,247,405 genes (based on 129 metagenomes and a total of 1.9 terabasepairs of sequencing data). Based on this catalog and taxonomic abundance profiling, we show that the dog microbiome is closer to the human microbiome than the microbiome of either pigs or mice. To investigate this similarity in terms of response to dietary changes, we report on a randomized intervention with two diets (high-protein/low-carbohydrate vs. lower protein/higher carbohydrate). We show that diet has a large and reproducible effect on the dog microbiome, independent of breed or sex. Moreover, the responses were in agreement with those observed in previous human studies.
Conclusions
We conclude that findings in dogs may be predictive of human microbiome results. In particular, a novel finding is that overweight or obese dogs experience larger compositional shifts than lean dogs in response to a high-protein diet.
Single-molecule localization microscopy (SMLM) aims for maximized precision and a high signal-to-noise ratio1. Both features can be provided by placing the emitter in front of a metal-dielectric nanocoating that acts as a tuned mirror2,3,4. Here, we demonstrate that a higher photon yield at a lower background on biocompatible metal-dielectric nanocoatings substantially improves SMLM performance and increases the localization precision by up to a factor of two. The resolution improvement relies solely on easy-to-fabricate nanocoatings on standard glass coverslips and is spectrally and spatially tunable by the layer design and wavelength, as experimentally demonstrated for dual-color SMLM in cells.
Rhodopsin-cyclases for photocontrol of cGMP/cAMP and 2.3 Å structure of the adenylyl cyclase domain
(2018)
The cyclic nucleotides cAMP and cGMP are important second messengers that orchestrate fundamental cellular responses. Here, we present the characterization of the rhodopsinguanylyl cyclase from Catenaria anguillulae (CaRhGC), which produces cGMP in response to green light with a light to dark activity ratio > 1000. After light excitation the putative signaling state forms with tau = 31 ms and decays with tau = 570 ms. Mutations (up to 6) within the nucleotide binding site generate rhodopsin-adenylyl cyclases (CaRhACs) of which the double mutated YFP-CaRhAC (E497K/C566D) is the most suitable for rapid cAMP production in neurons. Furthermore, the crystal structure of the ligand-bound AC domain (2.25 angstrom) reveals detailed information about the nucleotide binding mode within this recently discovered class of enzyme rhodopsin. Both YFP-CaRhGC and YFP-CaRhAC are favorable optogenetic tools for non-invasive, cell-selective, and spatio-temporally precise modulation of cAMP/cGMP with light.
Despite their popularity as enzyme engineering targets structural information about Sucrose Phosphorylases remains scarce. We recently clarified that the Q345F variant of Bifidobacterium adolescentis Sucrose Phosphorylase is able to accept large polyphenolic substrates like resveratrol via a domain shift. Here we present a crystal structure of this variant in a conformation suitable for the accommodation of the donor substrate sucrose in excellent agreement with the wild type structure. Remarkably, this conformation does not feature the previously observed domain shift which is therefore reversible and part of a dynamic process rather than a static phenomenon. This crystallographic snapshot completes our understanding of the catalytic cycle of this useful variant and will allow for a more rational design of further generations of Sucrose Phosphorylase variants.
Forest biodiversity conservation requires precise, area-wide information on the abundance and distribution of key habitat structures at multiple spatial scales. We combined airborne laser scanning (ALS) data with color-infrared (CIR) aerial imagery for identifying individual tree characteristics and quantifying multi-scale habitat requirements using the example of the three-toed woodpecker (Picoides tridactylus) (TTW) in the Bavarian Forest National Park (Germany). This bird, a keystone species of boreal and mountainous forests, is highly reliant on bark beetles dwelling in dead or dying trees. While previous studies showed a positive relationship between the TTW presence and the amount of deadwood as a limiting resource, we hypothesized a unimodal response with a negative effect of very high deadwood amounts and tested for effects of substrate quality. Based on 104 woodpecker presence or absence locations, habitat selection was modelled at four spatial scales reflecting different woodpecker home range sizes. The abundance of standing dead trees was the most important predictor, with an increase in the probability of TTW occurrence up to a threshold of 44–50 dead trees per hectare, followed by a decrease in the probability of occurrence. A positive relationship with the deadwood crown size indicated the importance of fresh deadwood. Remote sensing data allowed both an area-wide prediction of species occurrence and the derivation of ecological threshold values for deadwood quality and quantity for more informed conservation management.
Soft tissue tumors of infancy encompass an overlapping spectrum of diseases that pose unique diagnostic and clinical challenges. We studied genomes and transcriptomes of cryptogenic congenital mesoblastic nephroma (CMN), and extended our findings to five anatomically or histologically related soft tissue tumors: infantile fibrosarcoma (IFS), nephroblastomatosis, Wilms tumor, malignant rhabdoid tumor, and clear cell sarcoma of the kidney. A key finding is recurrent mutation of EGFR in CMN by internal tandem duplication of the kinase domain, thus delineating CMN from other childhood renal tumors. Furthermore, we identify BRAF intragenic rearrangements in CMN and IFS. Collectively these findings reveal novel diagnostic markers and therapeutic strategies and highlight a prominent role of isolated intragenic rearrangements as drivers of infant tumors.
