title: Single-chain and condensed-state behavior of hnRNPA1 from molecular simulations

abstract:
Intrinsically disordered proteins (IDPs) are essential components for the formation of membraneless organelles, which play key functional and regulatory roles within biological systems. These complex assemblies form and dissolve spontaneously over time via liquid-liquid phase separation of IDPs. Mutations in their amino acid sequence can alter their phase behavior, which has been linked to the emergence of severe diseases such as cancer and neurodegenerative diseases including amyotrophic lateral sclerosis. In this work, we study the conformation and phase behavior of a low-complexity domain of heterogeneous nuclear ribonucleoprotein A1 (hnRNPA1), using coarse-grained implicit solvent molecular dynamics simulations. We systematically analyze how these properties are affected by the number of aromatic residues within the examined sequences. We find a significant compaction of the chains and an increase in the critical temperature with increasing number of aromatic residues within the IDPs. Both observations strongly support the hypothesis that aromatic residues play a dominant role for condensation, which is further corroborated by a detailed analysis of the intermolecular contacts. Interestingly, we observe density inhomogeneities within the condensates near criticality, which are driven by electrostatic interactions. Comparing single-chain and condensed state simulations, we find distinct differences in the conformations of individual chains. By establishing quantitative comparisons to the experimental phase behavior, we start to critically assess the reliability of coarse-grained IDP models.

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title: Integrated AlphaFold2 and DEER investigation of the conformational dynamics of a pH-dependent APC antiporter

abstract:
The Amino Acid-Polyamine-Organocation transporter GadC contributes to the survival of pathogenic bacteria under extreme acid stress by exchanging extracellular glutamate for intracellular GABA. Its structure, determined exclusively in an inward-facing conformation at alkaline pH, consists of the canonical LeuT-fold of a conserved five-helix inverted repeat, thereby resembling functionally divergent transporters such as the serotonin reuptake transporter SERT and the glucose-sodium symporter transporter SGLT1. However, despite this structural similarity, it is unclear if the conformational dynamics of antiporters such as GadC follows the blueprint of these or other well-studied LeuT-fold transporters. Here, we used double electron-electron resonance (DEER) spectroscopy to monitor the conformational dynamics of GadC in lipid bilayers in response to acidification and substrate binding. To guide experimental design and facilitate the interpretation of the DEER data, we generated an ensemble of structural models in multiple conformations using a recently introduced AlphaFold2 methodology. Our experimental results reveal acid-induced conformational changes that dislodge the C-terminus from the permeation pathway coupled with rearrangement of helices that enable isomerization between both inward- and outward-facing states. The substrate glutamate, but not GABA, modulates the dynamics of an extracellular thin gate without shifting the equilibrium between inward- and outward-facing conformations. In addition to introducing an integrated methodology for probing transporter conformational dynamics, the congruence of the DEER data with patterns of structural rearrangements deduced from ensembles of AlphaFold2 models illuminate the conformational cycle of GadC underpinning transport and exposes yet another example of the divergence between the dynamics of different functional families in the LeuT-fold.

SIGNIFICANCE STATEMENTThe transporter GadC contributes to acid resistance in bacterial pathogens by exchanging two substrates, glutamate and GABA, using a mechanism termed alternating access. In this study, the conformational dynamics underlying alternating access was studied using a combination of spectroscopy and computational modeling. A conformationally diverse ensemble of models, generated using AlphaFold2, guided the design and interpretation of double electron-electron resonance spectroscopy experiments. We found that whereas GadC was inactive and conformationally homogeneous at neutral pH, low pH induced isomerization between two conformations. From our integrated computational/experimental investigation emerges a transport model that may be relevant to eukaryotic homologs that are involved in other cellular processes.

category: biophysics


title: Genomic analyses of wild and cultivated bacanora agave (Agave angustifolia var. pacifica) reveal inbreeding, few signs of cultivation history and shallow population structure

abstract: Due to the recent increase in demand for agave-based beverages, many wild agave populations have experienced rapid decline and fragmentation; whereas cultivated plants are now managed at monocultural plantations, in some cases involving clonal propagation. We examined the relative effect of migration, genetic drift, natural selection and human activities on the genetic repertoire of Agave angustifolia var. pacifica, an agave used for bacanora (an alcoholic spirit similar to tequila) production in northwestern Mexico. We sampled 34 wild and cultivated sites and used over eleven thousand genome-wide SNPs. We found shallow genetic structure among wild samples, although, detected differentiation between coastal and inland sites. Surprisingly, no differentiation was found between cultivated and wild populations. Moreover, we detected moderate inbreeding (FIS [~] 0.13) and similar levels of genomic diversity in wild and cultivated agaves. Nevertheless, the cultivated plants had almost no private alleles and presented evidence of clonality. The overall low genetic structure in A. angustifolia var. pacifica is apparently the result of high dispersibility promoted by pollinators and possibility of clonal reproduction. Incipient cultivation history and reliance on wild seeds and plants are probably responsible for the observed patterns of high genetic connectivity and considerable diversity in cultivated samples.

category: plant biology


title: Low-efficiency conversion of proliferative glia into induced neurons by Ascl1 in the postnatal mouse cerebral cortex in vivo

abstract: The proneural transcription factor Achaete-scute complex-like 1 (Ascl1) is a major regulator of neural progenitor fate, implicated both in neurogenesis and oligodendrogliogenesis. Ascl1 has been widely used to reprogram non-neuronal cells into induced neurons. In vitro, Ascl1 induces efficient reprogramming of proliferative astroglia from the early postnatal cerebral cortex into interneuron-like cells. Here, we examined whether Ascl1 can similarly induce neuronal reprogramming of glia undergoing proliferation in the postnatal mouse cerebral cortex in vivo. Toward this, we targeted cortical glia at the peak of proliferative expansion (i.e., postnatal day 5) by injecting a retrovirus encoding for Ascl1 into the mouse cerebral cortex. In sharp contrast to the very efficient reprogramming in vitro, Ascl1-transduced glial cells were converted into doublecortin-immunoreactive neurons only with low efficiency in vivo. Interfering with the phosphorylation of Ascl1 by mutation of six conserved proline-directed serine/threonine phosphorylation sites (Ascl1SA6) has been previously shown to increase its neurogenic activity in the early embryonic cerebral cortex. We therefore tested whether transduction of proliferative glia with a retrovirus encoding Ascl1SA6 improved their conversion into neurons. While in vitro glia-to-neuron conversion was markedly enhanced, in vivo reprogramming efficiency remained low. However, both wild-type and mutant Ascl1 reduced the relative number of cells expressing the astrocytic marker glial fibrillary acidic protein (GFAP) and increased the relative number of cells expressing the oligodendroglial marker Sox10 in vivo. Together, our results indicate that the enhanced neurogenic response of proliferative postnatal glia to Ascl1SA6 versus Ascl1 observed in vitro is not recapitulated in vivo.

category: neuroscience


title: In macrophages fatty acid oxidation spares glutamate for use in diverse metabolic pathways required for alternative activation

abstract: Fatty acid oxidation (FAO) is upregulated in IL-4-stimulated (alternatively activated) macrophages (M(IL-4)). We examined the effect of loss of function of the enzyme Cpt1a, which facilitates the entry of long chain fatty acids (FA) into mitochondria for FAO, on alternative activation. Expression of M(IL-4) markers ARG1, CD301 and RELM, was impaired in tamoxifen-treated ERT2Cre x Cpt1afl/fl macrophages and in macrophages expressing shRNA targeting Cpt1a (Cpt1a-shRNA). In contrast, VaviCre x Cpt1afl/fl and LysmCre x Cpt1afl/fl M(IL-4) responded normally to IL-4. Reduced alternative activation due to Cpt1a loss of function was linked to decreased cellular pools of -ketoglutarate, glutamate, and glutathione, diminished commitment of glucose carbon to serine/glycine synthesis, and decreased expression of genes in the Nrf2-oxidative stress response pathway. Consistent with this, reactive oxygen species were increased. Restoration of glutathione pools with N-acetyl cysteine normalized oxidative stress and allowed alternative activation in the face of Cpt1a-deficiency, pointing to a role for FAO in the control of ROS and as being important for alternative activation. In VaviCre x Cpt1afl/fl M(IL-4), glutamine uptake was increased, compensating for the loss of FAO to meet necessary metabolic demands, to allow alternative activation. The data indicate that macrophages are able to regulate glutamine metabolism to compensate for chronic disruption of FAO to meet metabolic needs.

category: immunology


title: Control of N2 fixation and NH3 excretion in Azorhizobium caulinodans ORS571

abstract:
Due to the costly energy demands of N2 fixation, diazotrophic bacteria have evolved complex regulatory networks that permit expression of the N2-fixing catalyst nitrogenase only under conditions of N starvation, whereas the same condition stimulates upregulation of high-affinity NH3 assimilation by glutamine synthetase (GS), preventing excess release of excess NH3 for plants. Diazotrophic bacteria can be engineered to excrete NH3 by interference with GS, however control is required to minimise growth penalties and prevent unintended provision of NH3 to non-target plants. Here, we attempted two strategies to control GS regulation and NH3 excretion in our model cereal symbiont Azorhizobium caulinodans AcLP, a derivative of ORS571. We first attempted to recapitulate previous work where mutation of both PII homologues glnB and glnK stimulated GS shutdown but found that one of these genes was essential for growth. Secondly, we expressed unidirectional adenylyltransferases (uATs) in a {Delta}glnE mutant of AcLP which permitted strong GS shutdown and excretion of NH3 derived from N2 fixation and completely alleviated negative feedback regulation on nitrogenase expression. We placed a uAT allele under control of the NifA-dependent promoter PnifH, permitting GS shutdown and NH3 excretion specifically under microaerobic conditions, the same cue that initiates N2 fixation, then deleted nifA and transferred a rhizopine-inducible nifAL94Q/D95Q-rpoN controller plasmid into this strain, permitting coupled rhizopine-dependent activation of N2 fixation with NH3 excretion. In future, this highly sophisticated and multi-layered control circuitry could be used to activate N2 fixation and NH3 excretion specifically by AcLP colonising transgenic rhizopine producing cereals, targeting delivery of fixed N to the crop, and preventing interaction with non-target plants.

