title: Single seed microbiota: assembly and transmission from parent plant to seedling

abstract: Gaining basic understanding of processes involved in seed microbiota assembly is a prerequisite for improving crop establishment. Investigation of microbiota structure during seed development revealed that individual seeds of bean and radish were associated with a dominant bacterial taxon representing more than 75% of all reads. The identity of this taxon was highly variable between plants and within seeds of the same plant. Succession of dominant taxa occurred during seed filling and maturation through Selection. In a second step, we evaluated seed to seedling transmission of these dominant seed-borne taxa. We showed that initial bacterial abundance on seeds was not a good predictor of seedling transmission and that the identity of seed-borne taxa can impact seedling phenotype. Altogether this work unveiled that seeds are colonized by few bacterial taxa of highly variable identity, which appears to be important for the early stages of plant development.


title: Time-resolved cryo-EM visualizes ribosomal translocation with EF-G and GTP

abstract: During translation, a conserved GTPase elongation factor--EF-G in bacteria or eEF2 in eukaryotes--translocates tRNA and mRNA through the ribosome. EF-G has been proposed to act as a flexible motor that propels tRNA and mRNA movement, as a rigid pawl that biases unidirectional translocation resulting from ribosome rearrangements, or by various combinations of motor- and pawl-like mechanisms. Using time-resolved cryo-EM, we visualized GTP-catalyzed translocation without inhibitors, capturing elusive structures of ribosome*EF-G intermediates at near-atomic resolution. Prior to translocation, EF-G binds near peptidyl-tRNA, while the rotated 30S subunit stabilizes the EF-G GTPase center. Reverse 30S rotation releases Pi and translocates peptidyl-tRNA and EF-G by [~]20 [A]. An additional 4-[A] translocation initiates EF-G dissociation from a transient ribosome state with highly swiveled 30S head. The structures visualize how nearly rigid EF-G rectifies inherent and spontaneous ribosomal dynamics into tRNA-mRNA translocation, whereas GTP hydrolysis and Pi release drive EF-G dissociation.

category: molecular biology


title: GFI1 tethers the NuRD complex to open and transcriptionally active chromatin in myeloid progenitors

abstract: GFI1 is a SNAG-domain, DNA binding transcriptional repressor which controls myeloid differentiation, in particular the formation of neutrophils. Here we show that GFI1 interacts with the chromodomain helicase CHD4 and other components of the "Nucleosome remodeling and deacetylase" (NuRD) complex. In granulo-monocytic precursors, GFI1, CHD4 or GFI1/CHD4 complexes occupy sites of open chromatin enriched for histone marks associated with active transcription suggesting that GFI1 recruits the NuRD complex to target genes that are regulated by active or bivalent promoters and active enhancers. Our data also show that GFI1 and GFI1/CHD4 complexes occupy promoters of different sets of genes that are either enriched for IRF1 or SPI-1 consensus sites, respectively. During neutrophil differentiation, overall chromatin closure and depletion of H3K4me2 occurs at different degrees depending on whether GFI1, CHD4 or both are present, indicating that GFI1 affects the chromatin remodeling activity of the NuRD complex. Moreover, GFI1/CHD4 complexes regulate chromatin openness and histone modifications differentially to enable regulation of target genes affecting the signaling pathways of the immune response or nucleosome organization or cellular metabolic processes.


title: End-of-life targeted auxin-mediated degradation of DAF-2 Insulin/IGF-1 receptor promotes longevity free from growth-related pathologies

abstract: Preferably, lifespan-extending therapies should work when applied late in life without causing undesired pathologies. However, identifying lifespan-extending interventions that are effective late in life and which avoid undesired secondary pathologies remains elusive. Reducing Insulin/IGF-1 signaling (IIS) increases lifespan across species, but the effects of reduced IIS interventions in extreme geriatric ages remains unknown. Using the nematode C. elegans, we engineered the conditional depletion of the DAF-2/insulin/IGF-1 transmembrane receptor using an auxin-inducible degradation (AID) system that allows for the temporal and spatial reduction in DAF-2 protein levels at time points after which interventions such as RNAi may lose efficacy. Using this system, we found that AID-mediated depletion of DAF-2 protein efficiently extends animal lifespan. Depletion of DAF-2 during early adulthood resulted in multiple adverse phenotypes, including growth retardation, germline shrinkage, egg-retention, and reducing offspring. By contrast, however, AID-mediated depletion of DAF-2 specifically in the intestine resulted in an extension of lifespan without these deleterious effects. Importantly, AID-mediated depletion of DAF-2 protein in animals past their median lifespan allowed for an extension of lifespan without affecting growth or behavioral capacity. Thus, both late-in-life targeting and tissue-specific targeting of IIS minimize the deleterious effects typically seen with interventions that reduced IIS, suggesting potential therapeutic methods by which longevity and healthspan can be increased in even geriatric populations.


title: Role of the G-Protein Coupled Receptor 3-Salt Inducible Kinase 2 Pathway in Human β CellProliferation

abstract: Loss of pancreatic {beta} cells is the hallmark of type 1 diabetes (T1D) 1, for which provision of insulin is the standard of care. While regenerative and stem cell therapies hold the promise of generating single-source or host-matched tissue to obviate immune-mediated complications2-4, these will still require surgical intervention and immunosuppression. Thus, methods that harness the innate capacity of {beta}-cells to proliferate to increase {beta} cell mass in vivo are considered vital for future T1D treatment5, 6. However, early in life {beta} cells enter what appears to be a permanent state of quiescence 7-10, directed by an evolutionarily selected genetic program that establishes a {beta} cell mass setpoint to guard against development of fatal endocrine tumours. Here we report the development of a high-throughput RNAi screening approach to identify upstream pathways that regulate adult human {beta} cell quiescence and demonstrate in a screen of the GPCRome that silencing G-protein coupled receptor 3 (GPR3) leads to human pancreatic {beta} cell proliferation. Loss of GPR3 leads to activation of Salt Inducible Kinase 2 (SIK2), which is necessary and sufficient to drive cell cycle entry, increase {beta} cell mass, and enhance insulin secretion in mice. Taken together, targeting the GPR3-SIK2 pathway represents a novel avenue to stimulate the regeneration of {beta} cells.

category: cell biology


title: Balanced mitochondrial and cytosolic translatomes underlie the biogenesis of human respiratory complexes

abstract: Oxidative phosphorylation (OXPHOS) complexes consist of nuclear and mitochondrial DNA-encoded subunits. Their biogenesis requires cross-compartment gene regulation to mitigate the accumulation of disproportionate subunits. To determine how human cells coordinate mitochondrial and nuclear gene expression processes, we established an optimized ribosome profiling approach tailored for the unique features of the human mitoribosome. Analysis of ribosome footprints in five cell types revealed that average mitochondrial synthesis rates corresponded precisely to cytosolic rates across OXPHOS complexes. Balanced mitochondrial and cytosolic synthesis did not rely on rapid feedback between the two translation systems. Rather, LRPPRC, a gene associated with Leighs syndrome, is required for the reciprocal translatomes and maintains cellular proteostasis. Based on our findings, we propose that human mitonuclear balance is enabled by matching OXPHOS subunit synthesis rates across cellular compartments, which may represent a vulnerability for cellular proteostasis.