Ras genes are among the most commonly mutated genes in human cancer; yet our understanding of their oncogenic activity at the molecular mechanistic level is incomplete. To identify downstream events that mediate ras-induced cellular transformation in vivo, we analyzed global microRNA expression in three different models of Ras-induction and tumor formation in zebrafish. Six microRNAs were found increased in Ras-induced melanoma, glioma and in an inducible model of ubiquitous Ras expression. The upregulation of the microRNAs depended on the activation of the ERK and AKT pathways and to a lesser extent, on mTOR signaling. Two Ras-induced microRNAs (miR-146a and 193a) target Jmjd6, inducing downregulation of its mRNA and protein levels at the onset of Ras expression during melanoma development. However, at later stages of melanoma progression, jmjd6 levels were found elevated. The dynamic of Jmjd6 levels during progression of melanoma in the zebrafish model suggests that upregulation of the microRNAs targeting Jmjd6 may be part of an anti-cancer response. Indeed, triple transgenic fish engineered to express a microRNA-resistant Jmjd6 from the onset of melanoma have increased tumor burden, higher infiltration of leukocytes and shorter melanoma-free survival. Increased JMJD6 expression is found in several human cancers, including melanoma, suggesting that the up-regulation of Jmjd6 is a critical event in tumor progression.
The following link has been created to allow review of record GSE37015: http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?token=jjcrbiuicyyqgpc&acc=GSE37015.
Fungi possess diverse photosensory proteins that allow them to perceive different light wavelengths and to adapt to changing light conditions in their environment. The biological and physiological roles of the green light-sensing rhodopsins in fungi are not yet resolved. The rice plant pathogen Fusarium fujikuroi exhibits two different rhodopsins, CarO and OpsA. CarO was previously characterized as a light-driven proton pump. We further analyzed the pumping behavior of CarO by patch-clamp experiments. Our data show that CarO pumping activity is strongly augmented in the presence of the plant hormone indole-3-acetic acid and in sodium acetate, in a dose-dependent manner under slightly acidic conditions. By contrast, under these and other tested conditions, the Neurospora rhodopsin (NR)-like rhodopsin OpsA did not exhibit any pump activity. Basic local alignment search tool (BLAST) searches in the genomes of ascomycetes revealed the occurrence of rhodopsin-encoding genes mainly in phyto-associated or phytopathogenic fungi, suggesting a possible correlation of the presence of rhodopsins with fungal ecology. In accordance, rice plants infected with a CarO-deficient F. fujikuroi strain showed more severe bakanae symptoms than the reference strain, indicating a potential role of the CarO rhodopsin in the regulation of plant infection by this fungus.
Species' functional traits set the blueprint for pair-wise interactions in ecological networks. Yet, it is unknown to what extent the functional diversity of plant and animal communities controls network assembly along environmental gradients in real-world ecosystems. Here we address this question with a unique dataset of mutualistic bird-fruit, bird-flower and insect-flower interaction networks and associated functional traits of 200 plant and 282 animal species sampled along broad climate and land-use gradients on Mt. Kilimanjaro. We show that plant functional diversity is mainly limited by precipitation, while animal functional diversity is primarily limited by temperature. Furthermore, shifts in plant and animal functional diversity along the elevational gradient control the niche breadth and partitioning of the respective other trophic level. These findings reveal that climatic constraints on the functional diversity of either plants or animals determine the relative importance of bottom-up and top-down control in plant-animal interaction networks.
Quantitative mass spectrometry has established proteome-wide regulation of protein abundance and post-translational modifications in various biological processes. Here, we used quantitative mass spectrometry to systematically analyze the thermal stability and solubility of proteins on a proteome-wide scale during the eukaryotic cell cycle. We demonstrate pervasive variation of these biophysical parameters with most changes occurring in mitosis and G1. Various cellular pathways and components vary in thermal stability, such as cell-cycle factors, polymerases, and chromatin remodelers. We demonstrate that protein thermal stability serves as a proxy for enzyme activity, DNA binding, and complex formation in situ. Strikingly, a large cohort of intrinsically disordered and mitotically phosphorylated proteins is stabilized and solubilized in mitosis, suggesting a fundamental remodeling of the biophysical environment of the mitotic cell. Our data represent a rich resource for cell, structural, and systems biologists interested in proteome regulation during biological transitions.
Obligate intracellular pathogenic Chlamydia trachomatis express several serine proteases whose roles in chlamydial development and pathogenicity are not completely understood. The chlamydial protease CPAF is expressed during the replicative phase of the chlamydial developmental cycle and is secreted into the lumen of the Chlamydia-containing vacuole called inclusion. How the secreted protease is activated in the inclusion lumen is currently not fully understood. We have identified human serine peptidase inhibitor PI15 as a potential host factor involved in the regulation of CPAF activation. Silencing expression as well as over expression of PI15 affected normal development of Chlamydia. PI15 was transported into the chlamydial inclusion lumen where it co-localized with CPAF aggregates. We show that PI15 binds to the CPAF zymogen and potentially induces CPAF protease activity at low concentrations. However, at high concentrations PI15 inhibits CPAF activity possibly by blocking its protease domain. Our findings shed light on a new aspect of chlamydial host co-evolution which involves the recruitment of host cell proteins into the inclusion to control the activation of bacterial proteases like CPAF that are important for the normal development of Chlamydia.