Author SummaryInoculation of cereal crops with associative "diazotrophic" bacteria that convert atmospheric N2 to NH3 could be used to sustainably improve delivery of nitrogen in agriculture. However, due to the costly energy demands of N2 fixation, natural diazotrophic bacteria have evolved to conserve energy by preventing excess production of NH3 and release to the plants. Diazotrophs can be engineered for excess NH3 production and release, however genetic control is required to minimise growth penalties and prevent unintended provision of NH3 to non-target weed species. Here, we engineer control of N2 fixation and NH3 excretion in response to the signalling molecule rhizopine which is produced by transgenic barley. This control could be used to establish plant host-specific activation of N2 fixation and NH3 release following root colonisation in the field, minimising bacterial energy requirements in the bulk soil and preventing provision of NH3 to non-target plants.

category: microbiology


title: Form follows function: Variable microtubule architecture in the malaria parasite

abstract: The malaria parasite undergoes a series of extensive morphological changes within its human host and mosquito vector. A scaffold of microtubules beneath a peripheral double membrane establishes and maintains the distinct shapes of all infectious forms, but the underlying structural basis remains unknown. Here we applied in situ electron cryo-tomography after focused ion beam milling to follow changes in the microtubule cytoskeleton throughout the Plasmodium life cycle. This revealed an unexpected level of structural and architectural diversity so far not observed in other organisms. Microtubules in migrating mosquito forms consist of 13 protofilaments reinforced by interrupted luminal helices. Conversely, gametocyte microtubules consist of 13 to 18 protofilaments with doublets, triplets and quadruplets of varying arrangements. We show the microtubule cytoskeleton within the native cellular context, highlighting structurally diverse apical rings which act as microtubule organising centres. This provides a unique view into a relevant human pathogen with an unusual microtubule cytoskeleton.


title: Heavy metal pollution impacts soil bacterial community structure and antimicrobial resistance at the Birmingham 35th Avenue Superfund Site

abstract: Heavy metals (HMs) are known to modify bacterial communities both in the laboratory and in situ. As a consequence, soils in HM contaminated sites like the U.S. Environmental Protection Agency (EPA) Superfund sites are predicted to have altered ecosystem functioning, with potential ramifications for the health of organisms, including humans, that live nearby. Further, several studies have shown that metal tolerant bacteria are often also resistant to antimicrobial agents (AMR), and therefore HM contaminated soils could potentially act as reservoirs that could disseminate AMR genes into human-associated pathogenic bacteria. To explore this possibility soil samples were collected from six public locations in the zip code 35207 (the home of the North Birmingham 35th Avenue Superfund site) and in six public areas in a neighboring reference zip code (35214). Sequencing of the V4 region of the bacterial 16S rRNA gene revealed that elevated concentrations of HMs As, Mn, Pb, and Zn reduced microbial diversity and altered community structure within each zip code. While there was no difference between zip codes in the proportion of total culturable microbes that survived antimicrobial or metal exposure, bacterial isolates with HMR almost always also exhibited AMR. Metagenomes inferred using PICRUST2 also predicted significantly higher mean relative frequencies in 35207 for several AMR genes related to both specific and broad-spectrum AMR phenotypes. Together, these results support the hypothesis that chronic HM pollution alters soil bacterial community structure in ecologically meaningful ways and may also select for bacteria with increased potential to contribute to AMR in human bacterial disease.


title: Extensive diversity and rapid turnover of phage defense repertoires in cheese-associated bacterial communities

abstract:
Phages are key drivers of genomic diversity in bacterial populations as they impose strong selective pressure on the evolution of bacterial defense mechanisms across closely related strains. The pan-immunity model suggests that such diversity is maintained because the effective immune system of a bacterial species is the one distributed across all strains present in the community. However, only few studies have analyzed the distribution of bacterial defense systems at the community-level, mostly focusing on CRISPR and comparing samples from complex environments. Here, we studied 2778 bacterial genomes and 158 metagenomes from cheese-associated communities, which are dominated by a few bacterial taxa and occur in relatively stable environments. We find that nearly identical strains of cheese-associated bacteria contain diverse and highly variable arsenals of innate and adaptive (i.e CRISPR-Cas) immunity suggesting rapid turnover of defense mechanisms in these communities. CRISPR spacer abundance correlated with the abundance of matching target sequences across the metagenomes providing evidence that the identified defense repertoires are functional and under selection. While these characteristics align with the pan-immunity model, the detected CRISPR spacers only covered a subset of the phages previously identified in cheese, suggesting that CRISPR does not provide complete immunity against all phages, and that the innate immune mechanisms may have complementary roles. Our findings show that the evolution of bacterial defense mechanisms is a highly dynamic process and highlight that experimentally tractable, low complexity communities such as those found in cheese, can help to understand ecological and molecular processes underlying phage-defense system relationships.

ImportanceBacteria are constantly exposed to phage predation and hence harbor highly diverse defense arsenals. According to the pan-immunity hypothesis the effective immune system of a bacterial species is not the one encoded in a single genome but in the entire community. However, few studies have investigated how defense systems are distributed within communities. Here, we carried out (meta)genomic analyses of bacterial communities used in cheesemaking. These are tractable communities of biotechnological interest which house few bacterial species and are exposed to high phage pressure. In line with the pan-immunity hypothesis, we find that nearly identical strains of cheese-associated bacteria contain highly variable arsenals of innate and adaptive immunity. We provide evidence for the functional importance of this diversity, and reveal that CRISPR alone does not provide complete immunity against all phages. Our findings can have implications for the design of robust synthetic communities used in biotechnology and the food industry.


title: Multiplex genome editing of human pluripotent stem cells using Cpf1

abstract: Targeted genome editing of human pluripotent stem cells (hPSC) is critical for basic and translational research, and can be achieved with site-specific endonucleases. Class 2 clustered regularly interspaced short palindromic repeats (CRISPR) Cpf1 represents a class of RNA programmable DNA endonucleases with features that could be complementary to the widely used Class 1 CRISPR-Cas9 system. Here we engineered a single vector system to deliver both Cpf1 and guide RNA, and achieved efficient knockin in transformed human cells. We then adopted the system to function in hPSCs by increasing expression levels of Cpf1, and showed that Cpf1 efficiently mediate homology directed repair in multiple loci (including knockin of a 6 kb cassette at the INS) in multiple hPSC lines, resulting in clones with transgenic cassettes integrated only at the targeted loci in the genome. Furthermore, with delivery of Cpf1 and a single U6 promoter-driven guide RNA array composed of an AAVS1-targeting guide and a MAFB-targeting guide, we demonstrated efficient multiplex genome editing in hPSCs at the AAVS1 (knockin) and MAFB (knockout) loci. Targeted hPSCs expressed pluripotency markers, and could be directed to differentiate to pancreatic {beta}-like cells, neural progenitor cells, and neurons in vitro. Generated INS reporter cells differentiated into {beta}-like cells that express tdTomato and luciferase specifically in insulin-expressing cells, allowing for in vivo tracking of human {beta}-like cells in humanized mice. By targeted screening of potential off-target sequences with most homology to guide RNA targeted sequences, we could not detect off-target mutations for all guide RNAs at the condition of efficient genome editing with Cpf1 in hPSCs. This work provides a system complementary to Cas9 for potentially precise genome editing in hPSCs.

category: molecular biology


title: Spatiotemporal proximity labeling tools to track GlcNAc sugar-modified functional protein hubs during cellular signaling

abstract:
A fundamental mechanism that all eukaryotic cells use to adapt to their environment is dynamic protein modification with monosaccharide sugars. In humans, O-linked N-acetylglucosamine (O-GlcNAc) is rapidly added to and removed from diverse protein sites as a response to fluctuating nutrient levels, stressors, and signaling cues. Two aspects remain challenging for tracking functional O-GlcNAc events with chemical strategies: spatial control over subcellular locations and time control during labeling. The objective of this study was to create intracellular proximity labeling tools to identify functional changes in O-GlcNAc patterns with spatiotemporal control. We developed a labeling strategy based on the TurboID proximity labeling system for rapid protein biotin conjugation that we directed to O-GlcNAc protein modifications inside cells, a set of tools we called "GlycoID." Localized variants to the nucleus and cytosol, nuc-GlycoID and cyt-GlycoID, labeled O-GlcNAc proteins and their interactomes in subcellular space. Labeling during insulin as well as serum stimulation revealed functional changes in O-GlcNAc proteins as soon as 30 minutes of signaling. We demonstrated using proteomic analysis that the GlycoID strategy captured O-GlcNAcylated "activity hubs" consisting of O-GlcNAc proteins and their associated protein-protein interactions. The ability to follow changes in O-GlcNAc hubs during physiological events like insulin stimulation poises these tools to be used for determining mechanisms of glycobiological cell regulation. Our functional O-GlcNAc datasets in human cells will be a useful resource for O-GlcNAc-driven mechanisms.