title: Craniofacial cartilage organoids from human embryonic stem cells via a neural crest cell intermediate

abstract: Severe birth defects or major injuries to the face require surgical reconstruction and rehabilitation. The ability to make bona fide craniofacial cartilage - cartilage of the head and face - from patient-derived induced pluripotent stem cells (iPSCs) to repair these birth defects and injuries has tremendous translational applications, but is not yet possible. The neural crest is the normal developmental pathway for craniofacial cartilage, however, the knowledge of cell signaling pathways that drive neural crest differentiation into craniofacial chondrocytes is limited. Here we describe a differentiation protocol that generated self-organizing craniofacial cartilage organoids from human embryonic stem cells (hESCs) and IPSCs through a neural crest stem cell (NCSC) intermediate. Histological staining of cartilage organoids revealed tissue architecture typical of hyaline cartilage. Organoids were composed of rounded aggregates of glassy, gray matrix that contained scattered small nuclei in lacunae. Mass spectrometry shows that the organoids express robust levels of cartilage markers including aggrecan, perlecan, proteoglycans, and many collagens. Organoids expressed markers indicative of neural crest lineage, as well as growth factors that are candidates for chondrocyte differentiation factors. The data suggest that chondrocyte differentiation is initiated by autocrine loops driven by a combination of secreted growth factors that bind to chondrocyte receptors. Craniofacial cartilage organoids were continuously cultured for one year, reaching up to one centimeter in diameter. The ability to grow craniofacial cartilage from NCSCs provides insights into the cell signaling mechanisms of differentiation into craniofacial cartilage, which lays the groundwork for understanding mechanistic origins of congenital craniofacial anomalies and repairing cartilaginous structures of the head and face.

category: developmental biology


title: Integrating ecological monitoring and local ecological knowledge to evaluate conservation outcomes

abstract: Successful conservation of long-lived species requires reliable understanding of long-term trends and historical baselines. Using a green turtle (Chelonia mydas) foraging aggregation in the northern Gulf of California, Mexico as case study, we integrated scientific monitoring data with historic catch rate reconstructions derived from Local Ecological Knowledge (LEK). Models fit to LEK and monitoring data indicate that turtle abundance is increasing, but only after ~40 years of safeguarding the species nesting and foraging habitats in Mexico. However, as population declines occurred 75% faster than increases, and current abundance is at ~60% of historical baseline levels, indicating the need for sustained, long-term conservation actions. This study demonstrates the potential of linking LEK and ecological science to provide critical information for conservation, by establishing reference baselines and gauging population status, while promoting equitable and sustainable futures for local communities.

category: ecology


title: The genetic impact of an Ebola outbreak on a wild gorilla population

abstract:
BackgroundNumerous Ebola virus outbreaks have occurred in Equatorial Africa over the past decades. Besides human fatalities, gorillas and chimpanzees have also succumbed to the fatal virus. The 2004 outbreak at the Odzala-Kokoua National Park (Republic of Congo) alone caused a severe decline in the resident western lowland gorilla (Gorilla gorilla gorilla) population, with a 95% mortality rate. Here, we explore the immediate genetic impact of the Ebola outbreak in the western lowland gorilla population.

ResultsAssociations with survivorship were evaluated by utilizing DNA obtained from fecal samples from 16 gorilla individuals declared missing after the outbreak (non-survivors) and 15 individuals observed before and after the epidemic (survivors). We used a target enrichment approach to capture the sequences of 123 genes previously associated with immunology and Ebola virus resistance and additionally analyzed the gut microbiome which could influence the survival after an infection. Our results indicate no changes in the population genetic diversity before and after the Ebola outbreak, and no significant differences in microbial community composition between survivors and non-survivors. However, and despite the low power for an association analysis, we do detect six nominally significant missense mutations in four genes that might be candidate variants associated with an increased chance of survival.

ConclusionThis study offers the first insight to the genetics of a wild great ape population before and after an Ebola outbreak using target capture experiments from fecal samples, and presents a list of candidate loci that may have facilitated their survival.

category: genomics


title: Distinct gene regulatory signatures of dominance rank and social bond strength in wild baboons

abstract: The social environment is a major determinant of morbidity, mortality, and Darwinian fitness in social animals. Recent studies have begun to uncover the molecular processes associated with these relationships, but the degree to which they vary across different dimensions of the social environment remains unclear. Here, we draw on a long-term field study of wild baboons to compare the signatures of affiliative and competitive aspects of the social environment in white blood cell gene regulation, under both immune stimulated and non-stimulated conditions. We find that the effects of dominance rank on gene expression are directionally opposite in males versus females, such that high-ranking males resemble low-ranking females, and vice-versa. Among females, rank and social bond strength are both reflected in the activity of cellular metabolism and proliferation genes. However, pronounced rank-related differences in baseline immune gene activity are near-absent for social bond strength, while only bond strength predicts the fold-change response to immune (lipopolysaccharide) stimulation. Together, our results indicate that the directionality and magnitude of social effects on gene regulation depend on the aspect of the social environment under study. This heterogeneity may help explain why social environmental effects on health and longevity can also vary between measures.


title: Digital embryos - A novel technical approach to investigate perceptual categorization in pigeons (Columba livia) using machine learning

abstract: Pigeons are classic model animals to study perceptual category learning. A theoretical understanding of the cognitive mechanisms of categorization requires a careful consideration of the employed stimulus material. Optimally, stimuli should not consist of real-world objects that might be associated with prior experience. The number of exemplars should be theoretically infinite and easy to produce. In addition, the experimenter should have the freedom to produce 2D- and 3D-versions of the stimuli and, finally, the stimulus set should provide the opportunity to identify the diagnostic elements that the animals use. To this end, we used the approach of "virtual phylogenesis" of "digital embryos" to produce two stimulus sets of objects that meet these criteria. In our experiment pigeons learned to categorize these stimuli in a forced-choice procedure. In addition, we used peck tracking to identify where on the stimulus the animals pecked to signal their choice. Pigeons learned the task and transferred successfully to novel exemplars. Using a k-nearest neighbor classifier, we were able to predict the presented stimulus class based on pecking location indicating that pecks are related to features of interest. We further identified potential strategies of the pigeons through this approach, namely that they were either learning one or two categories to discriminate between stimulus classes. These strategies remained stable during category transfer, but differed between individuals indicating that categorization learning is not limited to a single learning strategy.

category: animal behavior and cognition


title: A molecular toolbox for ADP-ribosyl binding proteins

abstract: Proteins interacting with ADP-ribosyl groups are often involved in disease-related pathways or in viral infections, which makes them attractive targets for the development of inhibitors. Our goal was to develop a robust and accessible assay technology that is suitable for high-throughput screening and applicable to a wide range of proteins acting as either hydrolysing or non-hydrolysing binders of mono- and poly-ADP-ribosyl groups. As a foundation of our work, we developed a C-terminal protein fusion tag based on a Gi protein alpha subunit peptide (GAP), which allows for site-specific introduction of cysteine-linked mono- and poly-ADP-ribosyl groups as well as chemical ADP-ribosyl analogs. By fusion of the GAP-tag and ADP-ribosyl binders to fluorescent proteins, we were able to generate robust FRET signals and the interaction with 22 previously described ADP-ribosyl-binders was confirmed. To demonstrate the applicability of this binding assay for high-throughput screening, we utilized it to screen for inhibitors of the SARS-CoV-2 nsp3 macrodomain and identified the drug suramin as a moderate yet unspecific inhibitor of this protein. To complement the binding technology, we prepared high-affinity ADP-ribosyl binders fused to a nanoluciferase, which enabled simple blot-based detection of mono- and poly-ADP-ribosylated proteins. These tools can be expressed recombinantly in E. coli using commonly available agents and will help to investigate ADP-ribosylation systems and aid in drug discovery.