Increasing evidence indicates that forest disturbances are changing in response to global change, yet local variability in disturbance remains high. We quantified this considerable variability and analyzed whether recent disturbance episodes around the globe were consistently driven by climate, and if human influence modulates patterns of forest disturbance. We combined remote sensing data on recent (2001–2014) disturbances with in-depth local information for 50 protected landscapes and their surroundings across the temperate biome. Disturbance patterns are highly variable, and shaped by variation in disturbance agents and traits of prevailing tree species. However, high disturbance activity is consistently linked to warmer and drier than average conditions across the globe. Disturbances in protected areas are smaller and more complex in shape compared to their surroundings affected by human land use. This signal disappears in areas with high recent natural disturbance activity, underlining the potential of climate-mediated disturbance to transform forest landscapes.
Overwintering temperature and body condition shift emergence dates of spring-emerging solitary bees
(2018)
Solitary bees in seasonal environments must align their life-cycles with favorable environmental conditions and resources; the timing of their emergence is highly fitness relevant. In several bee species, overwintering temperature influences both emergence date and body weight at emergence. High variability in emergence dates among specimens overwintering at the same temperatures suggests that the timing of emergence also depends on individual body conditions. However, possible causes for this variability, such as individual differences in body size or weight, have been rarely studied. In a climate chamber experiment using two spring-emerging mason bees (Osmia cornuta and O. bicornis), we investigated the relationship between temperature, emergence date, body weight, and body size, the last of which is not affected by overwintering temperature. Our study showed that body weight declined during hibernation more strongly in warm than in cold overwintering temperatures. Although bees emerged earlier in warm than in cold overwintering temperatures, at the time of emergence, bees in warm overwintering temperatures had lower body weights than bees in cold overwintering temperatures (exception of male O. cornuta). Among specimens that experienced the same overwintering temperatures, small and light bees emerged later than their larger and heavier conspecifics. Using a simple mechanistic model we demonstrated that spring-emerging solitary bees use a strategic approach and emerge at a date that is most promising for their individual fitness expectations. Our results suggest that warmer overwintering temperatures reduce bee fitness by causing a decrease in body weight at emergence. We showed furthermore that in order to adjust their emergence dates, bees use not only temperature but also their individual body condition as triggers. This may explain differing responses to climate warming within and among bee populations and may have consequences for bee-plant interactions as well as for the persistence of bee populations under climate change.
Background:
The compound eyes of insects allow them to catch photons and convert the energy into electric signals. All compound eyes consist of numerous ommatidia, each comprising a fixed number of photoreceptors. Different ommatidial types are characterized by a specific set of photoreceptors differing in spectral sensitivity. In honey bees, males and females possess different ommatidial types forming distinct retinal mosaics. However, data are lacking on retinal ontogeny and the mechanisms by which the eyes are patterned. In this study, we investigated the intrinsic temporal and circadian expression patterns of the opsins that give rise to the ultraviolet, blue and green sensitive photoreceptors, as well as the morphological maturation of the retina during pupal development of honey bees.
Results:
qPCR and histological labeling revealed that temporal opsin mRNA expression differs between sexes and correlates with rhabdom elongation during photoreceptor development. In the first half of the pupal stage, when the rhabdoms of the photoreceptors are still short, worker and (dorsal) drone retinae exhibit similar expression patterns with relatively high levels of UV (UVop) and only marginal levels of blue (BLop) and green (Lop1) opsin mRNA. In the second half of pupation, when photoreceptors and rhabdoms elongate, opsin expression in workers becomes dominated by Lop1 mRNA. In contrast, the dorsal drone eye shows high expression levels of UVop and BLop mRNA, whereas Lop1 mRNA level decreases. Interestingly, opsin expression levels increase up to 22-fold during early adult life. We also found evidence that opsin expression in adult bees is under the control of the endogenous clock.
Conclusions:
Our data indicate that the formation of the sex-specific retinal composition of photoreceptors takes place during the second half of the pupal development, and that opsin mRNA expression levels continue to increase in young bees, which stands in contrast to Drosophila, where the highest expression levels are found during the late pupal stage and remain constant in adults. From an evolutionary perspective, we hypothesize that the delayed retinal maturation during the early adult phase is linked to the delayed transition from indoor to outdoor activities in bees, when vision becomes important.
The metabolic rewiring that occurs during cell transformation is a hallmark of cancer. It is diverse in different cancers as it reflects different combinations of oncogenic drivers, tumor suppressors, and the microenvironment. Metabolic rewiring is essential to cancer as it enables uncontrolled proliferation and adaptation to the fluctuating availability of nutrients and oxygen caused by poor access to the vasculature due to tumor growth and a foreign microenvironment encountered during metastasis. Increasing evidence now indicates that the metabolic state in cancer cells also plays a causal role in tumor growth and metastasis, for example through the action of oncometabolites, which modulate cell signaling and epigenetic pathways to promote malignancy. In addition to altering the metabolic state in cancer cells, some multifunctional enzymes possess non-metabolic functions that also contribute to cell transformation. Some multifunctional enzymes that are highly expressed in cancer, such as pyruvate kinase M2 (PKM2), have non-canonical functions that are co-opted by oncogenic signaling to drive proliferation and inhibit apoptosis. Other multifunctional enzymes that are frequently downregulated in cancer, such as fructose-bisphosphatase 1 (FBP1), are tumor suppressors, directly opposing mitogenic signaling via their non-canonical functions. In some cases, the enzymatic and non-canonical roles of these enzymes are functionally linked, making the modulation of non-metabolic cellular processes dependent on the metabolic state of the cell.