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category: cell biology


title: Evolutionary Origin of Vertebrate OCT4/POU5 Functions in Supporting Pluripotency

abstract: The support of pluripotent cells over time is an essential feature of development. In eutherian embryos, pluripotency is maintained from naive states in peri-implantation to primed pluripotency at gastrulation. To understand how these states emerged, we reconstruct the evolutionary trajectory of the Pou5 gene family, which contains the central pluripotency factor OCT4. By coupling evolutionary sequence analysis with functional studies in mouse Embryonic Stem Cells (ESCs), we found that the ability of POU5 proteins to support pluripotency originated in the gnathostome lineage, prior to the generation of two paralogues, Pou5f1 and Pou5f3 via gene duplication. In osteichthyans, retaining both genes, the paralogues differ in their support of naive and primed pluripotency. This specialization of these duplicates enables the diversification of function in self-renewal and differentiation. By integrating sequence evolution, ESC phenotypes, developmental contexts and structural modelling, we pinpoint OCT4 regions sufficient for naive pluripotency and describe their adaptation over evolutionary time.

category: developmental biology


title: Profiling the size-dependent heterogeneity of membrane proteins in a mixed population of small extracellular vesicle for potential cancer diagnosis

abstract: The heterogeneity in small extracellular vesicles (small EVs) introduces an extra level of complexity in small EV-based liquid biopsy for cancer diagnosis. Heterogeneous membrane protein expression is correlated with sizes of small EVs, but accessing this correlative information is limited by the precise isolation of size-dependent subpopulations. Herein, we present a single EV enumeration (SEVEN) approach to profile protein heterogeneity in size-dependent subpopulations, and demonstrate its potential in improving the accuracy of cancer diagnosis. The interferometric plasmonic microscopy (iPM) capable of imaging single biological nanoparticles with the diameter down to 30 nm is employed to detect small EVs at the single-particle level. Small EVs population with mixed sizes are directly imaged, individually sized and digitally counted during their binding onto different aptamer-coated iPM sensor surfaces. The protein expression levels and binding kinetics of three size-dependent subpopulations are analyzed, forming a multidimensional data matrix for cancer diagnosis. Using small EVs derived from different cancer cell lines, highly heterogeneous protein profiles are recorded in the three subpopulations. We further demonstrate that the cancer classification accuracy could be greatly improved by including the subpopulation level heterogeneous protein profiles as compared with conventional ensemble measurement.

category: biochemistry


title: Is It A Barrel? General Features of Transmembrane Beta Barrels from a Large Database

abstract: Large datasets for particular classes of proteins can contribute a birds-eye view that is more accurate than that gathered from studying particular exemplars. We used co-evolution data in the form of predicted contact maps to create a large, high-quality database of transmembrane {beta}-barrels (TMBB). We classify sequences for our database using IsItABarrel, an approach combining topology prediction and rule-based TMBB classification. By applying simple feature detection on generated contact maps, our method achieves 95.88% balanced accuracy when discriminating this structural fold from other protein classes. Our database contains almost two million bacterial TMBB proteins from 20 classes, respectively 17 and 2.2 times larger than the previous sets TMBB-DB and OMPdb. Moreover, comparison with IsItABarrel revealed a high rate of false positives in previous TMBB algorithms. We assessed the TMBB-content of 2,959 proteomes of bacterial organisms across several taxonomic categories and find tremendous variance among TMBB-containing organisms with some having as many as 300 TMBBs (6.79% of proteome) and others as few as 10 (0.27% of proteome). The distribution of the lengths of the TMBBs is suggestive of previously hypothesized duplication events. In addition, we find that the C-terminal {beta}-signal varies among different classes of bacteria though it is most commonly GHyGHyG{varphi}+{varphi}. The database is available through our IsItABarrel webapp and we anticipate that its quality and size will serve as a useful resource where high quality TMBB sequence data is required.

category: bioinformatics


title: Mutations in DNA polymerase δ subunit 1 mediate CMD2-type resistance to Cassava Mosaic Geminiviruses

abstract: Cassava mosaic disease suppresses cassava yields across the tropics. The dominant CMD2 locus confers resistance to the cassava mosaic geminiviruses. It has been reported that CMD2-type landraces lose resistance after regeneration through de novo morphogenesis. As full genome bisulfite sequencing failed to uncover an epigenetic mechanism for loss of resistance, we performed whole genome sequencing and genetic variant analysis and fine-mapped the CMD2 locus to a 190 kilobase interval. Data suggest that CMD2-type resistance is caused by a nonsynonymous, single nucleotide polymorphism in DNA polymerase {delta} subunit 1 (MePOLD1) located within this region. Virus-induced gene silencing of MePOLD1 in a Cassava mosaic disease-susceptible cassava variety produced a recovery phenotype typical of CMD2-type resistance. Analysis of other CMD2-type cassava varieties identified additional resistance alleles within MePOLD1. MePOLD1 resistance alleles represent important genetic resources for resistance breeding or genome editing, and elucidating mechanisms of resistance to geminiviruses.


title: Evaluation of oral baits and distribution methods for Tasmanian devils (Sarcophilus harrisii)

abstract:
ContextDiseases are increasingly contributing to wildlife population declines. Tasmanian devil (Sarcophilus harrisii) populations have locally declined by 82% largely due to the morbidity and mortality associated with two independent transmissible devil facial tumours (DFT1 and DFT2). Toxic baits are often used as a management tool for controlling vertebrate pest populations in Australia, but in other areas of the world oral baits are also used to deliver vaccines or pharmaceuticals to wildlife. Oral vaccine bait products have been distributed for more than 50 years at a landscape scale to protect wildlife from rabies virus and contributed to the elimination of fox rabies from more than ten European nations. An oral bait vaccine to protect against devil facial tumours has been proposed as a management tool to improve the population health, resiliency and fitness of wild Tasmanian devils.

AimOur goal was to evaluate the potential use of edible baits as vehicles for vaccine delivery to Tasmanian devils.

MethodWe tested placebo versions of baits that are already used in Australia.

Key resultsCaptive devils consumed all types of placebo baits but exhibited a preference for ruminant- and fish-based baits. Captive devils also consumed inert capsules inserted into placebo baits. Bait fate trials in the field revealed that baits were generally removed within 24 hours. Tasmanian pademelons (Thylogale billardierii), brushtail possums (Trichosurus vulpecula), and Eastern quolls (Dasyurus viverrinus) were the most common nontarget bait competitors at six private properties in southern Tasmania; wild devils removed approximately 5% of ground baits at these sites. We also evaluated an automated bait dispenser, which reduced nontarget uptake and resulted in over 50% of the baits being removed by devils.

ConclusionsThis study demonstrates that captive and wild devils will accept and consume placebo versions of commercial baits. Bait dispensers or modified baits or baiting strategies are needed to increase bait uptake by devils.

ImplicationsBait dispensers can be used at a regional scale to deliver baits to Tasmanian devils. These could act as vehicles for preventive or therapeutic vaccines to mitigate the impacts of disease on devil populations.

Short summaryThis study aimed to test oral baits as potential vaccine delivery vehicles for Tasmanian devils. Captive and wild devils consumed placebo versions of commercial baits used on mainland Australia. Abundant non-target species, such as brushtail possums, Tasmanian pademelons, and eastern quolls consumed most baits in the wild. Implementation of automated bait dispensers increased bait uptake by devils to over 50% at the same regional field sites.

category: zoology


title: Catecholaminergic cell type-specific expression of Cre recombinase in knock-in transgenic rats generated by the Combi-CRISPR technology

abstract:
BackgroundCell groups containing catecholamines provide a useful model to study the molecular and cellular mechanisms underlying the morphogenesis, physiology, and pathology of the central nervous system. For this purpose, it is necessary to establish a system to induce catecholaminergic group-specific expression of Cre recombinase. Recently, we introduced a gene cassette encoding 2A peptide fused to Cre recombinase into the site between the C-terminus and translational termination codons of the rat tyrosine hydroxylase (TH) open reading frame by the Combi-CRISPR technology, which is a genomic editing method to enable an efficient knock-in (KI) of long DNA sequence into a target site. However, the expression patterns of the transgene and its function as well as the effect of the mutation on the biochemical and behavioral phenotypes in the KI strains have not been characterized yet.

New MethodWe aimed to evaluate the usefulness of TH-Cre KI rats as an experimental model for investigating the structure and function of catecholaminergic neurons in the brain.

ResultsWe detected cell type-specific expression of Cre recombinase and site-specific recombination activity in the representative catecholaminergic groups in the TH-Cre KI rat strains. In addition, we measured TH expression level and catecholamine accumulation in the brain regions, and spontaneous locomotion, indicating that catecholamine metabolism and general behavior are apparently normal in these KI rats.

ConclusionsTH-Cre KI rat strains produced by the Combi-CRISPR system offer a beneficial model to study the molecular and cellular mechanics for the morphogenesis, physiology, and pathology of catecholamine-containing neurons in the brain.


title: Genomic analysis of population structure, antimicrobial resistance, and virulent factors of methicillin-resistant Macrococcus caseolyticus in global lineages

abstract:
Macrococcus caseolyticus is an opportunistic pathogen frequently detected in dairy products as well as veterinary infections. The present study examined the population structure, antimicrobial resistance, and virulent factors of methicillin-resistant M. caseolyticus isolates in retail meat from Shanghai (n=10) and global isolates from GenBank (n=87). All strains were divided into five lineages that distributed in Europe (82.4%, n = 80), Asia (11.3%, n = 11), North America (4.1%, n = 4), Oceania (1%, n = 1) and Africa (1%, n = 1). MLST typing revealed novel alleles in Chinese M. caseolyticus strains. Furthermore, a total of 24 AMR genes associated with 10 classes of antimicrobial agents were identified in the isolates from global lineages, carried by dominant plasmids such as rep7a, rep22 and repUS56. Comparing to other lineages, genomes from the Chinese lineage carried significantly more AMR genes (p<0.005) and less virulent factors (p<0.001), which may be explained by the local evolution of M. caseolyticus in China. Finally, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were enrolled for morphology comparison between M. caseolyticus and S. aureus, showing that M. caseolyticus has a larger diameter and thicker cell wall. The present study showed geographical variation with regards to MLST profiles, antimicrobial resistance, and virulent factors in global M. caseolyticus lineages. This study suggests that such local evolution of foodborne or livestock origin M. caseolyticus may serve as vehicles for domestic transmission of methicillin resistance in retail meat in China.

