title: Insertion of YFP at the Arabidopsis AFL1 and P5CS1 loci shows the potential, and potential limitations, of gene targeting for functional analysis of stress-related genes

abstract: Transgenic expression of a tagged protein under control of its native promoter is the typical strategy to investigate tissue-specific accumulation and subcellular localization of a protein of interest. In some cases, difficulties in promoter identification, altered promoter function when placed in a new genomic context, and variability between transgenic lines can complicate such experiments. Crispr/CAS9-enabled homologous recombination to insert a tag in frame with an endogenous gene could circumvent these problems. Apart from work at a few loci used to test gene targeting (GT) protocols, there have been relatively few reports of GT to knock-in protein tags in Arabidopsis thaliana, and little data of whether expression of the knock-in allele accurately reflects that of the wild-type gene. We investigated these questions using a two-stage GT procedure where a starter line expressing hyper-accurate CAS9 (hypaCAS9) under control of the Egg Cell1.1 (EC1.1) promoter was transformed with an mCitrine (YFP) donor sequence flanked by homology arms for homologous recombination at the N- or C-terminal ends of the stress-responsive genes At14a-Like 1 (AFL1) and {Delta}1-pyrroline-5-carboxylate synthetase 1 (P5CS1). We observed high frequency GT to generate in frame fusions of YFP at either the N- or C-terminus of AFL1 and at the N-terminus of P5CS1. AFL1-YFP was produced from the knock-in locus; however, the N-terminal knock-ins disrupted AFL1 and P5CS1 expression. These results show that the combination of EC1.1 promoter and hypaCAS9 along with selection of a starter line with strong hypaCAS9 expression produced good GT frequencies, indicating that this approach can be feasible to knock-in or replace sequence at many loci. However, we also offer a cautionary tale that insertions generated by GT can interfere with gene expression for reasons which are yet unclear. Further refinement of this approach is a potentially useful tool to produce tagged proteins from their endogenous genomic loci.


title: Differential correlation across subpopulations of single cells in subtypes of acute myeloid leukemia

abstract:
Mass cytometers can record 40-50 parameters per single cell for millions of cells in a sample, and in particular, for leukemic cells. Many methods have been developed to cluster phenotypically similar cells within cytometry data, but there are fewer methods to visualize activity and interactions of pairs of proteins across these populations. We have developed a workflow for analyzing correlations associated with predfined populations. By clustering blood samples from acute myeloid leukemia (AML) patients and normal controls using an established algorithm, we obtained a minimum spanning tree of clusters of single cells. Using surface marker expression, we identified clusters on the tree that belonged to phenotypes of interest. Next, we computed correlations between pairs of proteins in each cluster. We developed a novel, coherent, probability-based statistic to test differences between vectors of correlation coefficients. By comparing all combinations of the normal controls under the statistic, we created an empirical distribution that provided a conservative measure of differential correlation. Using this empirically-derived distribution to define significance, we compared pooled samples from AML subtypes and normal controls to detect differential correlations. Given the structure present within this cytometry data set, we found it natural to consider correlations in this manner versus aggregating all data and computing a single correlation. Our results have the advantage that we can localize the statistical measure to determine contributions from particular phenotypic populations. Differentially correlated pairs of proteins can be further explored by considering a populations distribution of correlation coefficients or biaxially plotting protein expressions within individual cells in a given population. Our approach leads to a better understanding of the nonlinear relationships that exist in the cytometry data.

Author summaryWe introduce a novel method for analyzing the abundance of single cell data collected by high-throughput technologies. Due to the high dimensionality of such datasets, there is a need for methods to identify significant interactions between genes or proteins. In particular, we are interested in statistical differences between correlations of proteins within populations of cells determined by traditional immunophenotyping techniques. In this paper, we have demonstrated the utility of this new framework in the case of blood samples from individuals with different subtypes of leukemia and compare to healthy controls. The motivation for this application is that differences between can illuminate potential drivers of the disease. We have illustrated an example of how intracellular events can be detected by our statistic. Finally, this method is flexible in that it can also be applied in a variety of contexts where there are vectors of correlation coefficients that need to be compared.

category: systems biology


title: Effects of chronic cannabidiol in a mouse model of naturally occurring neuroinflammation, neurodegeneration, and spontaneous seizures

abstract: Cannabidiol (CBD) has gained attention as a therapeutic agent and is purported to have immunomodulatory, neuroprotective, and anti-seizure effects. Here, we determined the effects of chronic CBD administration in a mouse model of CLN1 disease (Cln1-/-) that simultaneously exhibits neuroinflammation, neurodegeneration, and spontaneous seizures. Proteomic analysis showed that putative CBD receptors are expressed at similar levels in the brains of Cln1-/- mice compared to normal animals. Cln1-/- mice received an oral dose (100mg/kg/day) of CBD for six months and were evaluated for changes in pathological markers of disease and seizures. Chronic cannabidiol administration was well-tolerated, high levels of CBD were detected in the brain, and markers of astrocytosis and microgliosis were reduced. However, CBD had no apparent effect on seizure frequency or neuron survival. These data are consistent with CBD having immunomodulatory effects. It is possible that a higher dose of CBD could also reduce neurodegeneration and seizure frequency.

category: neuroscience


title: Bio-inspired neural networks implement different recurrent visual processing strategies than task-trained ones do

abstract: Behavioral studies suggest that recurrence in the visual system is important for processing degraded stimuli. There are two broad anatomical forms this recurrence can take, lateral or feedback, each with different assumed functions. Here we add four different kinds of recurrence--two of each anatomical form--to a feedforward convolutional neural network and find all forms capable of increasing the ability of the network to classify noisy digit images. Specifically, we take inspiration from findings in biology by adding predictive feedback and lateral surround suppression. To compare these forms of recurrence to anatomically-matched counterparts we also train feedback and lateral connections directly to classify degraded images. Counter-intuitively, we find that the anatomy of the recurrence is not related to its function: both forms of task-trained recurrence change neural activity and behavior similarly to each other and differently from their bio-inspired anatomical counterparts. By using several analysis tools frequently applied to neural data, we identified the distinct strategies used by the predictive versus task-trained networks. Specifically, predictive feedback de-noises the representation of noisy images at the first layer of the network and decreases its dimensionality, leading to an expected increase in classification performance. Surprisingly, in the task-trained networks, representations are not de-noised over time at the first layer (in fact, they become  noiser and dimensionality increases) yet these dynamics do lead to de-noising at later layers. The analyses used here can be applied to real neural recordings to identify the strategies at play in the brain. Our analysis of an fMRI dataset weakly supports the predictive feedback model but points to a need for higher-resolution cross-regional data to understand recurrent visual processing..


title: A descending inhibitory mechanism of nociception mediated by an evolutionarily conserved neuropeptide system in Drosophila

abstract: Nociception is a neural process that animals have developed to avoid potentially tissue-damaging stimuli. While nociception is triggered in the peripheral nervous system, its modulation by the central nervous system is a critical process in mammals, whose dysfunction has been extensively implicated in chronic pain pathogenesis. The peripheral mechanisms of nociception are largely conserved across the animal kingdom. However, it is unclear whether the brain-mediated modulation is also conserved in non-mammalian species. Here, we show that Drosophila has a descending inhibitory mechanism of nociception from the brain, mediated by the neuropeptide Drosulfakinin (DSK), a homolog of cholecystokinin (CCK) that plays an important role in the descending control of nociception in mammals. We found that mutants lacking dsk or its receptors are hypersensitive to noxious heat. Through a combination of genetic, behavioral, histological, and Ca2+ imaging analyses, we subsequently revealed neurons involved in DSK-mediated nociceptive regulation at a single-cell resolution and identified a DSKergic descending pathway that inhibits nociception. This study provides the first evidence for a descending modulatory mechanism of nociception from the brain in a non-mammalian species that is mediated by the evolutionarily conserved CCK system, raising the possibility that the descending inhibition is an ancient mechanism to regulate nociception.


title: Circuit-Specific Targeting of Astrocytes for Genetic Control

abstract: Astrocytes are integral functional components of brain circuits. They ensheath the connections between neurons to form tripartite synapses. They react to local neuronal activities and release signaling molecules to regulate synaptic transmission. Their dysfunctions impair synaptic functions and are implicated in neuropsychiatric disorders. Increasing evidence indicates that astrocytes are diverse and they have distinct features and functions in different circuits. However, selectively targeting and controlling astrocytes in a circuit-specific manner is technically challenging. Recently we constructed a series of anterograde transneuronal viral vectors based on the yellow fever vaccine YFV-17D. These YFV-17D derivatives express fluorescent proteins almost exclusively in neurons. However, we find that YFV-17D carrying DNA recombinase Cre infect astrocytes associated with the traced neuronal pathways and express Cre to turn on reporter genes. The targeting of astrocytes is at a whole-brain level but specific to the neuronal circuits traced. Therefore, YFV-17D vectors carrying DNA recombinases provide tools for selectively and genetically targeting pathway-specific astrocytes. This new technology will also allow us to reveal the roles of astrocytes in specific neuronal circuits in normal brain functions and diseases.


title: Organ- and function-specific organization and bioelectronic modulation of vagus fascicles and fibers

abstract: Afferent and efferent fibers in the vagus nerve travel inside fascicles and form branches to innervate organs and regulate organ functions. The spatial organization of fibers and fascicles, with respect to the functions they mediate and the organs they innervate, is unknown. Accordingly, it is unknown whether such organization can be leveraged by bioelectronic devices for function- and organ-specific neurostimulation. To characterize the functional microscopic anatomy of the vagus, we developed a pipeline consisting of micro-computed tomography imaging of fascicles and quantitative immunohistochemistry of single fibers. We found that, in swine, the vagus contains clusters of fascicles, organized along 2 overlapping functional axes: one axis specific to sensory vs. motor somatic and visceral functions, with clusters increasingly separating in the cephalad direction, and a second axis specific to innervated organs, including larynx, lungs and heart, with increasing separation in the caudad direction. Large myelinated afferents and efferents, and small unmyelinated efferents occupy large areas of specific clusters of fascicles; small unmyelinated afferents are numerous and occupy small areas across every fascicle. To test whether fiber populations can be selectively modulated, we developed a multi-contact cuff electrode accommodating the nerve fascicular structure. Stimulation targeting separate nerve sections evokes compound action potentials from fibers of distinct functional types and elicits differential organ responses, including laryngeal muscle contraction, cough reflex, and changes in breathing and heart rate. Our results indicate that vagus fibers are anatomically organized according to functions they mediate and organs they innervate and can be differentially modulated by spatially selective nerve stimulation.