Drosophila melanogaster is a long-standing model organism in the circadian clock research. A major advantage is the relative small number of about 150 neurons, which built the circadian clock in Drosophila. In our recent work, we focused on the neuroanatomical properties of the lateral neurons of the clock network. By applying the multicolor-labeling technique Flybow we were able to identify the anatomical similarity of the previously described E2 subunit of the evening oscillator of the clock, which is built by the 5th small ventrolateral neuron (5th s-LNv) and one ITP positive dorsolateral neuron (LNd). These two clock neurons share the same spatial and functional properties. We found both neurons innervating the same brain areas with similar pre- and postsynaptic sites in the brain. Here the anatomical findings support their shared function as a main evening oscillator in the clock network like also found in previous studies. A second quite surprising finding addresses the large lateral ventral PDF-neurons (l-LNvs). We could show that the four hardly distinguishable l-LNvs consist of two subgroups with different innervation patterns. While three of the neurons reflect the well-known branching pattern reproduced by PDF immunohistochemistry, one neuron per brain hemisphere has a distinguished innervation profile and is restricted only to the proximal part of the medulla-surface. We named this neuron “extra” l-LNv (l-LNvx). We suggest the anatomical findings reflect different functional properties of the two l-LNv subgroups.
Tropical peat swamp forests sequester globally significant stores of carbon in deep layers of waterlogged, anoxic, acidic and nutrient-depleted peat. The roles of microbes in supporting these forests through the formation of peat, carbon sequestration and nutrient cycling are virtually unknown. This study investigated physicochemical peat properties and microbial diversity between three dominant tree species: Shorea uliginosa (Dipterocarpaceae), Koompassia malaccensis (legumes associated with nitrogen-fixing bacteria), Eleiodoxa conferta (palm) and depths (surface, 45 and 90 cm) using microbial 16S rRNA gene amplicon sequencing. Water pH, oxygen, nitrogen, phosphorus, total phenolic contents and C/N ratio differed significantly between depths, but not tree species. Depth also strongly influenced microbial diversity and composition, while both depth and tree species exhibited significant impact on the archaeal communities. Microbial diversity was highest at the surface, where fresh leaf litter accumulates, and nutrient supply is guaranteed. Nitrogen was the core parameter correlating to microbial communities, but the interactive effects from various environmental variables displayed significant correlation to relative abundance of major microbial groups. Proteobacteria was the dominant phylum and the most abundant genus, Rhodoplanes, might be involved in nitrogen fixation. The most abundant methanogens and methanotrophs affiliated, respectively, to families Methanomassiliicoccaceae and Methylocystaceae. Our results demonstrated diverse microbial communities and provide valuable insights on microbial ecology in these extreme ecosystems.
Ambrosia beetles farm ascomycetous fungi in tunnels within wood. These ambrosia fungi are regarded asexual, although population genetic proof is missing. Here we explored the intraspecific genetic diversity of Ambrosiella grosmanniae and Ambrosiella hartigii (Ascomycota: Microascales), the mutualists of the beetles Xylosandrus germanus and Anisandrus dispar. By sequencing five markers (ITS, LSU, TEF1α, RPB2, β-tubulin) from several fungal strains, we show that X. germanus cultivates the same two clones of A. grosmanniae in the USA and in Europe, whereas A. dispar is associated with a single A. hartigii clone across Europe. This low genetic diversity is consistent with predominantly asexual vertical transmission of Ambrosiella cultivars between beetle generations. This clonal agriculture is a remarkable case of convergence with fungus-farming ants, given that both groups have a completely different ecology and evolutionary history.
The remarkable diversity of sex determination mechanisms known in fish may be fuelled by exceptionally high rates of sex chromosome turnovers or transitions. However, the evolutionary causes and genomic mechanisms underlying this variation and instability are yet to be understood. Here we report on an over 30-year evolutionary experiment in which we tested the genomic consequences of hybridisation and selection between two Xiphophorus fish species with different sex chromosome systems. We find that introgression and imposing selection for pigmentation phenotypes results in the retention of an unexpectedly large maternally derived genomic region. During the hybridisation process, the sex-determining region of the X chromosome from one parental species was translocated to an autosome in the hybrids leading to the evolution of a new sex chromosome. Our results highlight the complexity of factors contributing to patterns observed in hybrid genomes, and we experimentally demonstrate that hybridisation can catalyze rapid evolution of a new sex chromosome.
Background:
The treatment strategies for colorectal cancer located in the right side of the colon have changed dramatically during the last decade. Due to the introduction of complete mesocolic excision (CME) with central ligation of the vessels and systematic lymph node dissection, the long-term survival of affected patients has increased significantly. It has also been proposed that right-sided colon resection can be performed laparoscopically with the same extent of resection and equal long-term results.