HighlightsO_LIGlobal M. caseolyticus strains were divided into five lineages from A to E.
C_LIO_LIMLST typing revealed novel alleles in Chinese M. caseolyticus strains.
C_LIO_LIChinese lineage carried significantly more AMR genes and less virulent factors.
C_LIO_LIMacrococcus caseolyticus has a larger diameter and thicker cell wall compared with S.aureus.
C_LIO_LIMacrococcus caseolyticus may enhance the domestic transmission of methicillin resistance in China.
C_LI


title: The transcription factor RXRA regulates lipid metabolism in duck myoblasts by the CD36 network

abstract: Lipid metabolism is a highly complex metabolic process regulated at multiple levels. RXRA is a well-characterized factor that regulates lipid metabolism. To explore the function and mechanism of the transcription factor RXRA in myoblasts, and to further explore the key factors that RXRA regulates the target gene CD36 signaling network to regulate lipid metabolism. We found that the transcription factor RXRA inhibited the accumulation of triglycerides (TGs), cholesterol (CHOL) and non-esterified fatty acids (NEFA) in CS2 cells by regulating CD36 as well as the fatty acid beta oxidation pathway. CD36 functions similar to RXRA in myoblasts. CD36 overexpression reduced the levels of TGs, CHOL, NEFAs and saturated fatty acids (SFAs) in these cells, while CD36 knockout increased the levels of TGs, CHOL, NEFAs, SFAs and monounsaturated fatty acids (MUPAs) in these cells. GRB2, MAP1B, SLC38A1, SLC4A7, NCOA3, PKIA, MOB2, SAA2 and RXRA are involved in the CD36 promotion of lipid efflux through lipid metabolism, endocytosis and amino acid metabolism. Considering these results, we propose that RXRA regulates lipid metabolism in myoblasts by regulating the CD36 signaling network.


title: Molecular basis of indispensable accuracy of mammalian miRNA biogenesis

abstract: Mammalian Dicer is the gatekeeper into the essential gene-regulating miRNA pathway. What is committing mammalian Dicer to the miRNA pathway remains unknown. We report that Dicers highly conserved DExD/H helicase domain is the key structural element supporting accurate miRNA biogenesis. While ATPase activity of the domain is non-essential, its loss is lethal in mice. It is required during canonical miRNA biogenesis for efficient recognition, high-fidelity cleavage, and strand selection. Structure of Dicer-miRNA precursor complexes showed that the DExD/H domain acquired helicase-unrelated function defining Dicer conformations, which affect substrate loading and facilitate pre-selection of miRNA precursors. Dicer lacking the DExD/H domain favors conformations enabling reduced substrate selectivity and supporting RNA interference, a different small RNA pathway. Therefore, Dicers DExD/H domain ensures indispensable high-fidelity precursor processing of mammalian miRNAs, which constitutes a structural "mold" for adaptive miRNA evolution.


title: Generation and characterization of iPSC lines (UOHi001-A, UOHi002-A)From a patient with SHANK3 mutation and her healthy mother

abstract: Phelan-McDermid syndrome (PMS) is a rare genetic condition that causes global developmental disability, delayed or absent speech, and autism spectrum disorder. The loss of function of one copy of SHANK3, which codes for a scaffolding protein found in the postsynaptic density of synapses, has been identified as the main cause of PMS. We report the generation and characterization of two induced pluripotent stem cell (iPSC) lines derived from one patient with a SHANK3 mutation and the patients mother as a control. Both lines expressed pluripotency markers, differentiated into the three germ layers, retained the disease-causing mutation, and displayed normal karyotypes.


title: ATF6 Activation Reduces Amyloidogenic Transthyretin Secretion Through Increased Interactions with Endoplasmic Reticulum Proteostasis Factors

abstract: The extracellular aggregation of destabilized transthyretin (TTR) variants is implicated in the onset and pathogenesis of familial TTR-related amyloid diseases. One strategy to reduce toxic, extracellular aggregation of TTR is to decrease the population of aggregation-prone protein secreted from mammalian cells. Stress-independent activation of the unfolded protein response (UPR)-associated transcription factor ATF6 preferentially decreases secretion and subsequent aggregation of destabilized, aggregation-prone TTR variants. However, the mechanism of this reduced secretion was previously undefined. Here, we implement a mass spectrometry-based interactomics approach to identify endoplasmic reticulum (ER) proteostasis factors involved in ATF6-dependent reductions of destabilized TTR secretion. We show that ATF6 activation reduces amyloidogenic TTR secretion and subsequent aggregation through a mechanism involving ER retention that is mediated by increased interactions with ATF6-regulated ER proteostasis factors including BiP and PDIA4. Intriguingly, the PDIA4-dependent retention of TTR is independent of both the single TTR cysteine residue and the redox activity of PDIA4, indicating that PDIA4 retains destabilized TTR in the ER through a redox-independent chaperoning activity. Our results define a mechanistic basis to explain the ATF6 activation-dependent reduction of destabilized, amyloidogenic TTR secretion that could be therapeutically accessed to improve treatments of TTR-related amyloid diseases.


title: Microbial landscape ecology: landscape structure better predicts soil protist diversity than local edaphic variables in alpine meadows

abstract:
ContextHuman-induced changes in landscape structure are among the main causes of biodiversity loss. However, despite their important contribution to biodiversity and ecosystem functioning, microbes - and particularly protists - remain spatially understudied. Furthermore, soil microbiota are most often related to the local soil properties, whereas the influence of the surrounding landscape is rarely assessed, calling for microbial landscape ecology studies.

ObjectivesHere, we assessed for the first time the effect of changes in landscape composition and configuration on soil protist diversity in meadows in the Swiss western Alps.

MethodsWe sampled 178 plots along an elevation gradient representing a broad range of climatic, edaphic, and land use conditions, and measured landscape structures in several neighbourhood windows of increasing size.

ResultsOur results showed that landscape structures between 500 m and 2000 m around plot centres better predicted microbial diversity than local edaphic and climatic descriptors, with the percentage cover of open habitat and meadows and, to a lesser extent, edge density being the most important descriptors. The patch level descriptors explained the least amount of variation in protist diversity, what is in contradiction with macro-organims where patch size and shape strongly impact the population dynamics. This study also highlights different responses of protist functional groups to changes in landscape structure.

ConclusionsOur results importantly show the significance of landscape structure for microbial diversity and the potentially negative consequences of land use change on soil microbial communities and their associated functions.

category: ecology


title: N-terminal helices and A domain of archaeal FtsY facilitate SRP54 binding and the association with cell membrane

abstract: The process of protein translocation is essential to the maintenance of cellular life and has been critically addressed in eukaryotes and bacteria. However, little information is available regarding protein translocation across archaeal membranes. The signal recognition particle (SRP) plays an important role in this process. It binds the signal peptide at the N-terminus of the polypeptide chain and interacts with the cognate SRP receptor (FtsY) located on the target membrane to form a targeting complex (TC). Concomitant GTP hydrolysis by SRP and FtsY delivers the polypeptide to the adjacent protein-conducting channel. The present study aims to characterize the structural domains of FtsY contributing to the targeting complex (TC) formation in Sulfolobus acidocaldarius, a thermo-acidophilic crenarchaeon. The contacting residues between SRP54 and FtsY were mapped along the N1-N3 helices. Interestingly, the previously reported crystal structure did not take the N-terminal A domain into account - a region rich in negatively charged residues. Such observation led us to investigate the contribution of each of the three participating helices (N1-3) in terms of membrane association and functional TC formation. Through biophysical analyses of SRP-FtsY and FtsY-membrane interaction, and biochemical characterization of the reciprocal GTPase activity, this work sought to elucidate the minimal structural motif controlling the archaeal TC assembly.


title: Efficient and sensitive profiling of RNA-protein interactions using TLC-CLIP

abstract: RNA-binding proteins are instrumental for post-transcriptional gene regulation, yet transcriptomewide methods to profile RNA-protein interactions remain technically challenging. We present an improved library preparation strategy for crosslinking and immunoprecipitation (CLIP) that involves tailing and ligation of cDNA molecules (TLC) for increased sensitivity and efficiency. TLC-CLIP eliminates time-consumingpurifications, reduces sample loss and minimises experimental steps, allowing precise profiling of RNA-protein interactions from limited starting material at nucleotide resolution.


title: BCL-xL antagonizes the deleterious effects of KRAS on mitochondrial scaffolding

abstract: In addition to its canonical role as a regulator of mitochondrial outer membrane permeabilization, BCL-xL exerts diverse non canonical functions contributing to cancer cell aggressiveness. In particular it regulates KRAS intracellular activation levels. We herein explored the mechanistic basis for this effect by a spatially restricted biotin-labelling proteomic approach designed to characterize proteins whose proximity to KRAS, used as a bait, is BCL-xL dependant. BCL-xL loss relocalizes KRAS to the vicinity of mitochondrial proteins. Proximal proteins include the mitochondrial scaffold prohibitin 2 (PHB2), which also interacts with BCL-xL and the downregulation of which prevents BCL-xL sensitive effects of KRAS induced contacts between mitochondria and endosomes, and mitochondrial mass decrease. These results argue that BCL-xL prevents a negative feedback regulation of KRAS canonical signaling by KRAS interference with mitochondrial quality control.


title: Attention Samples Items in Visual Working Memory Rhythmically

abstract: Attention allows us to selectively enhance the processing of specific locations or features in our external environment while simultaneously filtering out momentarily irrelevant information. It is currently hypothesized that this is achieved through the boosting of relevant sensory signals which biases the competition between competing neural representations. Recent neurophysiological and behavioral studies have revealed that attention is a fundamentally rhythmic process, tightly linked to neural oscillations in fronto-parietal networks. Instead of continuously highlighting a single object or location, attention rhythmically alternates between multiple relevant representations at a frequency of 3 - 6 Hz. However attention can not only be directed towards the external world but also towards internal visual working memory (VWM) representations, e.g. when selecting one of several search templates to find corresponding objects in the external world. Two recent studies have revealed that objects in VWM are attended in a similarly rhythmic fashion as perceived objects. We add to the current literature by showing that retro-cues towards multi-feature gratings in VWM initiate a similar theta-rhythmic competition, modulating reaction times in an anti-phasic manner. Our findings add to the converging body of evidence that external and internal visual representations are accessed by highly similar, rhythmic attentional mechanisms.


title: Effects of local versus global competition on reproductive skew and sex differences in social dominance behaviour

abstract: Social hierarchies are often found in group-living animals. The hierarchy position can influence reproductive success (RS), with a skew towards high-ranking individuals. The amount of aggression in social dominance varies greatly, both between species and between males and females within species. Using game theory we study this variation by taking into account the degree to which reproductive competition in a social group is mainly local to the group, emphasising within-group relative RS, or global to a larger population, emphasising an individuals absolute RS. Our model is similar to recent approaches in that reinforcement learning is used as a behavioural mechanism allowing social-hierarchy formation. We test two hypotheses. The first is that local competition should favour the evolution of mating or foraging interference, and thus of reproductive skew. Second, decreases in reproductive output caused by an individuals accumulated fighting damage, such as reduced parenting ability, will favour less intense aggression but should have little influence on reproductive skew. From individual-based simulations of the evolution of social dominance and interference, we find support for both hypotheses. We discuss to what extent our results can explain observed sex differences in reproductive skew and social dominance behaviour.