title: Organ- and function-specific anatomical organization and bioelectronic modulation of the vagus nerve


title: Gliotransmission of D-serine promotes thirst-directed behaviors in Drosophila

abstract: Thirst emerges from a range of cellular changes that ultimately motivate an animal to consume water. Although thirst-responsive neuronal signals have been reported, the full complement of brain responses is unclear. Here we identify molecular and cellular adaptations in the brain using single-cell sequencing of water deprived Drosophila. Water deficiency primarily altered the glial transcriptome. Screening the regulated genes revealed astrocytic expression of the astray-encoded phosphoserine phosphatase to bi-directionally regulate water consumption. Astray synthesizes the gliotransmitter D-serine and vesicular release from astrocytes is required for drinking. Moreover, dietary D-serine rescues aay-dependent drinking deficits while facilitating water consumption and expression of water-seeking memory. D-serine action requires binding to neuronal NMDA-type glutamate receptors. Fly astrocytes contribute processes to tripartite synapses and the proportion of astrocytes that are themselves activated by glutamate increases with water deprivation. We propose that thirst elevates astrocytic D-serine release, which awakens quiescent glutamatergic circuits to enhance water procurement.


title: A comparison of hard and soft direct methods for DNA extraction from soil

abstract:
Nucleic acid extraction is the first step in molecular biology studies of soil bacterial communities. The most common used soil DNA extraction method is the direct, hard extraction Mobio method, which uses bead beating to lyse bacteria. In this study we compared the Mobio method with a soft, enzymatic lysis extraction method. Next generation sequencing (Illumina and Pyrosequencing) of amplicons generated from four 16S primer pairs and DNA from 12 soils and 3 composts was used to compare the two extraction methods.

Four bacterial orders, the delta proteobacterial Desulfuromonadales and gamma proteobacterial Pseudomonadales, Enterobacteriales, and Alteromonadales were more common in amplicons from soft extracted DNA, sometimes by two orders of magnitude. These groups can be a significant fraction of the bacterial population. For example the Pseudomonadales made up to 16 % and Enterobacteriales 10% of amplicons from Soft extracted DNA. The JG30-KF-CM45 order was under extracted by the enzymatic lysis extraction method. Results differed more by primer choice than extraction method and the phylogenetic resolution of differences between extraction methods changed with primer choice.

Given how often Mobio extraction is used, these proteobacterial orders are probably under-represented in the studies of soil bacteria that use nucleic acid methods. Further improvements in soil DNA extraction are needed. Amplicons sequencing studies should use a range of different primers to confirm the phylogenetic resolution of their results.

ImportanceSeveral large scale studies of soil bacteria that compare thousands of soil samples across continents have used the Mobio method for DNA extraction. Large scale studies like these are increasing with the recent establishment of the Global Soil Biodiversity Observation Network (Soil BON), which also uses the Mobio method. The results of this work will be used to make policy decisions about how to manage the soil and may be a guide for bioprospectors. As the Mobio method is so widely used, it is important to know its limitations. Studies that use the Mobio method underestimate the fraction of several proteobacterial groups. Most notably the Enterobacteria and Pseudomonas can be under extracted by 10-100 fold. The degree of under extraction varies with different soils.

category: microbiology


title: Beer ethanol and iso-α-acid level affect microbial community establishment and beer chemistry throughout wood maturation of beer

abstract: Sour beers produced by barrel-aging of conventionally fermented beers are becoming increasingly popular. However, as the intricate interactions between the wood, the microbes and the beer are still unclear, wood maturation often leads to inconsistent end products with undesired sensory properties. Previous research on industrial barrel-aging of beer suggests that beer parameters like the ethanol content and bitterness play an important role in the microbial community composition and beer chemistry, but their exact impact still remains to be investigated. In this study, an experimentally tractable lab-scale system based on an in-vitro community of four key bacteria (Acetobacter malorum, Gluconobacter oxydans, Lactobacillus brevis and Pediococcus damnosus) and four key yeasts (Brettanomyces bruxellensis, Candida friedrichii, Pichia membranifaciens and Saccharomyces cerevisiae) that are consistently associated with barrel-aging of beer, was used to test the hypotheses that beer ethanol and bitterness impact microbial community composition and beer chemistry. Experiments were performed using different levels of ethanol (5.2 v/v%, 8 v/v% and 11 v/v%) and bitterness (13 ppm, 35 ppm and 170 ppm iso--acids), and beers were matured for 60 days. Samples were taken after 0, 10, 20, 30 and 60 days to monitor population densities and beer chemistry. Results revealed that all treatments and the maturation time significantly affected the microbial community composition and beer chemistry. More specifically, the ethanol treatments obstructed growth of L. brevis and G. oxydans and delayed fungal growth. The iso--acid treatments hindered growth of L. brevis and stimulated growth of P. membranifaciens, while the other strains remained unaffected. Beer chemistry was found to be affected by higher ethanol levels, which led to an increased extraction of wood-derived compounds. Furthermore, the distinct microbial communities also induced changes in the chemical composition of the beer samples, leading to concentration differences in beer- and wood-derived compounds like 4-ethyl guaiacol, 4-ethyl phenol, cis-oak lactone, vanillin, furfural and 5-methyl furfural. Altogether, our results indicate that wood-aging of beer is affected by biotic and abiotic parameters, influencing the quality of the final product. Additionally, this work provides a new, cost-effective approach to study the production of barrel-aged beers based on a simplified microbial community model.


title: New Microviridae isolated from Sulfitobacter reveals two cosmopolitan subfamilies of ssDNA phages infecting marine and terrestrial Alphaproteobacteria

abstract: The Microviridae family represents one of the major clades of ssDNA phages. Their cultivated members are lytic and infect Proteobacteria, Bacteroidetes, and Chlamydiae. Prophages have been predicted in genomes from Bacteroidales, Hyphomicrobiales, and Enterobacteraceae and cluster within the "Alpavirinae", "Amoyvirinae" and Gokushovirinae. We have isolated "Ascunsovirus oldenburgi" ICBM5, a novel phage distantly related to known Microviridae. It infects Sulfitobacter dubius SH24-1b and uses both a lytic and a carrier-state life strategy. Using ICBM5 proteins as a query, we uncovered in publicly available resources 65 new microviridae prophages and episomes in bacterial genomes and retrieved 47 environmental viral genomes (EVGs) from various viromes. Genome clustering based on protein content and phylogenetic analysis showed that ICBM5, together with Rhizobium phages, new prophages, episomes, and EVGs cluster within two new phylogenetic clades, here tentatively assigned the rank of subfamily and named "Tainavirinae" and "Occultatumvirinae". They both infect Rhodobacterales. Occultatumviruses also infect Hyphomicrobiales, including nitrogen-fixing endosymbionts from cosmopolitan legumes. A biogeographical assessment showed that tainaviruses and occultatumviruses are spread worldwide, in terrestrial and marine environments. The new phage isolated here shed light onto new and diverse branches of the Microviridae tree, suggesting that much of the ssDNA phage diversity remains in the dark.


title: In situ architecture of the lipid transport protein VPS13C at ER-lysosomes membrane contacts

abstract: VPS13 is a eukaryotic lipid transport protein localized at membrane contact sites. Previous studies suggested that it may transfer lipids between adjacent bilayers by a bridge-like mechanism. Direct evidence for this hypothesis from a full-length structure and from EM studies in situ, however, is still missing. Here we have capitalized on AlphaFold predictions to complement the structural information already available about VPS13 and to generate a full-length model of human VPS13C, the Parkinsons disease-linked VPS13 paralog localized at contacts between the ER and endo/lysosomes. Such model predicts a ~30-nm rod with a hydrophobic groove that extends throughout its length. We further investigated whether such a structure can be observed in situ at ER-endo/lysosome contacts. To this aim, we combined genetic approaches with cryo-focused-ion-beam (cryo-FIB) milling and cryo-electron tomography (cryo-ET) to examine HeLa cells overexpressing this protein (either full length or with an internal truncation) along with VAP, its anchoring binding partner at the ER. Using these methods we identified rod-like densities that span the space separating the two adjacent membranes and that match the predicted structures of either full length VPS13C or its shorter truncated mutant, thus providing the first in-situ evidence for a bridge-model of VPS13 in lipid transport. Intriguingly, the majority of the VPS13C rods were separated from the ER membranes by a narrow gap, suggesting that while VAP anchors the protein to the ER, direct contact of the VPS13C rod with the ER bilayer to allow lipid transport may be independently regulated.

category: cell biology


title: LIS1 RNA-binding orchestrates the mechanosensitive properties of embryonic stem cells in AGO2-dependent and independent ways

abstract: Lissencephaly-1 (LIS1) is associated with neurodevelopmental diseases and is known to regulate the activity of the molecular motor cytoplasmic dynein. Here we show that LIS1 is essential for the viability of mouse embryonic stem cells (mESCs), and it regulates the physical properties of these cells. LIS1 dosage substantially affects gene expression, and we uncovered an unexpected interaction of LIS1 with RNA and RNA-binding proteins, most prominently the Argonaute complex. We demonstrate that LIS1 overexpression partially rescued the expression of extracellular matrix (ECM) and mechanosensitive genes conferring stiffness to Argonaute null mESCs. Collectively, our data transforms the current perspective on the roles of LIS1 in post- transcriptional regulation underlying development and mechanosensitive processes.


title: PtdIns(3,4)P2, Lamellipodin, and VASP coordinate cytoskeletal remodeling during phagocytic cup formation in macrophages.