category: biochemistry


title: EnTrance: Exploration of Entropy Scaling Ball Cover Search in Protein Sequences

abstract: Biological sequence alignment using computational power has received increasing attention as technology develops. It is important to predict if a novel DNA sequence is potentially dangerous by determining its taxonomic identity and functional characteristics through sequence identification. This task can be facilitated by the rapidly increasing amounts of biological data in DNA and protein databases thanks to the corresponding increase in computational and storage costs. Unfortunately, the growth in biological databases has caused difficulty in exploiting this information. EnTrance presents an approach that can expedite the analysis of this large database by employing entropy scaling. This allows scaling with the amount of entropy in the database instead of scaling with the absolute size of the database. Since DNA and protein sequences are biologically meaningful, the space of biological sequences demonstrates the structure exploited by entropy scaling. As biological sequence databases grow, taking advantage of this structure can be extremely beneficial for reducing query times. EnTrance, the entropy scaling search algorithm introduced here, accelerates the biological sequence search exemplified by tools such as BLAST. EnTrance does this by utilizing a two step search approach. In this fashion, EnTrance quickly reduces the number of potential matches before more exhaustively searching the remaining sequences. Tests of EnTrance show that this approach can lead to improved query times. However, constructing the required entropy scaling indices beforehand can be challenging. To improve performance, EnTrance investigates several ideas for accelerating index build time that supports entropy scaling searches. In particular, EnTrance makes full use of the concurrency features of Go language greatly reducing the index build time. Our results identify key tradeoffs and demonstrate that there is potential in using these techniques for sequence similarity searches. Finally, EnTrance returns more matches and higher percentage identity matches when compared with existing tools.

category: bioinformatics


title: Foregut organ progenitors and their niche display distinct viscoelastic properties in vivo during early morphogenesis stages

abstract: Material properties of living matter play an important role for biological function and development. Yet, quantification of material properties of internal organs in vivo, without causing physiological damage, remains challenging. Here, we present a non-invasive approach based on modified optical tweezers for quantifying sub-cellular material properties deep inside living zebrafish. Material properties of cells within the gut region of living zebrafish are quantified as deep as 150 m into the biological tissue. The measurements demonstrate differential mechanical properties of the developing foregut organs progenitors: Gut progenitors are more elastic than any of the neighboring cell populations at the time when the developing organs undergo substantial displacements during morphogenesis. The higher elasticity of gut progenitors correlates with an increased cellular concentration of microtubules. The results infer a role of material properties during morphogenesis and the approach paves the way for quantitative material investigations in vivo of embryos, explants, or organoids.

category: biophysics


title: Calsyntenin-3, an atypical cadherin, suppresses inhibitory basket- and stellate-cell synapses but boosts excitatory parallel-fiber synapses in cerebellum

abstract: Cadherins contribute to the organization of nearly all tissues, but the functions of several evolutionarily conserved cadherins, including those of calsyntenins, remain enigmatic. Puzzlingly, two distinct, non-overlapping functions for calsyntenins were proposed: As postsynaptic neurexin ligands in synapse formation, or as presynaptic adaptors for kinesin-mediated vesicular transport. Here, we show that acute CRISPR-mediated deletion of calsyntenin-3 in cerebellar Purkinje cells in vivo causes a large decrease in inhibitory synapses, but a surprisingly robust increase in excitatory parallel-fiber synapses. No changes in the dendritic architecture of Purkinje cells or in climbing-fiber synapses were detected. Thus, by promoting formation of an excitatory type of synapses and decreasing formation of an inhibitory type of synapses in the same neuron, calsyntenin-3 functions as a postsynaptic adhesion molecule that regulates the excitatory/inhibitory balance in Purkinje cells. No similarly opposing function of a synaptic adhesion molecule was previously observed, suggesting a new paradigm of synaptic regulation.

category: neuroscience


title: One shot generalization in humans revealed through a drawing task

abstract: Humans have the striking ability to learn and generalize new visual concepts from just a single exemplar. We suggest that when presented with a novel object, observers identify its significant features and infer a generative model of its shape, allowing them to mentally synthesize plausible variants. To test this, we showed participants abstract 2D shapes ( Exemplars) and asked them to draw new objects ( Variations) belonging to the same class. We show that this procedure created genuine novel categories. In line with our hypothesis, particular features of each Exemplar were preserved in its Variations and there was striking agreement between participants about which shape features were most distinctive. Also, we show that strategies to create Variations were strongly driven by part structure: new objects typically modified individual parts (e.g., limbs) of the Exemplar, often preserving part order, sometimes altering it. Together, our findings suggest that sophisticated internal generative models are key to how humans analyze and generalize from single exemplars.


title: High-fat diet induced loss of GABAergic inhibition decouples intrinsic and synaptic excitability in AgRP neurons

abstract: Obesity is a progressive, relapsing disease with few therapies. Diet and lifestyle interventions are effective but are often temporary and many individuals regain weight. High-fat diet increases the excitability of AgRP neurons, a critical neuronal population for the regulation of food intake and body weight. Here we investigate the plasticity of AgRP neurons and the impact of high-fat diet on modulation by synaptic input. We find that diet-induced hyperexcitability of AgRP neurons is not reversed by a lower-fat diet intervention. High-fat diet is associated with changes in the synaptic modulation of AgRP neurons, with a paradoxical increase in inhibitory input accompanied by a loss of GABA-mediated inhibition due to a depolarizing shift in the reversal potential of the GABA-evoked Cl- current. These findings reveal that high-fat diet leads to decoupling of intrinsic and synaptic excitability in AgRP neurons, such that hyperexcitability of AgRP neurons persists despite an increase in inhibitory input, revealing a mechanism for the difficulty in sustaining weight loss.


title: Center-surround interactions underlie bipolar cell motion sensing in the mouse retina

abstract: Motion is a critical aspect of vision. We studied the representation of motion in mouse retinal bipolar cells and found, surprisingly, that some bipolar cells possess motion-sensing capabilities that rely on their center-surround receptive fields. Using a glutamate sensor, we directly observed motion-sensitive bipolar cell synaptic output, which was strongest for local motion and dependent on the motions origin. We characterized bipolar cell receptive fields and found that there are motion and non-motion sensitive bipolar cell types, the majority being motion sensitive. Next, we used these bipolar cell receptive fields along with connectomics to design biophysical models of downstream cells. The models and experiments demonstrated that bipolar cells pass motion-sensitive excitation to starburst amacrine cells through direction-specific signals mediated by bipolar cells center-surround receptive field structure. As bipolar cells provide excitation to most amacrine and ganglion cells, their motion sensitivity may contribute to motion processing throughout the visual system.


title: Neuromodulators enable overlapping synaptic memory regimes and nonlinear transition dynamics in recurrent neural networks

abstract: Mood, arousal, and other internal states can drastically alter behavior, even in identical external circumstances -- a cold glass of water when you are thirsty is much more desirable than when you are sated. Neuromodulators are critical controllers of such neural states, with dysfunctions linked to various neuropsychiatric disorders. Although biological aspects of neuromodulation have been well studied, the computational principles underlying how large-scale neuromodulation of distributed neural populations shifts brain states remain unclear. We use recurrent neural networks to model how synaptic weight modulation -- an important function of neuromodulators -- can achieve nuanced alterations in neural computation, even in a highly simplified form. We find that under structural constraints like those in brains, this provides a fundamental mechanism that can increase the computational capability and flexibility of a neural network by enabling overlapping storage of synaptic memories able to generate diverse, even diametrically opposed, behaviors. Our findings help explain how neuromodulators "unlock" specific behaviors by creating task-specific hypertubes in the space of neural activities and motivate more flexible, compact and capable machine learning architectures.