category: animal behavior and cognition


title: The cilia enriched oxysterol 7β,27-DHC is required for polycystin activation

abstract: PC-1 and PC-2 form a heteromeric ion channel complex (hereafter called the Polycystin complex) that is abundantly expressed on primary cilia of renal epithelial cells. Mutations within the polycystin complex cause Autosomal Dominant Polycystic Kidney Disease (ADPKD). The Polycystin complex forms a non-selective cation channel, yet the spatial and temporal regulation of the polycystin complex within the ciliary membrane remains poorly understood, partially due to technical limitations posed by the tiny ciliary compartment. Here, we employ our novel assays to functionally reconstitute the polycystin complex in the plasma membrane. Using whole-cell and ciliary patch-clamp recordings we identified a ciliary enriched oxysterol, 7{beta},27-DHC, as a critical component required for activation of the polycystin complex. We identified a novel oxysterol binding pocket in PC-2 using molecular docking simulation. We also identified two amino acids within the PC-2 oxysterol binding pocket, E208 and R581, to be critical for 7{beta},27-DHC dependent polycystin activation in both the plasma membrane and ciliary compartment. Further, we can show that the pharmacological and genetic inhibition of oxysterol synthesis by carbenoxolone (CNX) reduces channel activity in primary cilia. Our findings identified a unique second messenger that regulates the polycystin complex. We hypothesize that cilia-enriched lipids license the polycystin complex to be functional only in the ciliary organelle, thus providing novel insights into the spatial regulation of the polycystin complex. Our results also establish a framework to target the same allosteric regulatory site in the polycystin complex to identify activators of the polycystin channels as novel therapeutic strategies for ADPKD.

category: physiology


title: Multiphasic calcium signatures in the cytosol and nucleus of Lotus japonicus roots unlock the early discrimination of fungal signals

abstract:
Changes in intracellular calcium concentration are known to mediate plant responses to both beneficial and detrimental microorganisms. Increasing evidence suggests that boundaries between early signalling events in plant pathogenesis and symbiosis are less defined than previously thought, given the occurrence of Ca2+ signals partially sharing subcellular location and dynamics.

In this study we used differentially targeted aequorin-based Ca2+ reporters to measure cytosolic and nuclear Ca2+ signatures triggered in Lotus japonicus roots by putative symbiotic and pathogenic fungal molecules.

By using mutants impaired in the Common Symbiotic Signalling Pathway, we dissected the multiphasic nature of the stimulus-specific Ca2+ transients, highlighting significant differences in the genetic components underlying the cytosolic and nuclear Ca2+ response to short-chain (CO4), long-chain (CO8) and lipidated (mycLCO) chitin oligomers. Moreover, a pharmacological approach shed light on the origin, modulation and interdependence of the observed Ca2+ fluxes. Our analysis of the overall plant root response in terms of cytosolic and nuclear Ca2+ changes helps disentangling the complex communication circuits in the pre-contact signal exchange phase and enhances our quantitative understanding of how plants can perceive and discriminate different fungal molecules.


title: Building a Tissue-unbiased Brain Template of Fibre Orientation Distribution and Tractography with Multimodal Registration

abstract: A brain template provides a standard space for statistical analysis of brain structure and function. For decades, the T1- and T2-weighted brain templates have been widely used for brain grey matter anatomical and functional analysis. However, T1- and T2-weighted templates provide very limited information about the axonal organization within the white matter. Recent advances in Diffusion MRI have enabled the detailed modelling of the axonal fibre orientation distribution (FOD) in white matter. Therefore, building a FOD template is essential for more robust white matter anatomy related analysis; however, it is important that this template aligns well with the cortical and subcortical structures. From such a FOD template, a tractography template can be also generated by fibre tracking algorithms, which can be used for subsequent applications, such as to perform the joint structural and functional analysis while ensuring rigorous fibre-to-fibre correspondence. In this paper, we explore the potential of generating the FOD template based on multimodal registration, in order to constrain the tempalte unbiased to both white and grey matter. We combine the information from T1-weighted, T2-weighted and Diffusion MRI to generate a coherent transformation for FOD registration and template generation. Our FOD template preserves the structural details at the white-grey matter boundary. To illustrate the benefit of this new approach, the resulting tractography template was used for joint structural-functional connectivity analysis.


title: Prenatal development of neonatal vocalizations

abstract: Human and non-human primates produce rhythmical sounds as soon as they are born. These early vocalizations are important for soliciting the attention of caregivers. How they develop, remains a mystery. The orofacial movements necessary for producing these vocalizations have distinct spatiotemporal signatures. Therefore, their development could potentially be tracked over the course of prenatal life. We densely and longitudinally sampled fetal head and orofacial movements in marmoset monkeys using ultrasound imaging. We show that orofacial movements necessary for producing rhythmical vocalizations differentiate from a larger movement pattern that includes the entire head. We also show that signature features of marmoset infant contact calls emerge prenatally as a distinct pattern of orofacial movements. Our results establish that aspects of the sensorimotor development necessary for vocalizing occur prenatally, even before the production of sound.


title: Multi-omics analyses cannot identify true-positive novel associations from underpowered genome-wide association studies of four brain-related traits

abstract:
BackgroundThe integration of multi-omics information (e.g., epigenetics and transcriptomics) can be useful for interpreting findings from genome-wide association studies (GWAS). It has additionally been suggested that multi-omics may aid in novel variant discovery, thus circumventing the need to increase GWAS sample sizes. We tested whether incorporating multi-omics information in earlier and smaller sized GWAS boosts true-positive discovery of genes that were later revealed by larger GWAS of the same/similar traits.

MethodsWe applied ten different analytic approaches to integrating multi-omics data from twelve sources (e.g., Genotype-Tissue Expression project) to test whether earlier and smaller GWAS of 4 brain-related traits (i.e., alcohol use disorder/problematic alcohol use [AUD/PAU], major depression [MDD], schizophrenia [SCZ], and intracranial volume [ICV]) could detect genes that were revealed by a later and larger GWAS.

ResultsMulti-omics data did not reliably identify novel genes in earlier less powered GWAS (PPV<0.2; 80% false-positive associations). Machine learning predictions marginally increased the number of identified novel genes, correctly identifying 1-8 additional genes, but only for well-powered early GWAS of highly heritable traits (i.e., ICV and SCZ). Multi-omics, particularly positional mapping (i.e., fastBAT, MAGMA, and H-MAGMA), was useful for prioritizing genes within genome-wide significant loci (PPVs = 0.5 - 1.0).

ConclusionsAlthough the integration of multi-omics information, particularly when multiple methods agree, helps prioritize GWAS findings and translate them into information about disease biology, it does not substantively increase novel gene discovery in brain-related GWAS. To increase power for discovery of novel genes and loci, increasing sample size is a requirement.

category: genetics


title: Moss functional traits are important drivers for moss and underlying soil bacterial communities: evidence from a chronosequence in an Icelandic glacier forefield

abstract:
Mosses are among the first colonizing organisms after glacier retreat and can develop into thick moss mats during later successional stages. They are key players in N2 fixation through their microbiome, which is an important process for nutrient build-up during primary succession. How these moss-microbe interactions develop during succession is not well-studied and is relevant in the light of climate change and increased glacier retreat.

We examined how the bacterial communities associated with two moss species of the genus Racomitrium and the underlying substrate, as well as moss traits and nitrogen fixation, develop along a successional gradient in the glacier forefield of Flaajokull in southeast Iceland. In addition, tested whether moss functional traits, such as total carbon (TC) and nitrogen contents (TN) are drivers of moss and underlying soil bacterial communities.

Although time since deglaciation did not affect TN and moisture content, TC and shoot length increased with time since deglaciation. Moss and underlying soil bacterial communities were distinct. While the soil bacterial community structure was driven by the time since deglaciation and moss C/N ratios, the moss bacterial community structure was linked to time since deglaciation and moss moisture content. Moss N2-fixation rates were linked to bacterial community composition and nifH gene abundance rather than moss TN or time since deglaciation. This was accompanied by a shift from autotrophic to heterotrophic diazotrophs.

Overall, our results suggest that there is little lateral transfer between moss and soil bacterial communities and that moss traits and time since deglaciation affect moss and soil bacterial community structure. In addition, moss N2-fixation rates are determined by bacterial community structure, rather than moss traits or time since deglaciation.


title: MoDLE: High-performance stochastic modeling of DNA loop extrusion interactions

abstract: DNA loop extrusion emerges as a key process establishing genome structure and function. We introduce MoDLE, a computational tool for fast, stochastic modeling of molecular contacts from DNA loop extrusion capable of simulating realistic contact patterns genome wide in a few minutes. MoDLE accurately simulates contact maps in concordance with existing molecular dynamics approaches and with Micro-C data, and does so orders of magnitude faster than existing approaches. MoDLE runs efficiently on machines ranging from laptops to high performance computing clusters, and opens up for exploratory and predictive modeling of 3D genome structure in a wide range of settings.


title: Rats exhibit higher hepatic stress response pathway capacities compared with mice and humans

abstract: In order to minimise the occurrence of unexpected toxicities as a new medicine transitions from the preclinical to clinical phases of development, it is vital to understand fundamental similarities and differences between preclinical species and humans. The well-known difference in sensitivity of mice and rats to acetaminophen (APAP) liver injury has been related to differences in the fraction that is bioactivated to the reactive metabolite N-acetyl-p-benzoquinoneimine (NAPQI). We have used physiologically-based pharmacokinetic modelling to identify doses of APAP (300 and 1000 mg/kg in mice and rats, respectively) yielding similar hepatic burdens of NAPQI to enable the comparison of temporal liver tissue responses under conditions of equivalent chemical insult. Despite pharmacokinetic and biochemical verification of the equivalent NAPQI insult, serum biomarker and tissue histopathology analyses revealed that mice still exhibited a greater degree of liver injury than rats. Transcriptomic and proteomic analyses highlighted the stronger activation of stress response pathways (including the Nrf2 oxidative stress response and autophagy) in the livers of rats, indicative of a more robust transcriptional adaptation to the equivalent insult. Using RNA-Seq data, we also found that genes associated with these stress responses are expressed at a higher basal level in the livers of rats compared with both mice and humans. Taken together, these data indicate that rats possess a greater basal and adaptive capacity for hepatic stress responses than mice and humans. This has important implications for species selection in preclinical safety testing of drugs associated with reactive metabolite formation and other forms of chemical stress.