abstract: Phosphoinositides are pivotal regulators of vesicular traffic and signaling during phagocytosis. Phagosome formation, the initial step of the process, is characterized by local membrane remodelling and reorganization of the actin cytoskeleton that leads to formation of the pseudopods that drive particle engulfment. Using genetically-encoded fluorescent probes we found that upon particle engagement a localized pool of PtdIns(3,4)P2 is generated by the sequential activities of class I phosphoinositide 3-kinases and phosphoinositide 5-phosphatases. Depletion of the enzymes responsible for this locally generated pool of PtdIns(3,4)P2 blocks pseudopod progression and ultimately phagocytosis. We show that the PtdIns(3,4)P2 effector Lamellipodin (Lpd) is recruited to nascent phagosomes by PtdIns(3,4)P2. Furthermore, we show that silencing of Lpd inhibits phagocytosis and produces aberrant pseudopodia with disorganized actin filaments. Lastly, vasodilator-stimulated phosphoprotein (VASP) was identified as a key actin-regulatory protein mediating phagosome formation downstream of Lpd. Mechanistically, our findings imply that a pathway involving PtdIns(3,4)P2, Lpd and VASP mediates phagocytosis at the stage of particle engulfment.


title: Predicting plant-related and leaf-related parameters of winter wheat using vegetation index: Should phenological correction be applied across growth stages?

abstract: Most existing plant-related and leaf-related parameters models of winter wheat vary across growing seasons, but it is an open question whether a unified statistical model can be developed to predict plant-related and leaf-related parameters using VI across multiple growing seasons, or whether the phenological correction is necessary for each parameter across multiple growing seasons. To explore this question, we measured two plant-related parameters and four leaf-related parameters over five growth stages during the 2017-2021 growing seasons. A hierarchical linear model (HLM) automatically adapts the relationship between VIs and their corresponding parameter across growing seasons and assesses the contribution of phenological variables by applying a sensitivity analysis. The estimates of VI-plant-related parameters [aboveground dry biomass (AGB) and plant nitrogen concentration (PNC)] were scattered over a given growing season, unlike the relationship between VI-leaf-related parameters [leaf dry biomass (LGB), leaf nitrogen concentration (LNC), leaf area index (LAI) and soil and plant analysis development (SPAD)]. In contrast, the AGB, PNC, LGB, LNC, LAI, and SPAD HLM models are stable and can be popularized across growing seasons, with the determination coefficient R2 ranging from 0.84 to 0.86, 0.79 to 0.87, 0.70 to 0.71, 0.68 to 0.86, 0.75 to 0.81, and 0.68 to 0.70, respectively, and the root mean square error ranging from 0.13 to 0.50 t/ha, 0.01 to 0.07%, 0.01 to 0.03 t/ha, 0.02 to 0.03%, 0.02 to 0.07, and 0.47 to 0.69, respectively. The sensitivity index of the phenological information in the AGB and PNC models was 0.56-0.78 and 0.66-0.72, respectively, whereas that in the LGB, LNC, LAI, and SPAD models was 0.01-0.06, 0.01-0.10, 0.02-0.06, and 0.00-0.01, respectively. Although phenological effects have little effect on leaf-related indicators, HLM has a strong potential for application to other crops and regions.

category: ecology


title: Community diversity is associated with intra-species genetic diversity and gene loss in the human gut microbiome

abstract: The human gut microbiome contains a diversity of microbial species that varies in composition over time and across individuals. These species are comprised of diverse strains, which are known to evolve by mutation and recombination within hosts. How the ecological process of community assembly interacts with sub-species diversity and evolutionary change is a longstanding question. Two hypotheses have been proposed based on ecological observations and theory: Diversity Begets Diversity (DBD), where taxa tend to become more diverse in already diverse communities, and Ecological Controls (EC), where higher community diversity impedes diversification within taxa. Recently we showed with 16S rRNA gene amplicon data from the Earth Microbiome Project that DBD is detectable in natural bacterial communities from a range of environments at high taxonomic levels (ranging from phylum to species-level), but that this positive relationship between community diversity and within-taxon diversity plateaus at high levels of community diversity. Whether increasing community diversity is associated with sub-species genetic diversity within microbiomes, however, is not yet known. To test the DBD and EC hypotheses at a finer genetic resolution, we analyzed sub-species strain and nucleotide variation in static and temporally sampled shotgun sequenced fecal metagenomes from a panel of healthy human hosts. We find that both sub-species single nucleotide variation and strain number are positively correlated with community diversity, supporting DBD. We also show that higher community diversity predicts gene loss in a focal species at a future time point and that community metabolic pathway richness is inversely correlated with the pathway richness of a focal species. These observations are consistent with the Black Queen Hypothesis, which posits that genes with functions provided by the community are less likely to be retained in a focal species genome. Together, our results show that DBD and Black Queen may operate simultaneously in the human gut microbiome, adding to a growing body of evidence that these eco-evolutionary processes are key drivers of biodiversity and ecosystem function.

category: evolutionary biology


title: On-Farm Genetic Diversity of Wheat (Triticum aestivum spp.) In Digalu Tijo District, Arsi Zone, Ethiopia

abstract:
IntroductoryThis study was undertaken to assess the on-farm genetic diversity of wheat Landraces (Triticum aestivum spp) in Digalu and Tijo District.

MethodsFor this study, three farmer households kebeles were selected by cluster sampling (area sampling) method because they are found in the most dominant wheat landraces growing agro-ecological areas. Data for this study was collected using questionnaires, field observation, interviews, focus group discussion, and document analysis.

Results and DiscussionsIn total, 27 wheat landraces were reported to be grown before two decades to the present day. Of these, only 9 of wheat landraces are still cultivated by farmers. Among these,  Qamadi guracha is the most common growing landraces in the study area. The Shannon diversity index (H) of growing wheat landraces ranges from 1.09 to 1.37 among the groups. The estimated genetic erosion for wheat landraces was found to be 66.7% due to the major factors: improved wheat varieties; the introduction of other more productive crops and wheat landraces low productivity.

ConclusionsFood quality, Pests (example, birds), diseases resistance, market value, and straw quality were factors that initiated the farmers to maintain the genetic diversity of landraces on their farmlands. However, the preservation of wheat landraces are influenced by bottlenecks like the seed selection system, and insufficient crop yield.

RecommendationRegeneration of soil fertility, restoring of lost landraces, improvement of landraces, on-farm conservation by re-sowing, saving of seeds for future, and ex-situ conservation is suggested for the restoration of wheat landraces diversity in Digalu and Tijo District.

category: genetics


title: Diversity of Learning to Control Complex Rehabilitation Robots Using High-Dimensional Interfaces

abstract: Learning to perform everyday tasks, using a complex robot, presents a nested problem. It is nested, because, on the surface, there is a problem of robot control--but within it, there lies a deeper, more challenging problem that demands the control nuances necessary to perform complicated functional tasks. For individuals with limited mobility, such as those with cervical spinal cord injuries, the addition of physical burden is added to this motor learning burden. An explicit training regime can be designed to accelerate and aid the learning process, with the long-term aim to help individuals (injured or uninjured) acquire the skill of complex robot control. However, such training regimes are not well-established nor are the methods of evaluation. In this paper, we gain a baseline understanding of how humans learn to control a 7 degree-of-freedom assistive robotic arm, using a novel high-dimensional interface, in the absence of explicit training. We examine how participants transition between distinct workspace zones to extract their learning possibilities. We gain additional granularity in individual learning, based on how participants spend their time in the workspace, with the robot, and how the time spent is distributed across trials. These analyses highlight the high diversity of learning. Lastly, we provide benefits and opportunities for targeted training regimes that are explicit and heavily favor individualized support.

category: bioengineering


title: Powerful and interpretable control of false discoveries in differential expression studies

abstract:
MotivationThe standard approach for statistical inference in differential expression (DE) analyses is to control the False Discovery Rate (FDR). However, controlling the FDR does not in fact imply that the proportion of false discoveries is upper bounded. Moreover, no statistical guarantee can be given on subsets of genes selected by FDR thresholding. These known limitations are overcome by post hoc inference, which provides guarantees of the number of proportion of false discoveries among arbitrary gene selections. However, post hoc inference methods are not yet widely used for DE studies.

ResultsIn this paper, we demonstrate the relevance and illustrate the performance of adaptive interpolation-based post hoc methods for DE studies. First, we formalize the use of permutation-based methods to obtain sharp confidence bounds that are adaptive to the dependence between genes. Then, we introduce a generic linear time algorithm for computing post hoc bounds, making these bounds applicable to large-scale DE studies. The use of the resulting Adaptive Simes bound is illustrated on a RNA sequencing study. Comprehensive numerical experiments based on real microarray and RNA sequencing data demonstrate the statistical performance of the method.

AvailabilityA cross-platform open source implementation within the R package sanssouci is available at https://pneuvial.github.io/sanssouci/.

category: bioinformatics


title: RUNX1 isoform disequilibrium in the development of trisomy 21 associated myeloid leukemia

abstract:
Aneuploidy is a hallmark of cancer, but its complex nature limits our understanding of how it drives oncogenesis. Gain of chromosome 21 (Hsa21) is among the most frequent aneuploidies in leukemia and is associated with markedly increased leukemia risk. Here, we propose that disequilibrium of the RUNX1 isoforms is key to the pathogenesis of trisomy 21 (i.e. Down syndrome)-associated myeloid leukemia (ML-DS). Hsa21-focused CRISPR-Cas9 screens uncovered a strong and specific RUNX1 dependency in ML-DS. Mechanistic studies revealed that excess of RUNX1A isoform - as seen in ML-DS patients - synergized with the pathognomonic Gata1s mutation in leukemogenesis by displacing RUNX1C from its endogenous binding sites and inducing oncogenic programs in complex with the MYC cofactor MAX. These effects were reversed by restoring the RUNX1A:RUNX1C equilibrium or pharmacological interference with MYC:MAX dimerization. Our study highlights the importance of alternative splicing in leukemogenesis, even on a background of aneuploidies, and opens new avenues for developing specific and targeted therapies.