title: Neuromodulators generate multiple context-relevant behaviors in a recurrent neural network by shifting activity hypertubes


title: Loss of Stk11 in basolateral amygdalar projection neurons impairs CTA learning by altering the temporal pattern of taste response plasticity in GC

abstract: Gustatory cortex (GC) responds to tastes on the tongue with dynamic ensemble activity that represents first the presence, then the identity, and finally the hedonic value (palatability) of tastes. This final state of the taste response is uniquely altered by conditioned taste aversion (CTA) - a powerful one-trial learning paradigm in which a taste becomes aversive after association with gastric malaise - a process requiring coordination between GC and basolateral amygdala (BLA). Previously, we found that one key requirement for learning in this circuit is expression of the serine/threonine kinase 11 (Stk11) gene (a tumor suppression gene which has only recently been associated with learning; Levitan et al., 2020). When Stk11 is knocked out in BLA projection neurons (BLApn), CTA learning fails to occur. Here we have examined how learning-related response plasticity in GC taste responses is impacted by the knockout of Stk11 in BLApn. Contrary to the commonly held assumption that a lack of learning means a lack of such plasticity, but consistent with the fact that Stk11KO has been shown to increase the excitability of BLApn, our data reveal that the knockout of Stk11 in BLApn does not eliminate plasticity; rather, it shifts the impact of CTA training on GC taste responses to an earlier, learning-inappropriate epoch. Even naive taste representations are altered--specifically, the pattern of similarities and differences among the different taste responses are rendered abnormal by Stk11 KO, and these relationships fail to change with training. Finally, the latency of behavior-related dynamic ensemble features of the GC taste response, which is also abnormal in naive KO mice, is rendered disorganized by CTA. Together, these results suggest that Stk11 plays a role in governing the coordination of GC activity by BLA, and demonstrate that alterations in the function of BLApn caused by Stk11KO inhibit learning not by inhibiting plasticity but by changing its temporal properties.


title: Loss of Stk11 in basolateral amygdalar projection neurons impairs taste aversion learning by altering the temporal pattern of taste response plasticity in gustatory cortex


title: Mediobasal hypothalamic FKBP51 acts as a molecular switch linking autophagy to whole-body metabolism

abstract: The mediobasal hypothalamus (MBH) is the central region in the physiological response to metabolic stress. The FK506-binding protein 51 (FKBP51) is a major modulator of the stress response and has recently emerged as a scaffolder regulating metabolic and autophagy pathways. However, the detailed protein-protein interactions linking FKBP51 to autophagy upon metabolic challenges remain elusive. We performed mass spectrometry-based metabolomics of FKBP51 knockout (KO) cells revealing an increased amino acid and polyamine metabolism. We identified FKBP51 as a central nexus for the recruitment of the LKB1/AMPK complex to WIPI4 and TSC2 to WIPI3, thereby regulating the balance between autophagy and mTOR signaling in response to metabolic challenges. Furthermore, we demonstrated that MBH FKBP51 deletion strongly induces obesity, while its overexpression protects against high-fat diet (HFD) induced obesity. Our study provides an important novel regulatory function of MBH FKBP51 within the stress-adapted autophagy response to metabolic challenges.


title: U-RISC: An Ultra-high Resolution EM Dataset Challenging Existing Deep Learning Algorithms

abstract: Connectomics is a developing field aiming at reconstructing the connection of the neural system at nanometer scale. Computer vision technology, especially deep learning methods used in image processing, has promoted connectomic data analysis to a new era. However, the performance of the state-of-the-art methods still falls behind the demand of scientific research. Inspired by the success of ImageNet, we present the U-RISC, an annotated Ultra-high Resolution Image Segmentation dataset for Cell membrane, which is the largest cell membrane annotated Electron Microscopy (EM) dataset with a resolution of 2.18nm/pixel. Multiple iterative annotations ensured the quality of the dataset. Through an open competition, we reveal that the performance of current deep learning methods still has a considerable gap with human-level, different from ISBI 2012, on which the performance of deep learning is close to human. To explore the causes of this discrepancy, we analyze the neural networks with a visualization method, attribution analysis. We find that in U-RISC, it requires a larger area around a pixel to predict whether the pixel belongs to the cell membrane or not. Finally, we integrate currently available methods to provide a new benchmark (0.67, 10% higher than the leader of competition, 0.61) for cell membrane segmentation on U-RISC and propose some suggestions in developing deep learning algorithms. The U-RISC dataset and the deep learning codes used in this paper will be publicly available.


title: Drosophila architectural protein CTCF is not essential for fly survival and is able to function independently of CP190

abstract:
CTCF is the most likely ancestor of proteins that contain large clusters of C2H2 zinc finger domains (C2H2) and is conserved among most bilateral organisms. In mammals, CTCF functions as the main architectural protein involved in the organization of topology-associated domains (TADs). In vertebrates and Drosophila, CTCF is involved in the regulation of homeotic genes. Previously, null mutations in the dCTCF gene were found to result in death during the stage of pharate adults, which failed to eclose from their pupal case. Here, we obtained several new null dCTCF mutations and found that the complete inactivation of dCTCF appears to be limited to phenotypic manifestations of the Abd-B gene and fertility of adult flies. Many modifiers that are not associated with an independent phenotypic manifestation can significantly enhance the expressivity of the null dCTCF mutations, indicating that other architectural proteins are able to functionally compensate for dCTCF inactivation in Drosophila. We also mapped the 715-735 aa region of dCTCF as being essential for the interaction with the BTB (Broad-Complex, Tramtrack, and Bric a brac) and microtubule-targeting (M) domains of the CP190 protein, which binds to many architectural proteins. However, the mutational analysis showed that the interaction with CP190 was not essential for the functional activity of dCTCF in vivo.

HighlightsO_LIThe dCTCF 715-735 aa region interacts with the BTB and M domains of CP190
C_LIO_LIInteraction with CP190 is not essential for dCTCF function in vivo
C_LIO_LINull dCTCF mutants are viable but show severely reduced fertility
C_LI


title: Systematic identification of a CDYL1-dependent decrease in lysine crotonylation at DNA double-strand break sites functionally uncouples transcriptional silencing and repair

abstract:
Previously, we showed that CDYL1 is recruited to DNA double-strand breaks (DSBs) to promote homology-directed repair (HDR) and foster transcriptional silencing. Yet, how CDYL1 elicits DSB-induced silencing is not fully understood. Here, we systematically identify a CDYL1-dependent local decrease in the transcriptionally active marks lysine crotonylation (PanKcr) and crotonylated histone residue H3K9cr at AsiSI-induced DSBs, which correlates with transcriptional silencing. Mechanistically, we reveal that CDYL1 crotonyl-CoA hydratase activity counteracts PanKcr and H3K9cr at AsiSI sites, which triggers the eviction of the transcriptional elongation factor ENL and foster transcriptional silencing. Furthermore, genetic inhibition of CDYL1 hydratase activity blocks the reduction in H3K9cr and alleviates DSB-induced silencing, while HDR efficiency unexpectedly remains intact. Therefore, our results functionally uncouple the repair and silencing activity of CDYL1 at DSBs. In a broader context, we address a long-standing question concerning the functional relationship between HDR and DSB-induced transcriptional silencing, suggesting that they may occur independently.