Methods:
A retrospective evaluation of a prospectively expanded database on right-sided colorectal cancer or adenoma treated at the University Hospital of Wuerzburg between 2009 and 2016 was performed. All patients underwent CME. This data was analyzed alone and in comparison to the published data describing laparoscopic right-sided colon resection for colon cancer.
Results:
The database contains 279 patients, who underwent right-sided colon resection due to colorectal cancer or colorectal adenoma (255 open; 24 laparoscopic). Operation data (time, length of stay, time on ICU) was equal or superior to laparoscopy, which is comparable to the published results. Surprisingly, the surrogate parameter for correct CME (the number of removed lymph nodes) was significantly higher in the open group. In a subgroup analysis only including patients who were feasible for laparoscopic resection and had been operated with an open procedure by an experienced surgeon, operation time was significantly shorter and the number of removed lymph nodes is significantly higher in the open group.
Conclusion:
So far, several studies demonstrate that laparoscopic right-sided colon resection is comparable to open resection. Our data suggests that a consequent CME during an open operation leads to significantly more removed lymph nodes than in laparoscopically resected patients and in several so far published data of open control groups from Europe. Further prospective randomized trials comparing the long-term outcome are urgently needed before laparoscopy for right-sided colon resection can be recommended ubiquitously.
Crop diversification has been proposed as farm management tool that could mitigate the externalities of conventional farming while reducing productivity-biodiversity trade-offs. Yet evidence for the acclaimed biodiversity benefits of landscape-level crop diversity is ambiguous. Effects may strongly depend on spatial scale and the level of landscape heterogeneity (e.g. overall habitat diversity). At the same time, contrasting within-taxon responses obscure benefits to specific functional groups (i.e. species with shared characteristics or requirements) if studied at the community level. The objectives of this study were to 1) disentangle the relative effects of crop diversity and landscape heterogeneity on avian species richness across five spatial scales ranging from 250 to 3000 m radii around focal winter wheat fields; and 2) assess whether functional groups (feeding guild, conservation status, habitat preference, nesting behaviour) determine the strength and direction of responses to crop diversity and landscape heterogeneity. In central Germany, 14 landscapes were selected along independent gradients of crop diversity (annual arable crops) and landscape heterogeneity. Bird species richness in each landscape was estimated using four point counts throughout the breeding season. We found no effects of landscape-level crop diversity on bird richness and functional groups. Instead, landscape heterogeneity was strongly associated with increased total bird richness across all spatial scales. In particular, insect-feeding and non-farmland birds were favoured in heterogeneous landscapes, as were species not classified as endangered or vulnerable on the regional Red List. Crop-nesting farmland birds, however, were less species-rich in these landscapes. Accordingly, crop diversification may be less suitable for conserving avian diversity and associated ecosystem services (e.g. biological pest control), although confounding interactions with management intensity need yet to be confirmed. In contrast, enhancement of landscape heterogeneity by increasing perennial habitat diversity, reducing field sizes and the amount of cropland has the potential to benefit overall bird richness. Specialist farmland birds, however, may require more targeted management approaches.
Epichloë endophytes associated with cool-season grass species can protect their hosts from herbivory and can suppress mycorrhizal colonization of the hosts’ roots. However, little is known about whether or not Epichloë endophyte infection can also change the foliar fungal assemblages of the host. We tested 52 grassland study sites along a land-use intensity gradient in three study regions over two seasons (spring vs. summer) to determine whether Epichloë infection of the host grass Lolium perenne changes the fungal community structure in leaves. Foliar fungal communities were assessed by Next Generation Sequencing of the ITS rRNA gene region. Fungal community structure was strongly affected by study region and season in our study, while land-use intensity and infection with Epichloë endophytes had no significant effects. We conclude that effects on non-systemic endophytes resulting from land use practices and Epichloë infection reported in other studies were masked by local and seasonal variability in this study’s grassland sites.
It’s a matter of design - how pitfall trap design affects trap samples and possible predictions
(2018)
Background:
Pitfall traps are commonly used to assess ground dwelling arthropod communities. The effects of different pitfall trap designs on the trapping outcome are poorly investigated however they might affect conclusions drawn from pitfall trap data greatly.
Methods:
We tested four pitfall trap types which have been used in previous studies for their effectiveness: a simple type, a faster exchangeable type with an extended plastic rim plate and two types with guidance barriers (V- and X-shaped). About 20 traps were active for 10 weeks and emptied biweekly resulting in 100 trap samples.
Results:
Pitfall traps with guidance barriers were up to five times more effective than simple pitfall traps and trap samples resulted in more similar assemblage approximations. Pitfall traps with extended plastic rim plates did not only perform poorly but also resulted in distinct carabid assemblages with less individuals of small species and a larger variation.
Discussion:
Due to the obvious trait filtering and resulting altered assemblages, we suggest not to use pitfall traps with extended plastic rim plates. In comprehensive biodiversity inventories, a smaller number of pitfall traps with guidance barriers and a larger number of spatial replicates is of advantage, while due to comparability reasons, the use of simple pitfall traps will be recommended in most other cases.