Graphical Abstract

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category: cancer biology


title: The importance of environmental parameters and mixing zone in shaping estuarine microbial communities along a freshwater-marine gradient.

abstract: Microbial communities are important elements in the marine environment, contributing to nutrient cycling and biogeochemical processes. Estuaries comprise environments exhibiting characteristics from freshwater to marine, leading to distinct microbial communities across this environmental gradient. Here, we examine the spatial dynamics of microbial communities in Macquarie Harbour, an estuarine system on the West coast of Tasmania, Australia. Water was sampled along the estuary to explore the structure and composition of the microbial communities using 16S/18S rRNA gene amplicon sequencing. Multivariate analyses showed environmental variables and community compositions varying along a longitudinal (river to adjacent ocean) gradient at the surface. In the harbour, differences in the microbial community were observed between surface (0-1 m) and intermediate depths (4.5-11 m depth). The results of differential abundance, network and Partial Least Square analyses suggest that Macquarie Harbour is a mixing zone, where the distributions of archaeal, bacterial and eukaryotic communities are influenced by oceanic and riverine inputs. Coupled with the natural characteristics of the Harbour, the heterotrophic component of this microbial communities inhabiting the surface and intermediate waters may play important roles in the nutrient cycle in the studied area. These results provide critical insights into the Macquarie Harbour environment and the importance of understanding the role of microbial communities for similar systems elsewhere.


title: Activation of the cGAS-STING innate immune response in cells with deficient mitochondrial topoisomerase TOP1MT

abstract: Systemic lupus erythematosus (SLE) is an autoimmune condition where the underlying dysfunction is often unknown. Here, we identify a novel link between the mitochondrial topoisomerase TOP1MT and SLE, as a novel variant predicted to affect TOP1MT function, P193L, was identified in a family with SLE and other autoimmune diseases. Although there was no previous genetic association between TOP1MT and SLE, the role of TOP1MT as a regulator of mtDNA led us to investigate whether TOP1MT could mediate the release of mtDNA to the cytosol, where it could then activate the cGAS-STING innate immune pathway. This pathway increases the expression of type-I interferons and is known to be activated in some patients with SLE. Through analysis of cells lacking TOP1MT, or re-expressing the P193L variant, we established a new association by which TOP1MT dysfunction leads to increased release of mtDNA to the cytosol, causing activation of the cGAS-STING pathway. We also characterized the P193L variant for its ability to rescue several TOP1MT functions in TOP1MT knockout cells. While re-expression of the P193L variant was able to rescue steady state mtDNA copy number, most functions investigated only showed partial rescue, including repletion of mtDNA replication following depletion, nucleoid size, steady state mtDNA transcripts levels, and mitochondrial morphology. Meanwhile, the P193L variant failed to rescue decreased mitochondrial respiration. Overall, these findings support the notion that P193L variant is not fully functional and likely influences susceptibility to SLE via cGAS-STING mediated activation of the innate immune system.


title: The influence of learning on robustness of working memory to distractors

abstract: Inhibition of irrelevant information is a crucial process within the working memory (WM) system. In the present study, we explored how inhibition of distractors is learned within a Sternberg-like WM task and how this learning process affects readout performance. We introduced distractors of varying strength as well as novel stimuli to a classic Sternberg task. Our findings show that the length of the training period affected obstructiveness of distractors as well as differences between WM loads. Significant differences between conditions observed early on in accuracy became void after general ceiling performance was reached. Moreover, accuracy and reaction times, which are commonly used as readouts of WM performance, were affected differently by training. While both improved significantly, only accuracy exhibited differential dependency of experimental conditions on training. It is essential to consider these dynamics when introducing a WM task which requires a long training period.


title: Hetero-bivalent Nanobodies Provide Broad-spectrum Protection against SARS-CoV-2 Variants of Concern including Omicron

abstract: Following Delta, Omicron variant triggered a new wave of SARS-CoV-2 infection globally, adaptive evolution of the virus may not stop, the development of broad-spectrum antivirals is still urgent. We previously developed two hetero-bivalent nanobodies with potent neutralization against original WT SARS-CoV-2, termed aRBD-2-5 and aRBD-2-7, by fusing aRBD-2 with aRBD-5 or aRBD-7, respectively. Here, we resolved crystal structures of these nanobodies in complex with RBD, and found the epitope of aRBD-2 differs from that of aRBD-5, aRBD-7. aRBD-2 binds to a conserved epitope which renders its binding activity to all variants of concern (VOCs) including Omicron. Interestingly, although monovalent aRBD-5 and aRBD-7 lost binding to some variants, they effectively improved the overall affinity when transformed into the hetero-bivalent form after being fused with aRBD-2. Consistent with the high binding affinities, aRBD-2-5-Fc and aRBD-2-7-Fc exhibited ultra-potent neutralization to all five VOCs; particularly, aRBD-2-5-Fc neutralized authentic virus of Beta, Delta and Omicron with the IC50of 5.98[~]9.65 ng/mL or 54.3[~]87.6 pM. Importantly, aRBD-2-5-Fc provided in vivo prophylactic protection for mice against WT and mouse-adapted SARS-CoV-2, and provided full protection against Omicron in hamster model when administrated either prophylactically or therapeutically. Taken together, we found a conserved epitope on RBD, and hetero-bivalent nanobodies had increased affinity for VOCs over its monovalent form, and provided potent and broad-spectrum protection both in vitro and in vivo against all tested major variants, and potentially future emerging variants. Our strategy provides a new solution in the development of therapeutic antibodies for COVID-19 caused by newly emergent VOCs.

category: immunology


title: Efficacy of plasmid-encoded CRISPR-Cas antimicrobial is affected by competitive factors found in wild Enterococcus faecalis isolates

abstract:
Enterococcus faecalis is a leading cause of hospital-acquired infections. These infections are becoming more difficult to treat due to the increasing emergence of E. faecalis strains resistant to last resort antibiotics. Over the past decade, multiple groups have engineered the naturally occurring bacterial defense system CRISPR-Cas as a sequence-specific antimicrobial to combat antibiotic-resistant bacteria. We have previously established that the type II CRISPR-Cas system of E. faecalis can be reprogrammed as a CRISPR-Cas antimicrobial and delivered to antibiotic-resistant recipients on a conjugative pheromone-responsive plasmid. Using a co-culture system, we showed sequence-specific depletion of antibiotic resistance from E. faecalis model strains, both in vitro and in vivo. Although this and other studies have demonstrated the potential use for CRISPR-Cas as an antimicrobial, most have deployed the system against model bacterial strains. Thus, there is limited knowledge on how effective these potential therapies are against recently isolated and uncharacterized strains with limited laboratory passage, which we refer to here as wild strains. Here, we compare the efficacy of our previously established CRISPR-Cas antimicrobials against both E. faecalis model strains and wild E. faecalis fecal isolates. We demonstrate that these wild isolates can antagonize the CRISPR-Cas antimicrobial donor strain via competitive factors like cytolysin. Furthermore, we show that the wild isolates can effectively prevent delivery of the CRISPR-Cas antimicrobial plasmids, consequently avoiding CRISPR-Cas targeting. Our results emphasize the requisite to study CRISPR-Cas antimicrobials against wild strains to understand limitations and develop delivery systems that can endure competitive interspecies interactions in the gut microenvironment and effectively deliver CRISPR-Cas antimicrobials to their intended targets.

IMPORTANCEEnterococcus faecalis is a major nosocomial pathogen. Traditional antibiotics continue to lose potency against these opportunistic pathogens as they become increasingly resistant to more antibiotics. We previously showed that our CRISPR-Cas antimicrobials can deplete drug resistance in or kill E. faecalis model strains. Here, we examined the efficacy of CRISPR-Cas antimicrobials against a recent collection of E. faecalis fecal isolates. We found that CRISPR-Cas delivery and efficacy is affected by competitive factors produced by the wild isolates. Our study emphasizes the need to study CRISPR-Cas antimicrobials in the context of wild bacterial isolates, which are the intended target for this potential therapy, in order to understand limitations and develop CRISPR-enhanced antimicrobials with effective clinical applications.


title: Extensive neutralization against SARS-CoV-2 variants elicited by Omicron-specific subunit vaccine booster

abstract: The currently dominant variant of SARS-CoV-2 Omicron, carrying a great number of mutations, has been verified its strong capacity of immune escape in COVID-19 convalescents and vaccinated individuals. An increased risk of SARS-CoV-2 reinfection or breakthrough infection should be concerned. Here we reported higher humoral immune response elicited by Delta and Omicron variants after breaking through previous infection and cross-neutralization against VOCs, compared to the ancestral wild-type (WT) virus infection. To overcome the immune escape of Omicron, Omicron-specific vaccine was considered as a novel and potential strategy. Mouse models were used to verify whether Omicron-specific RBD subunit boost immune response by immunizing Omicron-RBD recombinant proteins. Three doses of Omicron-RBD immunization elicit comparable neutralizing antibody (NAb) titers with three doses of WT-RBD immunization, but the neutralizing activity was not cross-active. By contrast, two doses of WT-RBD with an Omicron-RBD booster increased the NAb geometric mean titers against Omicron by 9 folds. Moreover, an additional boost vaccination with Omicron-RBD protein could increase humoral immune response against both WT and current VOCs. These results suggest that the Omicron-specific subunit booster shows its advantages in the immune protection from both WT and current VOCs, and that SARS-CoV-2 vaccines administration using two or more virus lineages as antigens might improve the NAb response.


title: Self-cleaving 2A peptides allow for expression of multiple genes in Dictyostelium discoideum

abstract: The social amoeba Dictyostelium discoideum is a model for a wide range of biological processes including chemotaxis, cell-cell communication, phagocytosis, and development. Interrogating these processes with modern genetic tools often requires the expression of multiple transgenes. While it is possible to transfect multiple transcriptional units, the use of separate promoters and terminators for each gene leads to large plasmid sizes and possible interference between units. In other eukaryotic systems this challenge has been addressed through polycistronic expression mediated by 2A viral peptides, permitting efficient, co-regulated gene expression. Here, we screen the most commonly used 2A peptides, porcine teschovirus-1 2A (P2A), Thosea asigna virus 2A (T2A), equine rhinitis A virus 2A (E2A), and foot-and-mouth disease virus 2A (F2A), for activity in D. discoideum. We find that these 2A sequences are effective but that, unexpectedly, combining the coding sequences of two proteins into a single transcript leads to notable decreases in expression level with strain-dependent magnitudes, suggesting unexplored constraints of gene expression in D. discoideum that merit further investigation. Our results show that P2A is the optimal sequence for polycistronic expression, opening up new possibilities for genetic engineering in this model system.