category: pharmacology and toxicology


title: Slow oscillations promote long range effective communication: the key for memory consolidation in a broken down network

abstract: One of the most prominent and robust findings in cognitive neuroscience is the strengthening of memories during non-rapid eye movement (NREM) sleep, with slow oscillations (SOs, < 1Hz) playing a critical role in systems-level consolidation. However, NREM is a sleep period generally showing a breakdown in connectivity and reduction of synaptic plasticity with increasing depth: a brain state seemingly unfavorable to memory consolidation. Here, we present a novel approach to address this apparent paradox that leverages an event-related causality measure to estimate directional information flow during NREM sleep in epochs with and without SOs. Our results confirm that NREM is generally a state of dampened neural communication, but reveals that SOs provide two windows of enhanced large-scale communication before and after the SO trough. These peaks in communication are significantly higher when SOs are coupled with sleep spindles, compared with uncoupled SOs. To probe the functional relevance of these SO-selective peaks of information flow, we tested the temporal and topographic conditions that predict overnight episodic memory improvement. Our results show that global, long-range communication during SOs promote sleep-dependent systems consolidation of episodic memories. A significant correlation between peaks of information flow and memory improvement lends a predictive validity to our measurements of effective connectivity. In other words, we were able to predict memory improvement on the basis of independent electrophysiological observations during sleep. This work introduces a non-invasive approach to understanding information processing during sleep, a behavioral stage whose function, until now, has been understood only after synaptic reorganization after waking from sleep. Our findings provide a mechanism for how systems-level brain communication can occur during an otherwise low connectivity sleep state, indicating that SOs are a gating mechanism for large-scale neural communication, which is the necessary substrate for systems consolidation and long-term memory formation.


title: Long-Short Term Memory Networks for Electric Source Imaging with Distributed Dipole Models

abstract: 1Magneto- and electroencephalography (M/EEG) are widespread techniques to measure neural activity in vivo at a high temporal resolution but relatively low spatial resolution. Locating the sources underlying the M/EEG poses an inverse problem, which is itself ill-posed. In recent years, a new class of source imaging methods was developed based on artificial neural networks. We present a long-short term memory (LSTM) network to solve the M/EEG inverse problem. It integrates several aspects essential for qualitative inverse solutions: Low computational cost, exploitation of both spatial and temporal information, input flexibility and robustness to noise. Using simulation data, the LSTM shows higher accuracy on multiple metrics and for varying numbers of neural sources, compared to classical inverse solutions but also compared to our alternative architectures without integration of temporal information. It successfully integrates the temporal context given by sequential data, which is particularly useful with noisy data. Real data of a median nerve stimulation paradigm was used to show that the LSTM predicts plausible sources that are in concordance with classical inverse solutions. The performance of the LSTM regarding its robustness to noise renders it a promising and easy-to-apply inverse solution to be considered in future source localization studies and for clinical applications.


title: Perceptual expectations and false percepts generate stimulus-specific activity in distinct layers of the early visual cortex

abstract: Popular theories suggest that hallucinations arise through excessive top-down perceptual expectations, whereas others have emphasised the role of spontaneous bottom-up activity. These theories make different predictions about how input and feedback layers in sensory regions contribute to hallucinations. Here, we used layer-specific fMRI to interrogate neural activity underlying hallucinations - high confidence false percepts - and perceptual expectations while healthy participants (N=25) performed a perceptual discrimination task. We found that false percepts were related to stimulus-like activity in the middle input layers of V2. On the other hand, perceptual expectations activated the deep feedback layers of V2, without influencing perception. The prevalence of high confidence false percepts was related to everyday hallucination severity, confirming their ecological validity. These results reveal that hallucinations can arise through spontaneous stimulus-specific activity in the input layers of the visual cortex, independent of top-down perceptual expectations.


title: Landscape of immune-related signatures induced by targeting of different epigenetic regulators in melanoma: implications for immunotherapy

abstract:
BackgroundInnovative cancer immunotherapy approaches aim at combining immune checkpoint inhibitors with other immunomodulatory agents. Epigenetic regulators can control immune-related genes, therefore targeting them with specific inhibitors may be a potential way forward. Here we identified immune-related signatures induced by four classes of epigenetic drugs in human melanoma cells to define the most promising agent and to understand its biological activity in-vitro, in-vivo and in clinical samples.

MethodsHuman melanoma cell lines were characterized for mutational and differentiation profile and treated with inhibitors of DNA methyltransferases (guadecitabine), histone deacetylases (givinostat), bromodomain and extraterminal domain proteins (JQ1 and OTX-015) and enhancer of zeste homolog 2 (GSK126). Drug-specific gene signatures were identified by Clariom S and Nanostring platforms. Modulation of 14 proteins was determined by quantitative western blot. Ingenuity Pathway Analysis (IPA) identified Upstream Regulator (UR) molecules explaining changes in gene expression and biological activity of drugs. Gene set enrichment and IPA were used to test modulation of guadecitabine-specific gene and UR signatures, respectively, in on-treatment tumor biopsies from melanoma patients enrolled in the Phase Ib NIBIT-M4 Guadecitabine + Ipilimumab Trial.

ResultsDrug-specific gene and UR signatures were identified for each of the four inhibitors. Immune-related genes were frequently upregulated by guadecitabine, to a lesser extent by givinostat, but downregulated by JQ1 and OTX-015. GSK126 was the least active drug. Treatment of melanoma cells with combination of two epigenetic drugs revealed a dominant effect of guadecitabine and JQ1 on immune-related gene modulation. Drug-specific modulatory profiles were confirmed at the protein level. The guadecitabine-specific UR signature was characterized by activated molecules of the TLR, NF-kB, and IFN innate immunity pathways and was induced in drug-treated melanoma, mesothelioma, hepatocarcinoma cell lines and human melanoma xenografts. Most of the guadecitabine-specific signature genes (n>160) were upregulated in on-treatment tumor biopsies from NIBIT-M4 trial. Progressive activation of guadecitabine UR signature molecules was observed in on-treatment tumor biopsies from responding compared to non-responding patients.

ConclusionsGuadecitabine was the most promising immunomodulatory agent among those investigated. This DNA methyltransferases inhibitor emerged as a strong inducer of innate immunity pathways, supporting the rationale for its use in combinatorial immunotherapy approaches.


title: Larger but younger fish when growth compensates for higher mortality in heated ecosystem

abstract: Ectotherms are often predicted to "shrink" with global warming, in line with general growth models and the temperature-size rule (TSR), both predicting smaller adult sizes with warming. However, they also predict faster juvenile growth rates, leading to larger size-at-age. Hence, the result of warming on population size-structure also depends on mortality rates and how much also adult size changes. We used data from an artificially heated (+8C) bay in comparison with an unheated area, to analyze how warming has affected body growth, mortality rates and population size-structure of Eurasian perch (Perca fluviatilis). In the heated bay, body size was larger for all ages and growth faster for all sizes, resulting in larger size-spectrum exponent (greater proportion of large fish), despite higher mortality. Hence, to understand how warming alters species size-structure, and thus ecological interactions and dynamics, it is critical to account for warming-induced changes in both mortality and growth rate.


title: Dimensionality of genomic information and its impact on GWA and variant selection: a simulation study

abstract:
BackgroundIdentifying true-positive variants in genome-wide associations (GWA) depends on several factors, including the number of genotyped individuals. The limited dimensionality of the genomic information may give insights into the optimal number of individuals to use in GWA. This study investigated different discovery set sizes in GWA based on the number of largest eigenvalues explaining a certain proportion of variance in the genomic relationship matrix (G). An additional investigation included the change in accuracy by adding variants, selected based on different set sizes, to the regular SNP chips used for genomic prediction.

MethodsSequence data were simulated containing 500k SNP with 200 or 2000 quantitative trait nucleotides (QTN). A regular 50k panel included one every ten simulated SNP. Effective population size (Ne) was 20 and 200. The GWA was performed with the number of genotyped animals equivalent to the number of largest eigenvalues of G (EIG) explaining 50, 60, 70, 80, 90, 95, 98, and 99% of the variance. In addition, the largest discovery set consisted of 30k genotyped animals. Limited or extensive phenotypic information was mimicked by changing the trait heritability. Significant and high effect size SNP were added to the 50k panel and used for single-step GBLUP with and without weights.

ResultsUsing the number of genotyped animals corresponding to at least EIG98 enabled the identification of QTN with the largest effect sizes when Ne was large. Smaller populations required more than EIG98. Furthermore, using genotyped animals with higher reliability (i.e., higher trait heritability) helped better identify the most informative QTN. The greatest prediction accuracy was obtained when the significant or the high effect SNP representing twice the number of simulated QTN were added to the 50k panel. Weighting SNP differently did not increase prediction accuracy, mainly because of the size of the genotyped population.