Transforming modern agriculture towards both higher yields and greater sustainability is critical for preserving biodiversity in an increasingly populous and variable world. However, the intensity of agricultural practices varies strongly between crop systems. Given limited research capacity, it is crucial to focus efforts to increase sustainability in the crop systems that need it most. In this study, we investigate the match (or mismatch) between the intensity of pesticide use and the availability of knowledge on the ecosystem service of natural pest control across various crop systems. Using a systematic literature search on pest control and publicly available pesticide data, we find that pest control literature is not more abundant in crops where insecticide input per hectare is highest. Instead, pest control literature is most abundant, with the highest number of studies published, in crops with comparatively low insecticide input per hectare but with high world harvested area. These results suggest that a major increase of interest in agroecological research towards crops with high insecticide input, particularly cotton and horticultural crops such as citrus and high value-added vegetables, would help meet knowledge needs for a timely ecointensification of agriculture.
Apoptosis is a physiological cell death process essential for development, tissue homeostasis, and for immune defense of multicellular animals. Inhibitors of apoptosis proteins (IAPs) regulate apoptosis in response to various cellular assaults. Using both genetic and pharmacological approaches we demonstrate here that the IAPs not only support opportunistic survival of intracellular human pathogens like Chlamydia pneumoniae but also control plasticity of iNOS+ M1 macrophage during the course of infection and render them refractory for immune stimulation. Treatment of Th1 primed macrophages with birinapant (IAP-specific antagonist) inhibited NO generation and relevant proteins involved in innate immune signaling. Accordingly, birinapant promoted hypoxia, angiogenesis, and tumor-induced M2 polarization of iNOS+ M1 macrophages. Interestingly, birinapant-driven changes in immune signaling were accompanied with changes in the expression of various proteins involved in the metabolism, and thus revealing the new role of IAPs in immune metabolic reprogramming in committed macrophages. Taken together, our study reveals the significance of IAP targeting approaches (Smac mimetic compounds) for the management of infectious and inflammatory diseases relying on macrophage plasticity.
The honeybee olfactory pathway comprises an intriguing pattern of convergence and divergence: ~60.000 olfactory sensory neurons (OSN) convey olfactory information on ~900 projection neurons (PN) in the antennal lobe (AL). To transmit this information reliably, PNs employ relatively high spiking frequencies with complex patterns. PNs project via a dual olfactory pathway to the mushroom bodies (MB). This pathway comprises the medial (m-ALT) and the lateral antennal lobe tract (l-ALT). PNs from both tracts transmit information from a wide range of similar odors, but with distinct differences in coding properties. In the MBs, PNs form synapses with many Kenyon cells (KC) that encode odors in a spatially and temporally sparse way. The transformation from complex information coding to sparse coding is a well-known phenomenon in insect olfactory coding. Intrinsic neuronal properties as well as GABAergic inhibition are thought to contribute to this change in odor representation. In the present study, we identified intrinsic neuronal properties promoting coding differences between PNs and KCs using in-situ patch-clamp recordings in the intact brain. We found very prominent K+ currents in KCs clearly differing from the PN currents. This suggests that odor coding differences between PNs and KCs may be caused by differences in their specific ion channel properties. Comparison of ionic currents of m- and l-ALT PNs did not reveal any differences at a qualitative level.
After a large outbreak in Brazil, novel drugs against Zika virus became extremely necessary. Evaluation of virus-based pharmacological strategies concerning essential host factors brought us to the idea that targeting the Axl receptor by blocking its dimerization function could be critical for virus entry. Starting from experimentally validated compounds, such as RU-301, RU-302, warfarin, and R428, we identified a novel compound 2′ (R428 derivative) to be the most potent for this task amongst a number of alternative compounds and leads. The improved affinity of compound 2′ was confirmed by molecular docking as well as molecular dynamics simulation techniques using implicit solvation models. The current study summarizes a new possibility for inhibition of the Axl function as a potential target for future antiviral therapies.
All over the world, pollinators are threatened by land-use change involving degradation of seminatural habitats or conversion into agricultural land. Such disturbance often leads to lowered pollinator abundance and/or diversity, which might reduce crop yield in adjacent agricultural areas. For West Africa, changes in bee communities across disturbance gradients from savanna to agricultural land are mainly unknown. In this study, we monitored for the impact of human disturbance on bee communities in savanna and crop fields. We chose three savanna areas of varying disturbance intensity (low, medium, and high) in the South Sudanian zone of Burkina Faso, based on land-use/land cover data via Landsat images, and selected nearby cotton and sesame fields. During 21 months covering two rainy and two dry seasons in 2014 and 2015, we captured bees using pan traps. Spatial and temporal patterns of bee species abundance, richness, evenness and community structure were assessed. In total, 35,469 bee specimens were caught on 12 savanna sites and 22 fields, comprising 97 species of 32 genera. Bee abundance was highest at intermediate disturbance in the rainy season. Species richness and evenness did not differ significantly. Bee communities at medium and highly disturbed savanna sites comprised only subsets of those at low disturbed sites. An across-habitat spillover of bees (mostly abundant social bee species) from savanna into crop fields was observed during the rainy season when crops are mass-flowering, whereas most savanna plants are not in bloom. Despite disturbance intensification, our findings suggest that wild bee communities can persist in anthropogenic landscapes and that some species even benefitted disproportionally. West African areas of crop production such as for cotton and sesame may serve as important food resources for bee species in times when resources in the savanna are scarce and receive at the same time considerable pollination service.