category: molecular biology


title: Superoxide Dismutase 1 translocates to the nucleus in spinal cord mixed cells from newborn rats

abstract: Superoxide dismutase 1 (SOD1) catalyzes the superoxide conversion to oxygen and hydrogen peroxide in general. SOD1 can translocate to the cell nucleus in response to oxidative stress in yeast and human fibroblasts. Here, we report the translocation of sod1 to the cell nucleus in primary co-cultures of neurons and astrocytes derived from newborn rats explants even in the absence of oxidative stress stimuli. The successful tissue explants from rats allowed simplistic and clean modeling of Amyotrophic lateral sclerosis (ALS)-simile tissue microenvironment. This mixed-cells-population was responsive to H2O2 (100 M) oxidative stress with a progressive dying process up to 6 h after stimuli. However, no differences in SOD1 nucleus translocation were observed in rat amyotrophic lateral sclerosis (ALS)-simile tissue microenvironment up to 6 hours under oxidative stress. These results altogether point to a robust cellular collaboration between neurons and astrocytes so that the tissue can equilibrate this SOD1 translocation during the oxidative stress response and in the early animal development.

category: biochemistry


title: Quantitative interactome proteomics identifies proteostasis network for GABAA receptors

abstract: Gamma-aminobutyric acid type A (GABAA) receptors, the primary inhibitory neurotransmitter-gated ion channels in the mammalian central nervous system, inhibit neuronal firing to preserve balanced neuronal activity. Maintenance of GABAA receptor protein homeostasis (proteostasis) in the cell utilizing its interacting proteins is essential for the function of GABAA receptors. However, how the proteostasis network orchestrates GABAA receptor biogenesis in the endoplasmic reticulum (ER) is not well understood. To address this question systematically, we employed a proteomics-based approach to identify the interactomes of GABAA receptors by carrying out a quantitative immunoprecipitation-tandem mass spectrometry (IP-MS/MS) analysis utilizing stable isotope labeling by amino acids in cell culture (SILAC). To enhance the coverage and reliability of the identified proteins, we performed comparative proteomics by using both wild type 1 subunit and a misfolding-prone 1 subunit carrying the A322D variant as the bait proteins. The wild type 1 interactome contains 125 proteins, the 1(A322D) interactome contains 105 proteins, and 54 proteins overlap within two interactomes. Bioinformatics analysis identified potential GABAA receptor proteostasis network components, including chaperones, folding enzymes, trafficking factors, and degradation factors. Further, their potential involvement is modelled in the cellular folding, degradation and trafficking pathways for GABAA receptors. In addition, we verified endogenous interactions between 1 subunit and their selected interactors by carrying out co-immunoprecipitation assay in mouse brain homogenates. This study paves the way for understanding the molecular mechanisms as well as fine-tuning of GABAA receptor proteostasis to ameliorate related neurological diseases such as epilepsy.


title: Inter- and intra-Subunit interactions driving the MukBEF ATPase

abstract: MukBEF, an SMC-like protein complex, is the bacterial condensin. It is likely that it compacts DNA via an ATP hydrolysis-dependent, DNA loop-extrusion reaction as has been demonstrated for the yeast SMC proteins condensin and cohesin. MukB also interacts with the ParC subunit of the cellular chromosomal decatenase topoisomerase IV, an interaction that is required for proper chromosome condensation and segregation in E. coli, although it suppresses the MukB ATPase activity. We have investigated the MukBEF ATPase activity, identifying inter- and intra-subunit interactions by protein-protein crosslinking and site-specific mutagenesis. We show that interactions between the hinge of MukB and its neck region are essential for the ATPase activity, that the ParC subunit of Topo IV inhibits the MukB ATPase by preventing this interaction, that MukE interaction with DNA is likely essential for viability, and that interactions between MukF and the MukB neck region are necessary for ATPase activity and viability.


title: A non-conventional archaeal fluorinase identified by in silico mining catalyzes the fastest natural nucleophilic fluorination

abstract:
Fluorinases, the only enzymes known to catalyze the transfer of fluorine to an organic molecule, are essential biocatalysts for the sustainable synthesis of valuable organofluorines. However, the few fluorinases identified so far have low turnover rates that hamper biotechnological applications. Here, we isolated and characterized putative fluorinases retrieved from systematic in silico mining and identified a non-conventional archaeal enzyme from Methanosaeta sp. that mediates the highest fluorination rate reported to date. Furthermore, we demonstrate enhanced production of fluoronucleotides in vivo in a bacterial host engineered with this archaeal fluorinase, paving the way towards synthetic metabolism for efficient biohalogenations.

Fastest biological SN2 fluorination

category: synthetic biology


title: Comparative Genomic Analysis of Salmonella enterica serovar Typhimurium from Passerines Reveals Two Lineages Circulating in Europe, New Zealand, and the United States

abstract:
Salmonella enterica serovar Typhimurium from passerines have caused wild bird mortality and human salmonellosis outbreaks in Europe, Oceania, and North America. Here, we performed comparative genomic analysis to explore the emergence, genetic relationship, and evolution of geographically dispersed passerine isolates. We found that passerine isolates from Europe and the United States clustered to form two lineages (EU and US passerine lineages), which were distinct from major S. Typhimurium lineages circulating in other diverse hosts (e.g., humans, cattle, pigs, chicken, other avian hosts such as pigeons and ducks). Further, passerine isolates from New Zealand clustered to form a sublineage (NZ passerine lineage) of the US passerine lineage. We inferred that the passerine isolates mutated at a rate of 3.2 x 10-7 substitutions/site/year, and the US, EU, and NZ passerine lineages emerged in ca. 1952, 1970, and 1996, respectively. Isolates from the three lineages presented genetic similarity such as lack of antimicrobial resistance genes and accumulation of same virulence pseudogenes. In addition, genetic diversity due to microevolution existed in the three passerine lineages. Specifically, pseudogenization in type 1 fimbrial gene fimC (deletion of G at position 87) was only detected in the US and NZ passerine isolates, while a single-base deletion in type 3 secretion system effector genes (i.e., gogB, sseJ, and sseK2) solely concurred in the EU passerine isolates. These findings provide insights into evolution, host adaptation, and epidemiology of S. Typhimurium in passerines.

IMPORTANCEPasserine-associated S. Typhimurium have been linked to human salmonellosis outbreaks in recent years. Here we investigated the phylogenetic relationship of globally distributed passerine isolates and profiled their genomic similarity and diversity. Our study reveals two passerine-associated S. Typhimurium lineages circulating in Europe, Oceania, and North America. Isolates from the two lineages presented phylogenetic and genetic signatures that were distinct from isolates of other hosts. The findings shed light on host adaptation of S. Typhimurium in passerines and are important for source attribution of S. Typhimurium to avian hosts. Further, we found S. Typhimurium definitive phage type (DT) 160 from passerines that caused decade-long human salmonellosis outbreaks in New Zealand and Australia formed a sublineage of the US passerine lineage, suggesting that DT160 may have originated from passerines outside Oceania. Our study demonstrates the importance of whole-genome sequencing and genomic analysis of historical microbial collections to modern day epidemiologic surveillance.


title: Complete identity of the cis-asRNA StfZ and its influence on FtsZ protein level and cell division in Escherichia coli

abstract: Bacteria regulate FtsZ protein levels through transcriptional and translational mechanisms for proper cell division. A cis-antisense RNA, StfZ, produced from the ftsA-ftsZ intergenic region was proposed to regulate the FtsZ level in Escherichia coli. However, its structural identity remained unknown. In the present study, we determined the complete sequence, identified the promoters, and effects of StfZ on the FtsZ level. We show that StfZ is expressed at 1:6 ratio of StfZ:ftsZ mRNA at all growth phases from three promoters as three isoforms of 366 nt, 474 nt, and 552 nt. Overexpression of StfZ reduces FtsZ level, increases cell size, and blocks cell division. Thus, the cis-encoded StfZ emerges as a novel factor for maintaining the FtsZ level at different growth phases in E. coli.


title: Dynamic changes of gut microbiota associated with blood glucose levels in sows from pregnancy through lactation

abstract: Gut microbiota plays important roles in metabolism and physiological homeostasis throughout the lifespan of mammals, including in pregnancy. To evaluate the effects of practically adopted restricted feeding during gestation on blood glucose level and fecal microbiota composition, 31 sows were used to identify the shifts in bacterial assemblages by 16S rRNA sequencing in their early pregnancy, late pregnancy and late lactation stages. We find that the blood glucose was elevated significantly (p < 0.001) in sows offered abundant feed in the late lactation as compared with those of the same cohort in the pregnancy stages when on limited feed. Five bacterial genera were differentially abundant among three stages, and the abundances of Streptococcus (padj < 0.001) and unclassified_f_Lachnospiraceae (padj < 0.001) were increased, whereas the abundances of Lactobacillus (padj = 0.035) and Escherichia-Shigella (padj< 0.001) were decreased in the latter two stages. The genus Prevotellaceae_NK3B31_group (padj = 0.026) displayed higher abundance in the late lactation stage. A significant correlation in the Pearsons analysis indicates that three out of these five differentially abundant genera were related to blood glucose change. Our data show that restricted-fed gestating sows have aberrant blood glucose level and may affect the intestinal microbiota.


title: Reprogrammed Schwann cells organize into dynamic tracks that promote pancreatic cancer invasion

abstract:
Nerves are a component of the tumor microenvironment contributing to cancer progression, but the role of cells from nerves in facilitating cancer invasion remains poorly understood. Here we show that Schwann cells (SCs) activated by cancer cells collectively function as Tumor Activated Schwann cell Tracks (TASTs) that promote cancer cell migration and invasion. Non-myelinating SCs form TASTs and have cell gene expression signatures that correlate with diminished survival in patients with pancreatic ductal adenocarcinoma. In TASTs, dynamic SCs form tracks that serve as cancer pathways and apply forces on cancer cells to enhance cancer motility. These SCs are activated by c-Jun, analogous to their reprogramming during nerve repair. This study reveals a mechanism of cancer cell invasion that co-opts a wound repair process and exploits the ability of SCs to collectively organize into tracks. These findings establish a novel paradigm of how cancer cells spread and reveal therapeutic opportunities.