HighlightsO_LISystematic identification of a local decrease in lysine crotonylation PanKcr and H3K9cr at AsiSI-induced DSBs that correlates with transcriptional silencing.
C_LIO_LICDYL1 crotonyl-CoA hydratase activity downregulates Kcr at DSBs.
C_LIO_LIKcr reduction at DSBs promotes ENL eviction and fosters transcriptional silencing.
C_LIO_LICDYL1 roles in DSB-induced transcriptional silencing and HDR are functionally uncoupled.
C_LI


title: Molecular foundations of Precambrian uniformitarianism

abstract: The earliest geochemical indicators of microbes--and the enzymes that powered them--extend back almost 3.8 billion years on our planet. Paleobiologists often attempt to understand these indicators by assuming that the behaviors of modern microbes and enzymes are consistent (uniform) with those of their predecessors. A uniformitarian assumption (i.e., the idea that fundamental geobiological processes have occurred in much the same manner over Earth history) seems at odds with our understanding of the inherent variability of living systems. Here, we examine whether a uniformitarian assumption for an enzyme thought to generate carbon isotope indicators of biological activity, RuBisCO, can be corroborated by independently studying the history of changes recorded within RuBisCOs genetic sequences. Specifically, we resurrected a Precambrian-age, ancient RuBisCO by engineering its ancient DNA inside a modern cyanobacterium genome and measured the engineered organisms fitness and carbon-isotope-discrimination profile. The envelope of ancestral RuBisCO isotopic fractionation observed here indicates that uniformitarian assumptions may be warranted, but with important caveats. Our results suggest that further inquiries that link molecule-level evolutionary changes with planet-level geochemical conditions are needed to discern whether enzyme-affected isotope fractionation trends extend deeper into the early Precambrian. Experimental studies illuminating lifes early molecular innovations are crucial to explore the foundations of Precambrian uniformitarian assumptions.

category: evolutionary biology


title: Resurrected Rubisco suggests uniform carbon isotope signatures over geologic time


title: Hematologic and systemic metabolic alterations due to Mediterranean type II G6PD deficiency in a novel murine model

abstract: Deficiency of Glucose 6 phosphate dehydrogenase (G6PD) is the single most common enzymopathy, present in approximately 400 million humans (e.g. 5% of humans). Its prevalence is hypothesized to be due to conferring resistance to malaria. However, G6PD deficiency also results in hemolytic sequelae from oxidant stress. Moreover, G6PD deficiency is associated with kidney disease, diabetes, pulmonary hypertension, immunological defects, and neurodegenerative diseases. To date, the only available mouse models have decreased levels of G6PD due to promoter mutations, but with stable G6PD. However, human G6PD mutations are missense mutations that result in decreased enzymatic stability. As such, this results in very low activity in red blood cells and platelets that cannot synthesize new protein. To generate a more accurate model, the human sequence for a severe form of G6PD deficiency (Med -) was knocked into the murine G6PD locus. As predicted, G6PD levels were extremely low in RBCs and deficient mice have increased hemolytic sequalae to oxidant stress. G6PD levels were mildly decreased in non-erythroid organs, consistent with what has been observed in humans. Juxtaposition of G6PD deficient and wild-type mice revealed altered lipid metabolism in multiple organ systems. Together, these findings both establish a new mouse model of G6PD deficiency that more accurately reflects human G6PD deficiency and also advance our basic understanding of altered metabolism in this setting.


title: A Family of Glycosylated Macrolides Selectively Target Energetic Vulnerabilities in Leukemia

abstract: Cancer cells have long been recognized to exhibit unique bioenergetic requirements. The apoptolidin family of glycomacrolides are distinguished by their selective cytotoxicity towards oncogene transformed cells, yet their molecular mechanism remains uncertain. We used photoaffinity analogs of the apoptolidins to identify the F1 subcomplex of mitochondrial ATP synthase as the target of apoptolidin A. CryoEM of apoptolidin and ammocidin-ATP synthase complexes revealed a novel shared mode of inhibition that was confirmed by deep mutational scanning of the binding interface to reveal resistance mutations which were confirmed using CRISPR-Cas9. Ammocidin A was found to suppress leukemia progression in vivo at doses that were tolerated with minimal toxicity. The combination of cellular, structural, mutagenesis, and in vivo evidence define the mechanism of action of apoptolidin family glycomacrolides and establish a path to address OXPHOS-dependent cancers.


title: SLP2/prohibitins aggregates and instability of the PHB complex are key elements in CHCHD10S59L-related disease

abstract: CHCHD10 is an ALS/FTD gene, also involved in a large clinical spectrum, that encodes a protein whose precise function within mitochondria is unclear. Here we show that CHCHD10 interacts with the Stomatin-Like Protein 2 (SLP2) to control the stability of the Prohibitin (PHB) complex in the inner mitochondrial membrane. In affected tissues, SLP2 forms aggregates with prohibitins and the instability of the PHB complex results in activation of OMA1 and accelerated OPA1 proteolysis leading to mitochondrial fragmentation, loss of mitochondrial cristae and apoptosis. Abnormal cristae morphogenesis depends on both the PHB complex destabilization leading to MICOS complex instability, via disruption of OPA1/Mitofilin interaction, and the activation of PINK1-mediated pathways. We also show that the increase of mitophagy found in both heart and hippocampus of Chchd10S59L/+ mito-QC mice is PINK1/Parkin-dependent. Thus, SLP2/PHBs aggregates and destabilization of the PHB complex with PINK1 activation are critical in the sequence of events leading to CHCHD10S59L-related disease.

category: pathology


title: Lessons from 20 years of plant genome sequencing: an unprecedented resource in need of more diverse representation

abstract: The field of plant genomics has grown rapidly in the past 20 years, leading to dramatic increases in both the quantity and quality of publicly available genomic resources. With this ever-expanding wealth of genomic data from an increasingly diverse set of taxa, unprecedented potential exists to better understand the genome biology and evolution of plants. Here, we provide a contemporary view of plant genomics, including analyses on the quality of existing plant genome assemblies, the taxonomic distribution of sequenced species, and how national participation has influenced the fields development. We show that genome quality has increased dramatically in recent years, that substantial taxonomic gaps exist, and that the field has been dominated by affluent nations in the Global North and China, despite a wide geographic distribution of sequenced species. We identify multiple disconnects between the native range of focal species and the national affiliation of the researchers studying the plants, which we argue are rooted in colonialism--both past and present. However, falling sequencing costs paired with widening availability of analytical tools and an increasingly connected scientific community provide key opportunities to improve existing assemblies, fill sampling gaps, and, most importantly, empower a more global plant genomics community.

category: plant biology


title: Tracking the Ancestry of Known and 'Ghost' Homeologous Subgenomes in Model Grass Brachypodium Polyploids

abstract: Unraveling the evolution of plant polyploids is a challenge when their diploid progenitor species are extinct or unknown or when their progenitor genome sequences are unavailable. The subgenome identification methods cannot adequately retrieve the homeologous genomes that are present in the allopolyploids if they do not take into account the potential existence of unknown progenitors. We addressed this challenge in the widely distributed dysploid grass genus Brachypodium, which is a model genus for temperate cereals and biofuel grasses. We used a transcriptome-based phylogeny and newly designed subgenome detection algorithms coupled with a comparative chromosome barcoding analysis. Our phylogenomic subgenome detection pipeline was validated in Triticum allopolyploids, which have known progenitor genomes, and was used to infer the identities of three extant and four  ghost subgenomes in six Brachypodium polyploids (B. mexicanum, B. boissieri, B. retusum, B. phoenicoides, B. rupestre and B. hybridum), of which five contain undescribed homeologous subgenomes. The existence of the seven Brachypodium progenitor genomes in the polyploids was confirmed by their karyotypic barcode profiles. Our results demonstrate that our subgenome detection method is able to uncover the ancestral genomic components of both allo- and autopolyploids.