Aspergillus fumigatus is a saprophytic, cosmopolitan fungus that attacks patients with a weak immune system. A rational solution against fungal infection aims to manipulate fungal metabolism or to block enzymes essential for Aspergillus survival. Here we discuss and compare different bioinformatics approaches to analyze possible targeting strategies on fungal-unique pathways. For instance, phylogenetic analysis reveals fungal targets, while domain analysis allows us to spot minor differences in protein composition between the host and fungi. Moreover, protein networks between host and fungi can be systematically compared by looking at orthologs and exploiting information from host–pathogen interaction databases. Further data—such as knowledge of a three-dimensional structure, gene expression data, or information from calculated metabolic fluxes—refine the search and rapidly put a focus on the best targets for antimycotics. We analyzed several of the best targets for application to structure-based drug design. Finally, we discuss general advantages and limitations in identification of unique fungal pathways and protein targets when applying bioinformatics tools.
HyphaTracker: An ImageJ toolbox for time-resolved analysis of spore germination in filamentous fungi
(2018)
The dynamics of early fungal development and its interference with physiological signals and environmental factors is yet poorly understood. Especially computational analysis tools for the evaluation of the process of early spore germination and germ tube formation are still lacking. For the time-resolved analysis of conidia germination of the filamentous ascomycete Fusarium fujikuroi we developed a straightforward toolbox implemented in ImageJ. It allows for processing of microscopic acquisitions (movies) of conidial germination starting with drift correction and data reduction prior to germling analysis. From the image time series germling related region of interests (ROIs) are extracted, which are analysed for their area, circularity, and timing. ROIs originating from germlings crossing other hyphae or the image boundaries are omitted during analysis. Each conidium/hypha is identified and related to its origin, thus allowing subsequent categorization. The efficiency of HyphaTracker was proofed and the accuracy was tested on simulated germlings at different signal-to-noise ratios. Bright-field microscopic images of conidial germination of rhodopsin-deficient F. fujikuroi mutants and their respective control strains were analysed with HyphaTracker. Consistent with our observation in earlier studies the CarO deficient mutant germinated earlier and grew faster than other, CarO expressing strains.
Many evolutionarily distant pathogenic organisms have evolved similar survival strategies to evade the immune responses of their hosts. These include antigenic variation, through which an infecting organism prevents clearance by periodically altering the identity of proteins that are visible to the immune system of the host1. Antigenic variation requires large reservoirs of immunologically diverse antigen genes, which are often generated through homologous recombination, as well as mechanisms to ensure the expression of one or very few antigens at any given time. Both homologous recombination and gene expression are affected by three-dimensional genome architecture and local DNA accessibility2,3. Factors that link three-dimensional genome architecture, local chromatin conformation and antigenic variation have, to our knowledge, not yet been identified in any organism. One of the major obstacles to studying the role of genome architecture in antigenic variation has been the highly repetitive nature and heterozygosity of antigen-gene arrays, which has precluded complete genome assembly in many pathogens. Here we report the de novo haplotype-specific assembly and scaffolding of the long antigen-gene arrays of the model protozoan parasite Trypanosoma brucei, using long-read sequencing technology and conserved features of chromosome folding4. Genome-wide chromosome conformation capture (Hi-C) reveals a distinct partitioning of the genome, with antigen-encoding subtelomeric regions that are folded into distinct, highly compact compartments. In addition, we performed a range of analyses—Hi-C, fluorescence in situ hybridization, assays for transposase-accessible chromatin using sequencing and single-cell RNA sequencing—that showed that deletion of the histone variants H3.V and H4.V increases antigen-gene clustering, DNA accessibility across sites of antigen expression and switching of the expressed antigen isoform, via homologous recombination. Our analyses identify histone variants as a molecular link between global genome architecture, local chromatin conformation and antigenic variation.
To study delayed genetic and epigenetic radiation effects, which may trigger radiation-induced carcinogenesis, we have established single-cell clones from irradiated and non-irradiated primary human fibroblasts. Stable clones were endowed with the same karyotype in all analyzed metaphases after 20 population doublings (PDs), whereas unstable clones displayed mosaics of normal and abnormal karyotypes. To account for variation in radiation sensitivity, all experiments were performed with two different fibroblast strains. After a single X-ray dose of 2 Gy more than half of the irradiated clones exhibited radiation-induced genome instability (RIGI). Irradiated clones displayed an increased rate of loss of chromosome Y (LOY) and copy number variations (CNVs), compared to controls. CNV breakpoints clustered in specific chromosome regions, in particular 3p14.2 and 7q11.21, coinciding with common fragile sites. CNVs affecting the FHIT gene in FRA3B were observed in independent unstable clones and may drive RIGI. Bisulfite pyrosequencing of control clones and the respective primary culture revealed global hypomethylation of ALU, LINE-1, and alpha-satellite repeats as well as rDNA hypermethylation during in vitro ageing. Irradiated clones showed further reduced ALU and alpha-satellite methylation and increased rDNA methylation, compared to controls. Methylation arrays identified several hundred differentially methylated genes and several enriched pathways associated with in vitro ageing. Methylation changes in 259 genes and the MAP kinase signaling pathway were associated with delayed radiation effects (after 20 PDs). Collectively, our results suggest that both genetic (LOY and CNVs) and epigenetic changes occur in the progeny of exposed cells that were not damaged directly by irradiation, likely contributing to radiation-induced carcinogenesis. We did not observe epigenetic differences between stable and unstable irradiated clones. The fact that the DNA methylation (DNAm) age of clones derived from the same primary culture varied greatly suggests that DNAm age of a single cell (represented by a clone) can be quite different from the DNAm age of a tissue. We propose that DNAm age reflects the emergent property of a large number of individual cells whose respective DNAm ages can be highly variable.