SIGNIFICANCEHow the tumor microenvironment participates in pancreatic cancer progression is not fully understood. Here, we show that Schwann cells are activated by cancer cells and collectively organize into tracks that dynamically enable cancer invasion in a c-Jun dependent manner.


title: Lipocalin 2 upregulation mediates the detrimental effects of therapy-induced senescence in breast cancer cells.

abstract: The acquisition of novel detrimental cellular properties following exposure to cytotoxic drugs leads to aggressive and metastatic tumors that often translates into an incurable disease. While the bulk of the primary tumor is eliminated upon exposure to chemotherapeutic treatment, residual cancer cells and non-transformed cells within the host can engage a stable cell cycle exit program named senescence. Senescent cells secrete a distinct set of pro-inflammatory factors, collectively termed the senescence-associated secretory phenotype (SASP). Upon exposure to the SASP, cancer cells undergo cellular plasticity resulting in increased proliferation, migration and epithelial-to-mesenchymal transition. The molecular mechanisms by which the SASP regulates these pro-tumorigenic features are poorly understood. Here, we report that breast cancer cells exposed to the SASP strongly upregulate Lipocalin 2 (LCN2). Furthermore, we demonstrate that LCN2 is critical for SASP-induced increased migration in breast cancer cells, and its inactivation potentiates the response to chemotherapeutic treatment in mouse models of breast cancer. Finally, we show that neoadjuvant chemotherapy treatment leads to LCN2 upregulation in residual human breast tumors, and correlates with worse overall survival. These findings provide the foundation for targeting LCN2 as an adjuvant therapeutic approach to prevent the emergence of aggressive tumors following chemotherapy.


title: Bayesian multi-model-based 13C15N-metabolic flux analysis quantifies carbon-nitrogen metabolism in mycobacteria

abstract: Metabolic flux is the final output of cellular regulation and has been extensively studied for carbon but much less is known about nitrogen, which is another important building block for living organisms. For the pathogen Mycobacterium tuberculosis (Mtb), this is particularly important in informing the development of effective drugs targeting Mtbs metabolism. Here we performed 13C15N dual isotopic labelling of mycobacterial steady state cultures and quantified intracellular carbon-nitrogen (CN) and nitrogen (N) fluxes in addition to carbon (C) fluxes and inferred their reaction bidirectionalities. The combination of 13C15N-MFA with a Bayesian multi-model approach allowed us to resolve C and N fluxes simultaneously which was not possible with classical 13C-MFA. We quantified CN fluxes for amino acid and, for the first time, nucleotide biosynthesis. Our analysis identified glutamate as the central CN and N node in mycobacteria, and improved resolution of the anaplerotic node. Our study describes a powerful platform to measure carbon and nitrogen metabolism in any biological system with statistical rigor.


title: Mitochondrial TFAM in the Regulation of FOXP3+ T cell Mediated Maintenance of CD4+ T Cell Landscapes

abstract: The immune system deteriorates as we age. Regulatory T cells contribute to immunosenescence as they critically mold antigenic CD4+ T cell subsets and maintain immunological homeostasis. Given the role of mitochondrial Transcription Factor A (TFAM) in oxidative stress, cellular senescence, and age-related pathologies, we explored the role of this factor in Regulatory T (Treg) cells. We employed a conditional deletion of FoxP3-specific TFAM to create Regulatory T (Treg) cells lacking mitochondrial TFAM. Using comprehensive single-cell RNA cell technology, we identified how mitochondrial TFAM in Treg cells controls the CD4+ T cell fate and trajectory for a heterologous pool of inflammatory CD4 cytotoxic, CD4 exhausted, and CD4 effector memory T cells, all features of aging. Additionally, our results show how TNF signalling and related cell death pathways exacerbate systemic inflammation, degradation, and acceleration of cell death in mice lacking TFAM. Furthermore, partial TFAM restoration or use of the antioxidative drug resveratrol alleviate the inflammaging cascade. These findings reveal TFAM as a critical regulator of Foxp3 Treg cell activity in sculpting the CD4 heterologous pool and point to TFAM as a possible target for pharmacological manipulation in age-related diseases.


title: Mechanical Ventilation induced P21 in an Acute Aging Model of Lung Injury

abstract: Acute respiratory distress syndrome (ARDS) is a form of acute lung injury which leads to a paucity of oxygen. To remedy ARDS, patients are put on mechanical ventilators; however, the stretch resulting from the mechanical ventilator can lead to ventilator-induced lung injury or VILI. VILI is exacerbated by age. The combined effects of ARDS and mechanical ventilation can result in a hostile environment which may lead to senescence (stable cell cycle arrest). The role of senescence in VILI is poorly understood. Senescence is characterized by increased cyclin-dependent kinase inhibitors P16 and P21; however, P21 has been shown to occur in early senescence. We hypothesized that mechanical ventilation would lead to DNA damage and senescence-like phenotype. Both in vivo and in vitro models of VILI were used to investigate senescence and its mechanism in VILI. Mechanical ventilation increased senescence-associated markers such as DNA damage marker characterized by {gamma}H2AX, P21, senescence-associated secretory phenotype IL6, and decreased proliferation. Moreover, mechanical ventilation led to increased apoptosis. Lung sections were stained for KRT8 proteins, markers of transiently differentiated alveolar type 2 (AT2) cells which were reported to be more prone to DNA damage. Age and mechanical ventilation increased KRT8 positive cells. Finally, we probed a potential mediator of the stretch induced senescence in vitro by inhibiting P38-MAPK, which can be activated by DNA damage response, leading to increased P21. Inhibiting P38 decreased in P21 but did not decrease {gamma}H2AXThese findings suggest that mechanical ventilation may lead to a senescence-like phenotype involving the P38-MAPK pathway.


title: Distinct phospho-variants of STAT3 regulate naïve pluripotency and developmental pace in vivo

abstract: STAT3 has been studied extensively in the context of self-renewal of naive pluripotent mouse embryonic stem cells. We uncovered acute roles for STAT3 and its target, TFCP2L1, in maintenance of epiblast and primitive endoderm during in vivo diapause. On an outbred genetic background, we observed consistent developmental retardation from implantation until embryonic day 11.5, beginning with significant reduction of epiblast cells at implantation in Stat3 null embryos. Remarkably, mutants closely resemble non-affected embryos from the previous day at all postimplantation stages examined. We attribute this phenotype to loss of the active serine phosphorylated form of STAT3 required for neural differentiation and implicated in growth defects in mice and humans. Bulk RNA-sequencing analysis of isolated epiblasts revealed compromised lipid metabolism in Stat3 null embryos by embryonic day 6.5. Furthermore, we demonstrate that gastruloids generated from Stat3 null ESCs failed to extend the posterior axis or maintain BRACHYURY expression and were underrepresented in this region when mixed with wild type cells in chimaeric gastruloids. Our study implicates a role for STAT3 in temporal control of embryonic progression and metabolic mechanisms.

category: developmental biology


title: Rescue by gene swamping as a gene drive deployment strategy

abstract: Gene drives are genetic constructs that can spread deleterious alleles with potential application to population suppression of harmful species. Given that a gene drive can potentially spill over to other populations or even other species, control measures and fail-safes strategies must be considered. Gene drives are designed to generate a rapid demographic decline, while at the same time generating a dynamic change in the populations genetics. Since these evolutionary and demographic processes are linked and are expected to occur at a similar time-scale during gene drive spread, feedback between these processes may significantly affect the outcome of deployment. To study this feedback and to understand how it affects gene drive spillovers, we developed a gene drive model that combines evolutionary and demographic dynamics in a two-population setting. The model demonstrates how feedback between evolutionary and demographic dynamics can generate additional outcomes to those generated by the evolutionary dynamics alone. We identify an outcome of particular interest, where the short-term suppression of the target population is followed by gene swamping and loss of the gene drive. This outcome could be useful for designing gene drive deployments that temporarily suppress the population, but ultimately do not remain in the population. Using our model, we demonstrate the robustness of this outcome to spillover and to the evolution of resistance, and suggest that it could be used as a fail-safe strategy for gene drive deployment.


title: AlphaFold2 fails to predict protein fold switching

abstract: AlphaFold2 has revolutionized protein structure prediction by leveraging sequence information to rapidly model protein folds with atomic-level accuracy. Nevertheless, previous work has shown that these predictions tend to be inaccurate for structurally heterogeneous proteins. To systematically assess factors that contribute to this inaccuracy, we tested AlphaFold2s performance on 98 fold-switching proteins, which assume at least two distinct-yet-stable secondary and tertiary structures. Topological similarities were quantified between five predicted and two experimentally determined structures of each fold-switching protein. Overall, 94% of AlphaFold2 predictions captured one experimentally determined conformation but not the other. Despite these biased results, AlphaFold2s estimated confidences were moderate-to-high for 74% of fold-switching residues, a result that contrasts with overall low confidences for intrinsically disordered proteins, which are also structurally heterogeneous. To investigate factors contributing to this disparity, we quantified sequence variation within the multiple sequence alignments used to generate AlphaFold2s predictions of fold-switching and intrinsically disordered proteins. Unlike intrinsically disordered regions, whose sequence alignments show low conservation, fold-switching regions had conservation rates statistically similar to canonical single-fold proteins. Furthermore, intrinsically disordered regions had systematically lower prediction confidences than either fold-switching or single-fold proteins, regardless of sequence conservation. AlphaFold2s high prediction confidences for one fold-switching conformation corroborate previous work showing that machine-learning-based structure predictors fail to capture other fundamental biophysical features of proteins such as their folding pathways. Our results emphasize the need to look at protein structure as an ensemble and suggest that systematic examination of fold-switching sequences may reveal propensities for multiple stable secondary and tertiary structures.


title: A CRISPR screen identifies redox vulnerabilities for KEAP1/NRF2 mutant non-small cell lung cancer

abstract: The redox regulator NRF2 is hyperactivated in a large percentage of non-small cell lung cancer (NSCLC) cases, which is associated with chemotherapy and radiation resistance. To identify redox vulnerabilities for KEAP1/NRF2 mutant NSCLC, we conducted a CRISPR-Cas9-based negative selection screen for antioxidant enzyme genes whose loss sensitized cells to sub-lethal concentrations of the superoxide (O2*-)-generating drug {beta}-Lapachone. While our screen identified expected hits in the pentose phosphate pathway, the thioredoxin-dependent antioxidant system, and glutathione reductase, we also identified the mitochondrial superoxide dismutase 2 (SOD2) as one of the top hits. Surprisingly, {beta}-Lapachone did not generate mitochondrial O2*- but rather SOD2 loss enhanced the efficacy of {beta}-Lapachone due to loss of iron-sulfur protein function, loss of mitochondrial ATP maintenance and deficient NADPH production. Importantly, inhibition of mitochondrial electron transport activity sensitized to {beta}-Lapachone, demonstrating these effects may be translated to increase ROS sensitivity therapeutically.