ConclusionsAccurately identifying causative variants from sequence data depends on the effective population size and, therefore, the dimensionality of genomic information. This dimensionality can help identify the suitable sample size for GWA and could be considered for variant selection. Even when variants are accurately identified, their inclusion in prediction models has limited implications.

category: genomics


title: Automated detection of sow posture changes with millimeter-wave radars and deep learning

abstract:
Automated behavioural monitoring is increasingly required for animal welfare and precision agriculture. In pig farming, detailed analyses of sow activity are essential to identify and reduce the risks of piglets being crushed during postural changes of their mothers. Here we introduce a new, non-invasive, fast and accurate method for monitoring sow behaviour based on millimeter-wave radars and deep learning analysis. We used our method to predict postural changes in crated sows and distinguish the dangerous one that lie down abruptly from those that lie down carefully using transient postures. Two radars were placed on a metal backing above the head and the upper part of the back of each of ten sows to monitor their activity during 5 hours. We analysed the radar data with a convolutional neural network and identified five postures. The average sensitivity was 96.9% for standing, 90.8% for lying, 91.4% for nursing, 87.6% for sitting, but only 11.9% for kneeling. However, the average specificity and accuracy were greater than 92% for the five postures. Interestingly, two of the ten sows occasionally moved directly from standing to lying, without using the transient postures sitting and kneeling, thereby displaying risky behaviours for their piglets. Our radar-based classifier is more accurate, faster and require less memory than current computer vision approaches. Using more sows will improve the algorithm performance and facilitate future applications for large scale deployment in animal farming.

HighlightsO_LIAutomated behavioural analysis is a major challenge for precision farming.
C_LIO_LIWe developed automated detection of lactating sow postures with radars and deep learning.
C_LIO_LIWe identified five postures, including transitions risky for the piglets.
C_LIO_LIOur method is accurate, fast and requires less memory than computer vision.
C_LIO_LIRadars thus hold considerable promises for high through-put recording of livestock activity.
C_LI


title: Onlinemeta: A Web Serve For Meta-Analysis Based On R-shiny

abstract: Meta-analysis is a common statistical method used to summarize multiple studies that cover the same topic. It can provide less biased results and explain heterogeneity between studies. Although there exists a variety of meta-analysis software, they are rarely both convenient to use and capable of comprehensive analytical functions. As a result, we established a meta-analysis web tool called Meta-Analysis Online (Onlinemeta). Onlinemeta includes both risk bias analysis and meta-analysis. The risk bias analysis tool can produce heatmaps and histograms, whereas the meta-analysis tool can be used to analyze dichotomous variables, single-armed dichotomous variables, continuous variables, single-armed continuous variables, survival data, and diagnostic experiments. In addition, it can be used to generate forest plots, funnel plots, SROC curve, and crosshair plots. Onlinemeta can be found at the following address: https://smuonco.Shinyapps.io/Onlinemeta/.


title: Onlinemeta: A Web Server For Meta-Analysis Based On R-shiny


title: Spatiotemporal patterning enabled by gene regulatory networks

abstract: Spatiotemporal pattern formation plays a key role in various biological phenomena including embryogenesis and neural network formation. Though the reaction-diffusion systems enabling pattern formation have been studied phenomenonlogically, the biomolecular mechanisms behind these processes has not been modelled in detail. Here, we study the emergence of spatiotemporal patterns due to simple synthetic commonly observed two- and three-node gene regulatory network motifs coupled with their molecular diffusion in one- and two-dimensional space. We investigate the patterns formed due to the coupling of inherent multistable and oscillatory behavior of toggle switch (two mutually repressing nodes), toggle switch with double self-activation, toggle triad (three mutually repressing nodes) and repressilator (three nodes repressing the other sequentially in a cyclic manner) with the effect of spatial diffusion of these molecules. We probe various parameter regimes corresponding to different regions of stability (monostable, multistable, oscillatory) and assess the impact of varying diffusion coefficients too. This analysis offers valuable insights into the design principles of pattern formation facilitated by these network motifs, and suggest mechanistic underpinnings of biological pattern formation.

category: biophysics


title: Patching Science - amending the literature through version control

abstract: The ideal of self-correction in science is not well served by the current culture and system surrounding amendments to published literature. Here we report on a survey (N = 132) that highlights academics dissatisfaction with the status quo and their support for an alternative approach. We then describe our view of how amendments could and should work by drawing on the idea of an author-led version control system. Here authors would include a link in their published manuscripts to an updatable website (e.g. a GitHub repository or similar) that could be disseminated in the event of any amendment. Such a system is already in place for computer code and, as such, requires nothing but buy-in from the scientific community - a community that is already evolving towards various open science frameworks. This would remove a number of frictions that discourage amendments thus leading to an improved scientific literature and a healthier academic climate.

category: scientific communication and education


title: Tree diversity effects on productivity depend on mycorrhizae and life strategies in a temperate forest experiment

abstract: Tree species are known to predominantly interact either with arbuscular mycorrhizal (AM) or ectomycorrhizal (EM) fungi. However, there is a knowledge gap whether these mycorrhizae differently influence biodiversity-ecosystem functioning (BEF) relationships and whether a combination of both can increase community productivity. In 2015, we established a tree-diversity experiment by growing tree communities with varying species-richness levels (1, 2, or 4 species), and either with AM or EM tree species, or a combination of both. We investigated basal area and annual basal area increment from 2015 to 2020 as proxy for community productivity. We found significant positive relationships between tree species richness and community productivity, which strengthened over time. Further, AM and EM tree species differently influenced productivity; however, there was no overyielding when AM and EM trees grew together. EM tree communities were characterized by low productivity in the beginning, but an increase of increment over time, and showed overall strong biodiversity effects. For AM tree communities the opposite was true. While young trees did not benefit from the presence of the other mycorrhizal type, dissimilar mechanisms underlying BEF relationships in AM and EM trees indicate that maximizing tree and mycorrhizal diversity may increase ecosystem functioning in the long run.


title: Spinosaurids as 'subaqueous foragers' undermined by selective sampling and problematic statistical inference

abstract: Fabbri et al.1 claim that the huge sail-backed dinosaur Spinosaurus aegyptiacus and the spinosaurid Baryonyx were "subaqueous foragers," diving underwater in pursuit of prey, based on their measure of bone "compactness." Using thin-sections and computed tomographic (CT) scans of thigh bone (femur) and trunk rib from various living and extinct vertebrates, they claim to be able to distinguish taxa with "aquatic habits" from others. Their conclusions are undermined by selective bone sampling, inaccuracies concerning spinosaurid bone structure, faulty statistical inferences, and novel redefinition of the term "aquatic."

category: paleontology


title: Coding of reward uncertainty and probability by orexin neurons

abstract: Appropriate arousal in our brains critically relies on hypothalamic hypocretin/orexin neurons (HONs). HON activity regulation has been a subject of intense research, uncovering slow responses to nutrients and hormones, and fast responses to external sensory inputs. However, it remains unclear whether HONs also respond to statistical patterns in the environment. Among these, reward probability and uncertainty are fundamental measures of information that enable evaluation of predictions, and guide diverse aspects of cognition such as learning and motivation. Using different cues to denote reward likelihood across the full probability range (0-1), we found that HON population responses to the reward-predicting cues co-vary with reward uncertainty. Single-cell analyses additionally revealed HON subsets specializing in monotonic representations of probability, as well as cells sharply tuned to uncertainty. Furthermore, simultaneous recordings of HONs and midbrain dopamine neurons (DANs) indicated that HONs and DANs diverge in their coding of reward probability, both in responses to reward-predicting cues and to rewards. These results demonstrate previously unseen coding of reward expectation in genetically-defined hypothalamic cells central to clinical conditions and neurobehavioral physiology, and suggest a possible role for HONs in converting reward statistics into arousal-based allocation of cognitive and autonomic resources.

category: neuroscience


title: Fast Kernel-based Association Testing of non-linear genetic effects for Biobank-scale data

abstract: Our knowledge of non-linear genetic effects on complex traits remains limited, in part, due to the modest power to detect such effects. While kernel-based tests offer a powerful approach to test for nonlinear relationships between sets of genetic variants and traits, current approaches cannot be applied to Biobank-scale datasets containing hundreds of thousands of individuals. We propose, FastKAST, a Kernel-based approach that can test for non-linear effects of a set of variants on a trait. FastKAST provides calibrated hypothesis tests while enabling analysis of Biobank-scale datasets with hundreds of thousands of individuals. We applied FastKAST to thirty quantitative traits measured across {approx} 300 K unrelated white British individuals in the UK Biobank to detect sets of variants with nonlinear effects at genome-wide significance.


title: The broad shell colour variation in common cockle (Cerastoderma edule) from Northeast Atlantic relies on a major QTL revealed by GWAS using a new high-density genetic map

abstract: Shell colour pattern shows broad diversity in molluscs, and both genetic and environmental factors seem to interact to some extent on the final phenotype. Despite information on the genetic component and pathways involved in shell construction and colour has increased in the last decade, more data are needed particularly to understand colour variation and its putative role on adaptation. The European common cockle (Cerastoderma edule) is a valuable species from ecological and commercial perspectives with important variation in colour pattern, but this diversity has never been characterized and the underlying genetic architecture is unknown. In this study, we constructed a high-density genetic map, as an essential tool for genomic screening in common cockle, that was applied to ascertain the genetic basis of colour pattern variation in the species. The consensus map, including 13,874 2b-RAD SNPs, was constituted by the 19 linkage groups (LGs) corresponding to the n = 19 chromosomes of its karyotype and spanned 1,073 cM (730 markers per LG; inter-marker distance of 0.13 cM). Five full-sib families showing segregation for several colour-associated traits were used to perform a GWAS analysis. A major QTL on chromosome 13 explained most of the variation for shell colour patterns. Mining on this genomic region revealed the presence of several candidate genes enriched on Gene Ontology terms such as anatomical structure development, ion transport, membrane transport and cell periphery, closely related to shell architecture, including six chitin-related, one ependymin, several ion binding and transporters, and others related to transit across the cell membrane. Interestingly, this major QTL overlaps with a genomic region previously reported associated with divergent selection in the distribution range of the species, suggesting a putative role on local adaptation.


title: Predicting Dog Phenotypes from Genotypes

abstract: We analyze dog genotypes (i.e., positions of dog DNA sequences that often vary between different dogs) in order to predict the corresponding phenotypes (i.e., unique observed characteristics). More specifically, given chromosome data from a dog, we aim to predict the breed, height, and weight. We explore a variety of linear and non-linear classification and regression techniques to accomplish these three tasks. We also investigate the use of a neural network (both in linear and non-linear modes) for breed classification and compare the performance to traditional statistical methods. We show that linear methods generally outperform or match the performance of non-linear methods for breed classification. However, we show that the reverse is true for height and weight regression. Finally, we evaluate the results of all of these methods based on the number of input features used in the analysis. We conduct experiments using different fractions of the full genomic sequences, resulting in input sequences ranging from 20 SNPs to [~]200k SNPs. In doing so, we explore the impact of using a very limited number of SNPs for prediction. Our experiments demonstrate that these phenotypes in dogs can be predicted with as few as 0.5% of randomly selected SNPs (i.e., 992 SNPs) and that dog breeds can be classified with 50% balanced accuracy with as few as 0.02% SNPs (i.e., 40 SNPs).