title: Mesolimbic dopamine adapts the rate of learning from errors in performance

abstract: Recent success in training artificial agents and robots derives from a combination of direct learning of behavioral policies and indirect learning via value functions. Policy learning and value learning employ distinct algorithms that depend upon evaluation of errors in performance and reward prediction errors, respectively. In animals, behavioral learning and the role of mesolimbic dopamine signaling have been extensively evaluated with respect to reward prediction errors; however, to date there has been little consideration of how direct policy learning might inform our understanding. Here we used a comprehensive dataset of orofacial and body movements to reveal how behavioral policies evolve as naive, head-restrained mice learned a trace conditioning paradigm. Simultaneous multi-regional measurement of dopamine activity revealed that individual differences in initial reward responses robustly predicted behavioral policy hundreds of trials later, but not variation in reward prediction error encoding. These observations were remarkably well matched to the predictions of a neural network based model of behavioral policy learning. This work provides strong evidence that phasic dopamine activity regulates policy learning from performance errors in addition to its roles in value learning and further expands the explanatory power of reinforcement learning models for animal learning.


title: 3D atlas of the pituitary gland of the model fish medaka

abstract:
In vertebrates, the anterior pituitary plays a crucial role in regulating several essential physiological processes via the secretion of at least seven peptide hormones by different endocrine cell types. Comparative and comprehensive knowledge of the spatial distribution of those endocrine cell types is required to better understand their role during the animal life. Using medaka as the model and several combinations of multi-color fluorescence in situ hybridization, we present the first 3D atlas revealing the gland-wide distribution of seven endocrine cell populations: lactotropes, thyrotropes, Lh and Fsh gonadotropes, somatotropes, and pomca-expressing cells (corticotropes and melanotropes) in the anterior pituitary of a teleost fish. By combining in situ hybridization and immunofluorescence techniques, we deciphered the location of corticotropes and melanotropes within the pomca-expressing cell population. The 3D localization approach reveals sexual dimorphism of tshba- and pomca-expressing cells in the adult medaka pituitary. Finally, we show the existence of bi-hormonal cells co-expressing lhb-fshb, fshb-tshba and lhb-sl using single-cell transcriptomics analysis and in situ hybridization. This study offers a solid basis for future comparative studies of the teleost pituitary and its developmental plasticity.

Highlights- We offer the first 3D atlas of a teleost pituitary, which presents a valuable resource to the endocrinology and model fish community.
- The atlas reveals the 3D spatial distribution of the seven endocrine cell types and blood vessels in the juvenile/adult male and female pituitary.
- Gene expression for tshba and pomca, as well as the population size of cells expressing these genes, displays obvious sexual dimorphism in the adult medaka pituitary.
- Multi-color in situ hybridization and single cell RNA-seq reveal the existence of bi-hormonal cells, co-expressing lhb-fshb, fshb-tshba, lhb-sl, and a few multi-hormonal cells.
- An online version of the atlas is available at https://www.nmbu.no/go/mpg-atlas.


title: Local field potentials reflect cortical population dynamics in a region-specific and frequency-dependent manner

abstract: The spiking activity of populations of cortical neurons is well described by a small number of population-wide covariance patterns, the "latent dynamics". These latent dynamics are largely driven by the same correlated synaptic currents across the circuit that determine the generation of local field potentials (LFP). Yet, the relationship between latent dynamics and LFPs remains largely unexplored. Here, we characterised this relationship for three different regions of primate sensorimotor cortex during reaching. The correlation between latent dynamics and LFPs was frequency-dependent and varied across regions. However, for any given region, this relationship remained stable across behaviour: in each of primary motor and premotor cortices, the LFP-latent dynamics correlation profile was remarkably similar between movement planning and execution. These robust associations between LFPs and neural population latent dynamics help bridge the wealth of studies reporting neural correlates of behaviour using either type of recordings.


title: A low repeated dose of Δ9-tetrahydrocannabinol affects memory performance through serotonergic signalling in mice

abstract: Cannabis is the most widely used illicit drug worldwide. Its principal psychoactive component, {Delta}9-tetrahydrocannabinol (THC), acts as a partial agonist of the main cannabinoid receptor in the brain, the cannabinoid type-1 receptor (CB1R), that is responsible for the central effects of THC including memory impairment. CB1Rs may form heterodimers with the serotonin 5-HT2A receptor (5-HT2AR) which were found responsible for the memory impairment produced by acute high dose of THC in mice. In this study we investigated whether a repeated low dose of THC (1 mg/kg), with no acute consequence on memory performance, could eventually have deleterious cognitive effects. We found that this dose of THC impaired novel object-recognition memory and fear conditioning memory 24 h after the last of 7 consecutive daily treatments. At that time, a general enhancement of c-Fos expression was also observed in several brain regions of THC-exposed animals, as well as a decreased dendritic spine density on hippocampal CA1 pyramidal neurons that was accompanied by reduced long-term potentiation (LTP) at Schaffer collateral-CA1 synapses. Interestingly, an up-regulation in the expression of CB1R/5-HT2AR heterodimers was observed in the hippocampus after THC exposure and pre-treatment with the 5-HT2AR antagonist MDL 100,907 (0.01 mg/kg) prevented the enhanced heterodimerization and the THC-associated memory impairment. Together, these results reveal the significance of serotonergic signalling through 5-HT2ARs in the memory-impairing effects of repeated low doses of THC.


title: COVID-19 Mortality is Associated with Impaired Innate Immunity in Pre-existing Health Conditions

abstract:
BackgroundCOVID-19 poses a life-threatening endangerment to individuals with chronic diseases. However, not all comorbidities affect COVID-19 prognosis equally. Some increase the risk of COVID-19 related death by more than six folds while others show little to no impact. To prevent severe outcomes, it is critical that we comprehend pre-existing molecular abnormalities in common health conditions that predispose patients to poor prognoses. In this study, we aim to discover some of these molecular risk factors by associating gene expression dysregulations in common health conditions with COVID-19 mortality rates in different cohorts.

MethodsWe focused on fourteen pre-existing health conditions, for which age-and-sex-adjusted hazard ratios of COVID-19 mortality have been documented. For each health condition, we analyzed existing transcriptomics data to identify differentially expressed genes (DEGs) between affected individuals and unaffected individuals. We then tested if fold changes of any DEG in these pre-existing conditions were correlated with hazard ratios of COVID-19 mortality to discover molecular risk factors. We performed gene set enrichment analysis to identify functional groups overrepresented in these risk factor genes and examined their relationships with the COVID-19 disease pathway.

ResultsWe found that upregulated expression of 70 genes and downregulated expression of 181 genes in pre-existing health conditions were correlated with increased risk of COVID-19 related death. These genes were significantly enriched with endoplasmic reticulum (ER) function, proinflammatory reaction, and interferon production that participate in viral transcription and immune responses to viral infections.