title: The oxytocin system regulates tearing

abstract: Tears are an exocrine physiological fluid secreted onto the ocular surface from the lacrimal apparatus in all mammals. Limited research has been conducted on the functional neuronal circuitry of tear production. In particular, the neuronal mechanisms of emotional tearing, which is a physiological reaction harmonized with enhanced emotional arousal and assumed to be unique to humans, remain unclear. We identified that the oxytocin neurons in the paraventricular hypothalamus is functionally projected to the oxytocin receptor-expressing neurons in the lacrimation center of the superior salivatory nucleus. Optogenetic activation or inhibition of these neurons and/or receptors can modulate the superior salivatory nucleus dependent tear secretion mediated through oxytocin. Moreover, we identified that maternal behavior, nociceptive behavior stimulation, and aversive memory retrieval are linked to tearing in mice, and that these emotional linked tearing are suppressed by optogenetic inhibition of the corresponding oxytocin system. Thus, tearing could be regulated through functional connections between central oxytocin systems in the paraventricular hypothalamus and the superior salivatory nucleus.

category: animal behavior and cognition


title: Social closeness modulates brain dynamics during distrust anticipation

abstract: Anticipation of trust from a partner with high social closeness generates the preconditions for cooperative and reciprocal interactions to occur. However, if there is uncertainty in the interaction because the partner is a stranger or because the person has mistrusted us on another occasion, an aversive experience is generated. Hence, low social closeness or mistrust makes people keep track of the others behavior. In case of experiencing deviations from social norms, the person will monitor the intentions of the partner and update their priors regarding their social preferences. The anterior insula (AIns) seems to be sensitive to these social norm violations in functional magnetic resonance imaging (fMRI) experiments, however, the monitoring of partners with different levels of social closeness has not been investigated. In our study we wanted to find the brain regions related to (dis)trust anticipation from partners who differ in their level of social closeness. For this, we designed an experiment in which participants played an economic decision game with three people (trustors): A computer, a stranger, and a real friend. We covertly manipulated their decisions in the game so that they unexpectedly distrusted our participants. Using task-fMRI, our whole-brain analysis found that the AIns was active during the anticipation of the decisions from human partners (humans vs computer), but not during anticipation between high and low social closeness (friend vs stranger). However, using a psychophysiological interaction analysis, we found increases in functional coupling between the AIns and regions in the "mentalizing" network (such as temporal regions and parieto-occipital cortices) during trust anticipation between a high versus low social closeness partner. These results suggest that there may be a modulation of the AIns activity, specifically for high social closeness trustors, by regions that encode the intentions underlying the truster behavior.


title: Necessity and recruitment of cue-specific neuronal ensembles within the basolateral amygdala during appetitive reversal learning

abstract:
Through Pavlovian appetitive conditioning, environmental cues can become predictors of food availability. Over time, however, the food, and thus the value of the associated cues, can change based on environmental variations. This change in outcome necessitates updating of the value of the cue to appropriately alter behavioral responses to these cues. The basolateral amygdala (BLA) is critical in updating the outcomes of learned cues. However, it is unknown if the same BLA neuronal ensembles that are recruited in the initial associative memory are required when the new cue-outcome association is formed during reversal learning. The current study used the Daun02 inactivation method that enables selective targeting and disruption of activated neuronal ensembles in Fos-lacZ transgenic rats. Rats were implanted with bilateral cannulas that target the BLA and underwent appetitive discriminative conditioning in which rats had to discriminate between two auditory stimuli. One stimulus (CS+) co-terminated with food delivery, and the other stimulus was unrewarded (CS-; counterbalanced). Rats were then tested for CS+ or CS- memory retrieval and infused with either Daun02 or a vehicle solution into the BLA to inactivate either CS+ or CS- neuronal ensembles that were activated during that test. To assess if the same neuronal ensembles are necessary to update the value of the new association when the outcomes are changed, rats underwent reversal learning: the CS+ was no longer followed by food (reversal CS-, rCS-), and the CS- was now followed by food (reversal CS+; rCS+). The group that received Daun02 following CS+ session showed a decrease in conditioned responding and increased latency to the rCS- (previously CS+) during the first session of reversal learning, specifically during the first trial. This indicates that neuronal ensembles that are activated during the recall of the CS+ memory are the same neuronal ensembles needed for learning the new outcome of the same CS, now rCS-. Additionally, the group that received Daun02 following CS- session was slower to respond to the rCS+ (previously CS-) during reversal learning. This indicates that neuronal ensembles that are activated during the recall of the CS- memory are the same neuronal ensembles needed for learning the new outcome of the same CS. These results demonstrate that different neuronal ensembles within the BLA mediate memory recall of CS+ and CS- cues and reactivation of each cue-specific neuronal ensemble is necessary to update the value of that specific cue to respond appropriately during reversal learning. These results also indicate substantial plasticity within the BLA for behavioral flexibility as both groups eventually showed similar terminal levels of reversal learning.

HighlightsO_LIChemogenetic inactivation of BLA neuronal ensembles activated by learned CS+ or CS-
C_LIO_LIExamined if specific ensembles needed when cues values change in reversal learning
C_LIO_LICS+ ensemble ablation reduced responding to the same cue in early reversal learning
C_LIO_LICS- ensemble inactivation slowed learning of the new value of the cue
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title: Structure of the IL-1 27 quaternary receptor signaling complex

abstract: Interleukin 27 (IL-27) is a heterodimeric cytokine that functions to constrain T cell-mediated inflammation and plays an important role in immune homeostasis. Binding of IL-27 to cell surface receptors IL-27R and gp130 results in activation of receptor-associated Janus Kinases and nuclear translocation of STAT1 and STAT3 transcription factors. Despite the emerging therapeutic importance of this cytokine axis in cancer and autoimmunity, a molecular blueprint of the IL-27 receptor signaling complex, and its relation to other gp130/IL-12 family cytokines, is currently unclear. We used cryogenic-electron microscopy (cryo-EM) to determine the quaternary structure of IL-27 (p28/Ebi3) bound to receptor subunits, IL-27R and gp130. The resulting 3.47 [A] resolution structure revealed a three-site assembly mechanism nucleated by the central p28 subunit of the cytokine. The overall topology and molecular details of this binding are reminiscent of IL-6 but distinct from related heterodimeric cytokines IL-12 and IL-23. These results indicate distinct receptor assembly mechanisms used by heterodimeric cytokines with important consequences for targeted agonism and antagonism of IL-27 signaling.


title: Structure of the IL-27 quaternary receptor signaling complex


title: A spatio-temporally constrained gene regulatory network directed by PBX1/2 acquires limb patterning specificity via HAND2

abstract: During development cell fates are specified by tightly controlled gene expression programs. PBX TALE transcription factors control gene regulatory networks (GRN) that direct vertebrate tissue patterning and organ morphogenesis. How PBX1/2 proteins acquire context-specific functions, despite widespread embryonic expression of Pbx1/2, remains elusive. In mouse limb buds, mesenchymal-specific loss of PBX1/2 or of the transcriptional regulator HAND2 results in similar phenotypes, suggesting that PBX1/2- and HAND2-dependent programs converge to control limb development. To investigate this scenario, we combined tissue-specific and temporally-controlled mutagenesis with multi-omics approaches using the murine hindlimb bud as a model. We reconstructed a GRN collaboratively directed by PBX1/2 and HAND2, demonstrating that Pbx1-Hand2 genetically interact in vivo during hindlimb patterning, with PBX1 concomitantly acting as an upstream regulator of Hand2. At organismal-level resolution the GRN is active within restricted subsets of posterior-proximal hindlimb mesenchymal cells, wherein Pbx1/2 and Hand2 are co-expressed with their target genes. Genome-wide profiling of PBX1 binding across multiple tissues further revealed that HAND2 selects a subset of PBX-bound regions to impart limb patterning functionality. This research elucidates mechanisms underlying limb bud-specific functions by PBX1/2, while informing general principles by which promiscuous transcription factors cooperate with select cofactors to instruct distinct developmental programs.


title: Ontogeny of superorganisms:Social control of queen specialization in ants

abstract: The functioning of biological systems relies on the cooperation of specialized components. Understanding the processes that produce such specialization - the ontogeny of biological systems - is a major challenge in biology. Although biological systems exist at multiple phenotypic scales, most studies of their ontogeny have focused on multicellular organisms. Here we expand our understanding of the ontogeny of biological systems to superorganisms (i.e., insect societies with specialized individuals). Using the black garden ant Lasius niger as a study system, we investigate how founding queens, the earliest developmental stage of ant colonies, transition from expressing a diverse repertoire of behaviors to being strictly specialized in egg production. We demonstrate that the presence of workers is sufficient and necessary to inhibit the behavioral pluripotency, and thus initiate the specialization of queens. We show that this specialization is reversible, as queens revert to expressing behavioral pluripotency upon removal of their workers, even after having been specialized for several years. We report a similar social control of queen behavior in another species, suggesting that it may be common in ants. This is supported by further indications that the behavior and physiology of founding queens may be regulated by ancestral mechanisms. In addition to presenting a novel approach to study the ontogeny of biological systems, our study has the potential to reshape our understanding of the specialization of ant queens, and thus of division of labor in insect societies.


title: A happy accident: a novel turfgrass reference genome

abstract: Poa pratensis, commonly known as Kentucky bluegrass, is a popular cool-season grass species used as turf in lawns and recreation areas globally. Despite its substantial economic value, a reference genome had not previously been assembled due to the genomes relatively large size and biological complexity that includes apomixis, polyploidy, and interspecific hybridization. We report here a fortuitous de novo assembly and annotation of a P. pratensis genome. The draft assembly consists of 6.09 Gbp with an N50 scaffold length of 65.1 Mbp, and a total of 118 scaffolds, generated using PacBio long reads and Bionano optical map technology. We annotated 256K gene models and found 58% of the genome to be composed of transposable elements. To demonstrate the applicability of the reference genome, we evaluated population structure and estimated genetic diversity in three North American wild P. pratensis populations. Our results support previous studies that found high genetic diversity and population structure within the species. The reference genome and annotation will be an important resource for turfgrass breeding and biologists interested in this complex species.