title: A Novel and Prevalent Pathway in Microbial Selenium Metabolism

abstract: Selenium is an essential micronutrient in diverse organisms. Two routes are known for its insertion into proteins and nucleic acids via selenocysteine and 2-selenouridine, respectively1. However, despite its importance, pathways for specific incorporation of selenium into small molecules have remained elusive. We herein use a genome mining strategy to uncover a widespread three-gene cluster in varied microorganisms that encodes a dedicated pathway for producing selenoneine, the selenium-analog of the multifunctional molecule ergothioneine2,3. We elucidate the reactions of all three proteins and uncover two novel selenium-carbon bond-forming enzymes and the first biosynthetic pathway for production of a selenosugar, an unexpected intermediate on route to the final product. Our findings expand the scope of biological selenium utilization, suggest that the selenometabolome is more diverse than previously thought, and set the stage for the discovery of other selenium-containing natural products.


title: CellDrift: Inferring Perturbation Responses in Temporally-Sampled Single Cell Data

abstract: Cells and tissues respond to perturbations in multiple ways that can be sensitively reflected in alterations of gene expression. Current approaches to finding and quantifying the effects of perturbations on cell-level responses over time disregard the temporal consistency of identifiable gene programs. To leverage the occurrence of these patterns for perturbation analyses, we developed CellDrift (https://github.com/KANG-BIOINFO/CellDrift), a generalized linear model-based functional data analysis method capable of identifying covarying temporal patterns of various cell types in response to perturbations. As compared to several other approaches, CellDrift demonstrated superior performance in the identification of temporally varied perturbation patterns and the ability to impute missing time points. We applied CellDrift to multiple longitudinal datasets, including COVID-19 disease progression and gastrointestinal tract development, and demonstrated its ability to identify specific gene programs associated with sequential biological processes, trajectories, and outcomes.


title: Heterogeneity in the gene regulatory landscape of leiomyosarcoma.

abstract: Soft-tissue sarcomas are group of rare but highly aggressive malignancies. It is a tremendously heterogeneous group of tumors. Characterizing inter-tumor heterogeneity is crucial for selecting suitable cancer therapy as the presence of diverse molecular subgroups of patients can be associated with disease outcome or response to treatment. However, no methods have been developed to characterize heterogeneity based on genome-wide patient-specific regulatory networks. In this work, we propose a simple but efficient approach to characterize inter-tumor regulatory network heterogeneity, which we call PORCUPINE (Principal Components Analysis to Obtain Regulatory Contributions Using Pathway-based Interpretation of Network Estimates). PORCUPINE uses as input individual patient regulatory networks, represented by estimated regulatory interactions between transcription factors and their target genes, and a list of genes assigned to biological pathways in order to identify key pathways that drive heterogeneity among individuals. We used PORCUPINE to model regulatory heterogeneity in leiomyosarcoma, one of the most common softtissue sarcomas subtypes. We applied it to 80 genome-wide leiomyosarcoma regulatory networks modeled on data from The Cancer Genome Atlas and validated the results in an independent dataset of 37 leiomyosarcoma cases from the German Cancer Research Center. PORCUPINE identified 37 pathways, including pathways that represent potential targets for treatment of subgroups of leiomyosarcoma patients, such as FGFR and CTLA4 inhibitory signaling. PORCUPINE thereby provides a robust way of analyzing and interpreting patient-specific regulatory networks and is the first step towards implementing network-informed personalized medicine in leiomyosarcoma.


title: Improved MALDI-TOF MS based antimicrobial resistance prediction through hierarchical stratification

abstract:
MotivationPredicting antimicrobial resistance using MALDI-TOF mass spectrometry based machine learning is a fast-growing field of research. Recent advances in machine learning methods specifically designed for MALDI-TOF mass spectra have outperformed established classification approaches. However, classification performance was observed to have a large standard deviation between different train-test splits. We hypothesise that this variance is caused by the underlying phylogenetic structure between microbial samples, which is implicitly reflected in their MALDI-TOF MS profiles, but not taken into account during the training of a model.

ResultsIn this paper, we propose to infer this structure from the dataset--using agglomerative hierarchical clustering--and consider it during the dataset splitting between train and test. We show that incorporating such phylogenetic structure into the antimicrobial resistance prediction scenario leads to an improved classification performance. Average precision was increased from 42.3 to 47.1 for ciprofloxacin resistance prediction in Escherichia coli and from 44.6 to 50.8 for amoxicillin-clavulanic acid resistance prediction in Staphylococcus aureus using a Gaussian process classifier with a MALDI-TOF MS specific kernel. We envision that these results will support the quick and reliable identification of antimicrobial resistances, thus increasing patient well-being and reducing healthcare costs.

AvailabilityAll data is available for download and code available as an easy-to-use Python package under https://github.com/BorgwardtLab/maldi_PIKE at branch maldi_stratification.

Contactcaroline.weis@bsse.ethz.ch, karsten.borgwardt@bsse.ethz.ch

Supplementary informationSupplementary information at the end of document.


title: Tumor cell-intrinsic HFE drives glioblastoma growth

abstract:
BackgroundGlioblastoma (GBM) tumor cells modulate expression of iron-associated genes to enhance iron uptake from the surrounding microenvironment, driving proliferation and tumor growth. The homeostatic iron regulator (HFE) gene, encoding the iron sensing HFE protein, is upregulated in GBM and correlates with poor survival outcomes. However, the molecular mechanisms underlying these observations remain unclear. Identification of pathways for targeting iron dependence in GBM tumors is therefore a critical area of investigation.

MethodsWe interrogated the impact of cell-intrinsic Hfe expression on proliferation and tumor growth through genetic loss and gain of function approaches in syngeneic mouse glioma models. We determined the expression of iron-associated genes and their relationship with survival in GBM using public datasets and identified differentially expressed pathways in Hfe knockdown cells through Nanostring transcriptional profiling.

ResultsLoss of Hfe induced apoptotic cell death in vitro and inhibited tumor growth in vivo while overexpression of Hfe accelerated both proliferation and tumor growth. Analysis of iron gene signatures in Hfe knockdown cells revealed alterations in the expression of several iron-associated genes, suggesting global disruption of intracellular iron homeostasis. Analyzing differentially expressed pathways further identified oxidative stress as the top pathway upregulated with Hfe loss. Enhanced 55Fe uptake and generation of reactive oxygen species (ROS) were found with Hfe knockdown, implicating toxic iron overload resulting in apoptotic cell death.

ConclusionsCollectively, these findings identify a novel role for HFE in regulating iron homeostasis in GBM tumors and provide a potential avenue for future therapeutic development.

Key PointsO_LIHFE is an iron sensor that is upregulated in GBM and negatively impacts survival.
C_LIO_LIHFE overexpression drives proliferation and tumor growth in vivo.
C_LIO_LILoss of HFE increases production of reactive oxygen species and induces apoptosis, extending survival in vivo.
C_LI

Importance of StudyDysregulation of iron metabolism is an important feature of GBM contributing to tumor growth and negatively impacting survival. The identification of key iron regulators controlling this process is therefore important for therapeutic targeting. We identify HFE as an important regulator of iron homeostasis in GBM and suggest a role for sexual dimorphism in HFE-mediated tumor iron regulation that ultimately results in differential survival outcomes. Our findings demonstrate that HFE drives tumor cell proliferation and survival in GBM and may be a viable target for modulating tumor iron flux and inducing apoptosis in tumor cells.

category: cancer biology


title: CRISPR screens reveal genetic determinants of PARP inhibitor sensitivity and resistance in prostate cancer

abstract: Prostate cancer (PCa) harboring BRCA1/2 mutations is often exquisitely sensitive to PARP inhibition. However, genomic alterations in other DNA damage response genes have not been consistently predictive of clinical response to PARP inhibitors (PARPis). Here, we perform genome-wide CRISPR-Cas9 knockout screens in BRCA1/2-proficient PCa cell lines and identify novel genes whose loss has a profound impact on PARPi sensitivity and resistance. Specifically, MMS22L deletion, frequently observed (up to 14%) in PCa, renders cells hypersensitive to PARPis by disrupting RAD51 loading required for homologous recombination repair, although this response is TP53-dependent. Unexpectedly, loss of CHEK2 confers resistance rather than sensitivity to PARPis in PCa cells through increased expression of BRCA2, a target of CHEK2-TP53-E2F7-mediated transcriptional repression. Combined PARP and ATR inhibition overcomes PARPi resistance caused by CHEK2 loss. Our findings may inform the use of PARPis beyond BRCA1/2-deficient tumors and support reevaluation of currently used biomarkers for PARPi treatment in PCa.


title: Transcriptomics based prediction of survival and response to therapy in malignant mesothelioma

abstract: Malignant mesothelioma is an aggressive cancer with limited treatment options and poor prognosis. Better understanding of mesothelioma genomics and transcriptomics could advance novel therapies. We performed whole-exome and RNA-sequencing of germline and tumors of 122 patients with pleural, peritoneal, and tunica-vaginalis mesothelioma. We identify a 48 gene prognostic signature that is highly predictive of mesothelioma patient survival including CCNB1, whose expression is highly predictive of patient survival on its own. Using a synthetic-lethality (SL) based pipeline for analyzing the patients transcriptomic data, we identified SL-based signatures predictive of response to an anti-PD1 immune checkpoint inhibitor and combination therapies with pemetrexed. These SL-profiles successfully predict the overall patient-response observed across targeted, immuno- and chemotherapies in 11 independent mesothelioma clinical trials spanning 7 different treatments. These findings lay a basis for future studies aimed specifically at testing the ability of these SL profiles to serve as treatment biomarkers in mesothelioma.