Due to intensive agriculture honeybees are threatened by various pesticides. The use of one group of them, the neonicotinoids, was recently restricted by the European Union. These chemicals bind to the nicotinic acetylcholine receptor (nAchR) in the honeybee brain. Recently, Bayer AG released a new pesticide by the name of “Sivanto” against sucking insects. It is assumed to be harmless for honeybees, although its active ingredient, flupyradifurone, binds nAchR similar to the neonicotinoids. We investigated if this pesticide affects the taste for sugar and cognitive performance in honeybee foragers. These bees are directly exposed to the pesticide while foraging for pollen or nectar. Our results demonstrate that flupyradifurone can reduce taste and appetitive learning performance in honeybees foraging for pollen and nectar, although only the highest concentration had significant effects. Most likely, honeybee foragers will not be exposed to these high concentrations. Therefore, the appropriate use of this pesticide is considered safe for honeybees, at least with respect to the behaviors studied here.
Endogenous molecular circadian clocks drive daily rhythmic changes at the cellular, physiological, and behavioral level for adaptation to and anticipation of environmental signals. The core molecular system consists of autoregulatory feedback loops, where clock proteins inhibit their own transcription. A complex and not fully understood interplay of regulatory proteins influences activity, localization and stability of clock proteins to set the pace of the clock. This study focuses on the molecular function of Ribosomal S6 Kinase (RSK) in the Drosophila melanogaster circadian clock. Mutations in the human rsk2 gene cause Coffin–Lowry syndrome, which is associated with severe mental disabilities. Knock-out studies with Drosophila ortholog rsk uncovered functions in synaptic processes, axonal transport and adult behavior including associative learning and circadian activity. However, the molecular targets of RSK remain elusive. Our experiments provide evidence that RSK acts in the key pace maker neurons as a negative regulator of Shaggy (SGG) kinase activity, which in turn determines timely nuclear entry of the clock proteins Period and Timeless to close the negative feedback loop. Phosphorylation of serine 9 in SGG is mediated by the C-terminal kinase domain of RSK, which is in agreement with previous genetic studies of RSK in the circadian clock but argues against the prevailing view that only the N-terminal kinase domain of RSK proteins carries the effector function. Our data provide a mechanistic explanation how RSK influences the molecular clock and imply SGG S9 phosphorylation by RSK and other kinases as a convergence point for diverse cellular and external stimuli.
The HECT-type ubiquitin ligase HECT, UBA and WWE Domain Containing 1, (HUWE1) regulates key cancer-related pathways, including the Myc oncogene. It affects cell proliferation, stress and immune signaling, mitochondria homeostasis, and cell death. HUWE1 is evolutionarily conserved from Caenorhabditis elegance to Drosophila melanogaster and Humans. Here, we report that the Drosophila ortholog, dHUWE1 (CG8184), is an essential gene whose loss results in embryonic lethality and whose tissue-specific disruption establishes its regulatory role in larval salivary gland development. dHUWE1 is essential for endoreplication of salivary gland cells and its knockdown results in the inability of these cells to replicate DNA. Remarkably, dHUWE1 is a survival factor that prevents premature activation of JNK signaling, thus preventing the disintegration of the salivary gland, which occurs physiologically during pupal stages. This function of dHUWE1 is general, as its inhibitory effect is observed also during eye development and at the organismal level. Epistatic studies revealed that the loss of dHUWE1 is compensated by dMyc proeitn expression or the loss of dmP53. dHUWE1 is therefore a conserved survival factor that regulates organ formation during Drosophila development.
Bees receive nectar and pollen as reward for pollinating plants. Pollen of different plant species varies widely in nutritional composition. In order to select pollen of appropriate nutritional quality, bees would benefit if they could distinguish different pollen types. Whether they rely on visual, olfactory and/or chemotactile cues to distinguish between different pollen types, has however been little studied. In this study, we examined whether and how Apis mellifera workers differentiate between almond and apple pollen. We used differential proboscis extension response conditioning with olfactory and chemotactile stimulation, in light and darkness, and in summer and winter bees. We found that honeybees were only able to differentiate between different pollen types, when they could use both chemotactile and olfactory cues. Visual cues further improved learning performance. Summer bees learned faster than winter bees. Our results thus highlight the importance of multisensory information for pollen discrimination.