category: plant biology


title: CD47-SIRPα controls ADCC killing of primary T cells by PMN through a combination of trogocytosis and NADPH oxidase activation.

abstract:
Immunotherapies targeting the "dont eat me" myeloid checkpoint constituted by CD47 SIRP interaction have promising clinical potential but are limited by toxicities associated with the destruction of non-tumor cells. These dose-limiting toxicities demonstrate the need to highlight the mechanisms of anti-CD47-SIRP therapy effects on non-tumor CD47-bearing cells. Given the increased incidence of lymphopenia in patients receiving anti-CD47 antibodies, and the strong ADCC effector function of Poly Morpho Nuclear Cells (PMNs), we investigated the behavior of primary PMNs cocultured with primary T cells in the presence of anti-CD47 mAbs. PMNs killed T cells in a CD47-mAb-dependent manner and at a remarkably potent PMN to T cell ratio of 1:1. The observed cytotoxicity was produced by a novel combination of both trogocytosis and a strong respiratory burst induced by classical ADCC and CD47-SIRP checkpoint blockade. The complex effect of the CD47 blocking mAb could be recapitulated by combining its individual mechanistic elements: ADCC, SIRP blockade and ROS induction. Although previous studies had concluded that disruption of SIRP signaling in PMNs was limited to trogocytosis-specific cytotoxicity, our results suggest that SIRP also tightly controls activation of NADPH oxidase, a function demonstrated during differentiation of immature PMNs but not so far in mature PMNs. Together, our results highlight the need to integrate PMNs in the development of molecules targeting the CD47-SIRP immune checkpoint and to design agents able to enhance myeloid cells function while limiting adverse effects to healthy cells able to participate in the anti-tumor immune response.

SynopsisDose-limiting toxicities demonstrate the need to investigate anti-CD47-SIRP therapy effects on non-tumor CD47-bearing cells. We demonstrate that anti-CD47 mAbs stimulate potent killing of T cells by PMN through a novel combination of trogocytosis and ROS regulated by SIRP.

Graphical Abstract

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title: Control of NK cell tolerance in MHC class I-deficiency by regulated SHP-1 localization to the activating immune synapse

abstract: Signaling via inhibitory KIR/Ly49 receptors preserves natural killer (NK) cell self-tolerance but also conveys NK cell reactivity towards MHC class-I low target cells in an education process. Here, we demonstrate that mouse NK cell education by H-2Dd regulates transcription of several genes in Ly49A+ NK cells including Ptpn6, encoding the phosphatase SHP-1. SHP-1 was highly expressed in uneducated NK cells, in which knock-out of Ptpn6 increased responsiveness. Following NKp46 triggering of uneducated NK cells, a higher synaptic abundance of phosphorylated SHP-1 was found relative to educated NK cells, concomitant with reduced phosphorylation of several signaling molecules, including PLC-g2, SLP-76, ZAP70/Syk and ERK1/2. SHP-1 overlapped extensively with F-actin and SLP-76 in the uneducated activating synapse of Ly49A+ NK cells, whereas a greater association between Ly49A and SHP-1 was observed in educated NK cells. Thus, our results indicate that in addition to transcriptional regulation, a distinct SHP-1 patterning in NK cell activating synapses can determine their tolerance.


title: ImputAccur: fast and user-friendly calculation of genotype-imputation accuracy-measures

abstract:
SummaryImputAccur is a software tool for genotype-imputation accuracy-measures. Imputation of untyped markers is a standard approach in genome-wide association studies to close the gap between directly genotyped and other known DNA variants. However high accuracy for imputed genotypes is fundamental. Several accuracy measures have been proposed, but unfortunately, they are implemented on different platforms, which is impractical. With ImputAccur the accuracy measures info, Iam-hiQ and r{superscript 2}based indices can be derived from standard output-files of imputation software. Sample/probe and marker filtering is possible. This allows e.g. accurate marker filtering ahead of data analysis. A Python code is available but also a stand-alone executable file.

Availability and ImplementationThe source code (Python version 3.9.4), a stand-alone executive file, and example data for ImputAccur are freely available at https://gitlab.gwdg.de/kolja.thormann1/imputationquality.git.

Supplementary InformationSupplementary information is available at Bioinformatics online.

Contactarosenb@gwdg.de or koljaalexander.thormann@uni-goettingen.de

category: genomics


title: Nucleotide augmentation for machine learning-guided protein engineering

abstract:
Machine learning-guided protein engineering is a rapidly advancing field. Despite major experimental and computational advances however, collecting protein genotype (sequence) and phenotype (function) data remains time and resource intensive. As a result, the quality and quantity of training data is often a limiting factor in developing machine learning models. Data augmentation techniques have been successfully applied to the fields of computer vision and natural language processing, however, there is a lack of such augmentation techniques for biological sequence data. Towards this end we develop nucleotide augmentation (NTA), which leverages natural nucleotide codon degeneracy to augment protein sequence data in a biologically meaningful way. As a proof of concept for protein engineering, we apply NTA to train machine learning models with benchmark data sets of protein genotype and phenotype, revealing performance gains on par and surpassing benchmarks models, even when only using a fraction of the training data. NTA also enables substantial improvements for classification tasks under heavy class imbalance.

Availability and implementationThe code to use NTA and to reproduce the analyses in this study is publicly available at https://github.com/minotm/NTA


title: ScrapPaper: A web scrapping method to extract journal information from PubMed and Google Scholar search result using Python.

abstract:
This paper introduces a program called ScrapPaper, a simple Python script that use web-scraping method to extract journal information from PubMed and Google Scholar search results page. Currently the motivation behind the program development is trying to solve a problem to obtain scientific literatures information especially the title and link and save as a list for further use such as in meta-analysis and comparative study of literatures. ScrapPaper advantage that it is simple to use with no prior programming experience and get the results ready within minutes (depending on the total search result). Web-scrapping is a very powerful method to extract information from the web and ScrapPaper employ several server friendly approaches accessing both PubMed and Google Scholar site.

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title: A universal pocket in FAALs ensures redirection of fatty acid pool away from CoA-based activation

abstract: Fatty acyl-AMP ligases (FAALs) channelize fatty acids towards biosynthesis of virulent lipids in mycobacteria and other pharmaceutically or ecologically important polyketides and lipopeptides in other microbes. They do so by bypassing the ubiquitous coenzyme A-dependent activation and rely on the acyl carrier protein-tethered 4-phosphopantetheine (holo-ACP). The molecular basis of how FAALs strictly reject chemically identical and abundant acceptors like coenzyme A (CoA) and accept holo-ACP unlike other members of the ANL superfamily remains elusive. We show FAALs have plugged the promiscuous canonical CoA-binding pockets and utilize highly selective alternative binding sites. These alternative pockets can distinguish adenosine 3, 5-bisphosphate-containing CoA from holo-ACP and thus FAALs can distinguish between CoA and holo-ACP. These exclusive features helped identify the omnipresence of FAAL-like proteins and their emergence in plants, fungi, and animals with unconventional domain organisations. The universal distribution of FAALs suggests they are parallelly evolved with FACLs for ensuring a CoA-independent activation and redirection of fatty acids towards lipidic metabolites.


title: Stem cell regulators control a G1 duration gradient in the plant root meristem

abstract: In meristems, where new plant organs initiate, key stem cell regulators have been identified, but their link to cell cycle progression remains unclear. Here, we show that the root meristem has a positional gradient of G1 duration that ranges from [~]2 h near the meristem boundary to more than 20 h in stem cells and early derivatives. Mutants in the PLETHORA (PLT) genes shortened G1 length and flattened its gradient. Computer modeling of an incoherent feed-forward loop (IFFL) predicted the inference of a negative regulatory pathway. We propose that PLT genes play opposing roles, maintaining meristem and stem cell activity and inhibiting G1 progression through the CDK inhibitor KRP5, a PLT target, and RBR1. This establishes a previously undescribed proximal-distal feature of the root meristem in which a G1 duration gradient is shaped by stem cell and meristem maintenance regulators.


title: Alcohol-sourced acetate reduces T cell filamentous actin branching and migration

abstract:
Alcohol is among the most widely consumed dietary substances. While excessive alcohol consumption damages the liver, heart and brain, clinical observations suggest that alcohol has strong immunoregulatory properties. Mechanisms by which alcohol undermines immune cell functions during autoimmunity and vaccination are currently elusive. Herein, we show that acetate, the main metabolite of alcohol, effectively inhibits the migratory capacity of T cells through acetylation of cortactin, a protein which stabilizes and promotes branching of filamentous actin. Acetate-induced cytoskeletal changes effectively inhibited activation, proliferation, and immune synapse formation in T cells at tissue concentrations reached by alcohol consumption. In summary, these data show that acetate inhibits T cell-mediated immune responses by modulating the T cell cytoskeletal biomechanics.

One-Sentence SummaryExposure to elevated acetate concentrations change T cell biomechanics, reducing cell migration and immune synapse formation.


title: Crops grown in mixtures show niche partitioning in spatial water uptake

abstract:
O_LIMore diverse plant communities generally produce more biomass than monocultures. This benefit of plant diversity is supposed to stem from resource complementarity of species in mixtures. Different plant species might use the resources spatially, temporally, or chemically in different ways. Along the same lines, for agricultural production crop mixtures outperform monocultures. Complementarity in water use of crop species in mixtures could result in higher yield. Therefore, complementarity in water use could be relevant for crop production in the future when water for agriculture becomes increasingly limited.
C_LIO_LIHere we used the stable isotopes of water and a Bayesian model to investigate the spatial water uptake patterns of six different crops species and how these patterns change upon increasing crop diversity. In addition, we calculated niche overlaps of water uptake and compared them among the different diversity levels.
C_LIO_LIThe spatial water uptake pattern differed among crop species. The effect of crop diversity had a minor effect on water uptake but varied strongly depending on the crop species. Niche overlap in water uptake was highest in monocultures and decreased strongly in mixtures. Additionally, we show that increased competition intensity caused stronger changes in water uptake patterns.
C_LIO_LISynthesis. We found evidence for niche partitioning of water uptake in crop mixtures which might lead to a more efficient use of the limited resource water in intercropping systems. Intercropping can therefore increase and stabilise crop yields under forthcoming drier climatic conditions.
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