ConclusionsImpaired innate immunity in pre-existing health conditions are associated with increased hazard of COVID-19 mortality. The discovered molecular risk factors are potential prognostic biomarkers and targets for therapeutic interventions.


title: Direct Energy Transfer from Allophycocyanin-Free Rod-Type CpcL-Phycobilisome to Photosystem I

abstract:
Phycobilisomes (PBSs) are photosynthetic antenna megacomplexes comprised of pigment-binding proteins (cores and rods) joined with linker proteins. A rod-type PBS that does not have a core is connected to photosystem I (PSI) by a pigment-free CpcL linker protein, which induces a red-shift of the absorption band of phycocyanobilin (PCB) in the rod (red-PCB). Herein, the isolated supercomplex of the rod-type PBS and the PSI tetramer from Anabaena sp. PCC 7120 were probed by picosecond laser spectroscopy at 77 K and by decay-associated spectral analysis to show that red-PCB mediates the fast (time constant = 90 ps) and efficient (efficiency = 95%) transfer of excitation energy from PCB in rod to chlorophyll a (Chl a) in PSI. According to the Forster energy transfer mechanism, this high efficiency corresponds to a 4-nm distance between red-PCB and Chl a, suggesting that {beta}-84 PCB in rod acts as red-PCB.

TOC GRAPHIC O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY


title: Functional organization of frontoparietal cortex in the marmoset investigated with awake resting-state fMRI

abstract:
Frontoparietal networks contribute to complex cognitive functions in humans and macaques such as working memory, attention, task-switching, response suppression, grasping, reaching, and eye movement control. However, little is known about the organization of frontoparietal networks in the New World common marmoset monkey (Callithrix jacchus) which is now widely recognized as a powerful nonhuman primate experimental animal. In this study, we employed hierarchical clustering of interareal BOLD signals to investigate the hypothesis that the organization of the frontoparietal cortex in the marmoset follows the organizational principles of the macaque frontoparietal system. We found that the posterior part of the lateral frontal cortex (premotor regions) was functionally connected to the anterior parietal areas while more anterior frontal regions (frontal eye field (FEF)) were connected to more posterior parietal areas (the area around lateral intraparietal area (LIP)). These overarching patterns of inter-areal organization are consistent with a recent macaque study. These findings demonstrate parallel frontoparietal processing streams in marmosets and support the functional homologies of FEF-LIP and premotor-anterior parietal pathways between marmoset and macaque.

Significant StatementFrontoparietal networks contribute to many cognitive functions in humans and macaques, but little is known about the organization of frontoparietal networks in the New World common marmoset monkey. Here, we investigated the hypothesis that the organization of the frontoparietal cortex in the marmoset follows the organizational principles of the macaque frontoparietal system. These frontoparietal connection showed overarching pattern of interareal organization consistent with Old world monkeys. These findings demonstrate parallel frontoparietal processing streams in marmosets.


title: Accelerated Antibody Discovery Targeting the SARS-CoV-2 Spike Protein for COVID-19 Therapeutic Potential

abstract: Rapid deployment of technologies capable of high-throughput and high-resolution screening is imperative for timely response to viral outbreaks. Risk mitigation in the form of leveraging multiple advanced technologies further increases the likelihood of identifying efficacious treatments in an aggressive timeline. In this study, we describe two parallel, yet distinct, in vivo approaches for accelerated discovery of antibodies targeting the SARS-CoV-2 spike protein. Working with human transgenic Alloy-GK mice, we detail a single B-cell discovery workflow to directly interrogate antibodies secreted from plasma cells for binding specificity and ACE2 receptor blocking activity. Additionally, we describe a concurrent accelerated hybridoma-based workflow utilizing a DiversimAb mouse model for increased diversity. The panel of antibodies isolated from both workflows revealed binding to distinct epitopes with both blocking and non-blocking profiles. Sequence analysis of the resulting lead candidates uncovered additional diversity with the opportunity for straightforward engineering and affinity maturation. By combining in vivo models with advanced integration of screening and selection platforms, lead antibody candidates can be sequenced and fully characterized within one to three months.

category: immunology


title: Spike mutation T403R allows bat coronavirus RaTG13 to use human ACE2

abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the cause of the COVID-19 pandemic, most likely emerged from bats1. A prerequisite for this devastating zoonosis was the ability of the SARS-CoV-2 Spike (S) glycoprotein to use human angiotensin-converting enzyme 2 (ACE2) for viral entry. Although the S protein of the closest related bat virus, RaTG13, shows high similarity to the SARS-CoV-2 S protein it does not efficiently interact with the human ACE2 receptor2. Here, we show that a single T403R mutation allows the RaTG13 S to utilize the human ACE2 receptor for infection of human cells and intestinal organoids. Conversely, mutation of R403T in the SARS-CoV-2 S significantly reduced ACE2-mediated virus infection. The S protein of SARS-CoV-1 that also uses human ACE2 also contains a positive residue (K) at this position, while the S proteins of CoVs utilizing other receptors vary at this location. Our results indicate that the presence of a positively charged amino acid at position 403 in the S protein is critical for efficient utilization of human ACE2. This finding could help to predict the zoonotic potential of animal coronaviruses.

category: microbiology


title: Correlation of vaccine-elicited antibody levels and neutralizing activities against SARS-CoV-2 and its variants

abstract: Both Pfizer-BNT162b2 and Moderna-mRNA-1273 vaccines can elicit an effective immune response against SARS-CoV-2 infection. However, the elicited serum antibody levels vary substantially and longitudinally decrease after vaccination. We examined the correlation of vaccination-induced IgG levels and neutralization titers against newly emerged variants remains and demonstrate a significant reduction of neutralization activities against the variants (B.1.1.7, B.1.525, and B.1.351) in Pfizer or Moderna vaccined sera. There was a significant and positive correlation between serum IgG levels and ID50 titers for not only SARS-CoV-2 WT but also the variants. These findings indicate that a high level of anti-spike IgG may offer better protection against infection from SARS-CoV-2 and its variants. Therefore, it is necessary to longitudinally monitor specific serum IgG level for evaluating the protective efficacy of the vaccines against SARS-CoV-2 and its new variants.


title: Revealing the host antiviral protein ZAP-S as an inhibitor of SARS-CoV-2 programmed ribosomal frameshifting

abstract: Programmed ribosomal frameshifting (PRF) is a fundamental gene expression event in many viruses including SARS-CoV-2, which allows production of essential structural and replicative enzymes from an alternative reading frame. Despite the importance of PRF for the viral life cycle, it is still largely unknown how and to what extent cellular factors alter mechanical properties of frameshifting RNA molecules and thereby impact virulence. This prompted us to comprehensively dissect the interplay between the host proteome and the SARS-CoV-2 frameshift element. Here, we reveal that zinc-finger antiviral protein (ZAP-S) is a direct and specific regulator of PRF in SARS-CoV-2 infected cells. ZAP-S overexpression strongly impairs frameshifting and viral replication. Using in vitro ensemble and single-molecule techniques, we further demonstrate that ZAP-S directly interacts with the SARS-CoV-2 RNA and ribosomes and interferes with the folding of the frameshift RNA. Together these data illuminate ZAP-S as de novo host-encoded specific inhibitor of SARS-CoV-2 frameshifting and expand our understanding of RNA-based gene regulation.


title: Prediction of residue-specific contributions to binding and thermal stability using yeast surface display

abstract: Accurate prediction of residue burial as well as quantitative prediction of residue-specific contributions to protein stability and activity is challenging, especially in the absence of experimental structural information. This is important for prediction and understanding of disease causing mutations, and for protein stabilization and design. Using yeast surface display of a saturation mutagenesis library of the bacterial toxin CcdB, we probe the relationship between ligand binding and expression level of displayed protein, with in vivo solubility in E.coli and in vitro thermal stability. We find that both the stability and solubility correlate well with the total amount of active protein on the yeast cell surface but not with total amount of expressed protein. We coupled FACS and deep sequencing to reconstruct the binding and expression mean fluorescent intensity of each mutant. The reconstructed mean fluorescence intensity (MFIseq) was used to differentiate between buried site, exposed non active-site and exposed active-site positions with high accuracy. The MFIseq was also used as a criterion to identify destabilized as well as stabilized mutants in the library, and to predict the melting temperatures of destabilized mutants. These predictions were experimentally validated and were more accurate than those of various computational predictors. The approach was extended to successfully identify buried and active-site residues in the receptor binding domain of the spike protein of SARS-CoV-2, suggesting it has general applicability.


title: Genomic evidence for a chemical link between redox conditions and microbial community composition

abstract: Environmental influences on community structure are often assessed through multivariate analyses in order to relate microbial abundances to separately measured physicochemical variables. However, genes and proteins are themselves chemical entities; in combination with genome databases, differences in microbial abundances directly encode for chemical variability. We predicted that the carbon oxidation state of inferred community proteomes, obtained by combining taxonomic abundances from published 16S rRNA gene sequencing datasets with predicted microbial proteomes from the NCBI Reference Sequence (RefSeq) database, would reflect environmental oxidation-reduction conditions in various natural and engineered settings including shale gas wells. Our analysis confirms the geobiochemical predictions for environmental redox gradients within and between hydrothermal systems and stratified lakes and marine environments. Where they are present, a common set of taxonomic groups (Gamma- and Deltaproteobacteria and Clostridia) act as drivers of the community-level differences in oxidation state, whereas Flavobacteria most often oppose the overall changes. The geobiochemical signal is largest for the steep redox gradients associated with hydrothermal systems and between surface water and produced fluids from shale gas wells, demonstrating the ability to determine the magnitude of redox effects on microbial communities from 16S sequencing alone.


title: Phenotypic and transcriptomic analyses reveal major differences between apple and pear scab nonhost resistance

abstract:
BackgroundNonhost resistance is the outcome of most plant/pathogen interactions, but it has rarely been described in Rosaceous fruit species. Apple (Malus x domestica Borkh.) is a nonhost for Venturia pyrina, the scab species attacking European pear (Pyrus communis L.). Reciprocally, P. communis is a nonhost for Venturia inaequalis, the scab species attacking apple. The major objective of our study was to compare the scab nonhost resistance in apple and in European pear, at the phenotypic and transcriptomic levels.

ResultsMacro- and microscopic observations after reciprocal scab inoculations indicated that, after a similar germination step, nonhost apple/V. pyrina interaction remained nearly symptomless, whereas hypersensitive reactions were observed during nonhost pear/V. inaequalis interaction. Comparative transcriptomic analyses of apple and pear nonhost interactions with V. pyrina and V. inaequalis, respectively, revealed considerable differences. Very few differentially expressed genes were detected during apple/V. pyrina interaction, which is consistent with a symptomless type I nonhost resistance. On the contrary, numerous genes were differentially expressed during pear/V. inaequalis interaction, as expected in a type II nonhost resistance involving visible hypersensitive reaction. Pre-invasive defense, such as stomatal closure, was detected, as well as several post-invasive defense mechanisms (apoplastic reactive oxygen species accumulation, phytoalexin production and alterations of the epidermis composition). In addition, a comparative analysis between pear scab host and nonhost interactions indicated that, although specificities were observed, two major defense lines were shared in these resistances: cell wall and cuticle modifications and phenylpropanoid pathway induction.

ConclusionThis first deciphering of the molecular mechanisms underlying a nonhost scab resistance in pear offers new possibilities for the genetic engineering of sustainable scab resistance in this species.


title: RNA surveillance controls 3D genome structure via stable cohesin-chromosome interaction

abstract: The 3D architecture that the genome is folded into is regulated by CTCF, which determines domain borders, and cohesin, which generates interactions within domains. However, organisms lacking CTCF have been reported to still have cohesin-mediated 3D structures with strong borders. How this can be achieved and precisely regulated are yet unknown. Using in situ Hi-C, we found that 3-end RNA processing factors coupled with proper transcription termination are a cis-acting determinant that regulates the localization and dynamics of cohesin on the chromosome arms. Loss of RNA processing factors, including nuclear exosome and Pfs2, destabilizes cohesin from the 3-ends of convergent genes and results in decreased cohesin-mediated domain boundaries. We observed the co-localization between Rad21 and a wide range of 3end RNA processing/termination factors. Further, deletion of Rrp6 leads to cohesin redistribution to facultative heterochromatin, resulting in improper domain boundaries. Importantly, we observed that knockdown of Rrp6Exosc10 caused a defect in cohesin binding and loss of local topologically associating domains (TADs) interactions in mouse embryonic stem cells. Based on these findings, we propose a novel function of the RNA surveillance pathway in 3D genome organization via cohesin complex, which provides the molecular basis underlying the dynamics of cohesin function.


title: Development, validation, and application of the ribosome separation and reconstitution system for protein translation in vitro

abstract: The conventional view regarding regulation of gene expression is based on transcription control. However, a growing number of recent studies has revealed the important additional impact of translational regulation. Eukaryotic translational machinery appears to be capable of reprogramming mRNA translation to generate proteins required to maintain a healthy cellular proteostasis under particular physiological conditions or to adapt to stress. Although the mechanisms of such remarkable regulation are beginning to emerge, recent studies have identified the ribosome as one of the major constituents of translation-dependent control of gene expression that is especially important during stress. Built of RNA and proteins, ribosomes are susceptible to environmental and intracellular stresses. How stress-modified ribosomes regulate translation and whether they play a role in stress-induced gene expression remain largely elusive. This knowledge gap is likely due to the lack of an appropriate experimental system. Canonical approaches based on exposing cells or cell-free extracts to stressors provide inconclusive results due to off-target effects of modifying agents. Here we describe a robust and simple in vitro assay that allows separation of yeast ribosomes from other translational machinery constituents, followed by reconstitution of the translation reaction. This ribosome separation and reconstitution assay (RSR) is highly advantageous, as it allows modification of ribosomes without compromising other key translational components, followed by supplementing the ribosomes back into translation reactions containing undamaged, translationally-competent yeast lysate. Besides addressing the impact of ribosome-derived stress on translation, RSR can also be used to characterize mutated ribosomes and ribosomes devoid of associated factors.


title: Phylogenetic context using phylogenetic outlines

abstract: AO_SCPLOWBSTRACTC_SCPLOWMicrobial studies typically involve the sequencing and assembly of draft genomes for individual microbes or whole microbiomes. Given a draft genome, one first task is to determine its phylogenetic context, that is, to place it relative to the set of related reference genomes. We provide a new interactive graphical tool that addresses this task using Mash sketches to compare against all bacterial and archaeal representative genomes in the GTDB taxonomy, all within the framework of SplitsTree5. The phylogenetic context of the query sequences is then displayed as a phylogenetic outline, a new type of phylogenetic network that is more general that a phylogenetic tree, but significantly less complex than other types of phylogenetic networks. We propose to use such networks, rather than trees, to represent phylogenetic context, because they can express uncertainty in the placement of taxa, whereas a tree must always commit to a specific branching pattern. We illustrate the new method using a number of draft genomes of different assembly quality.


title: Infection of mouse neural progenitor cells by Toxoplasma gondii affects in vitro proliferation, differentiation and migration

abstract: Congenital toxoplasmosis constitutes a major cause of pre- and post-natal complications. Fetal infection with Toxoplasma gondii influences development and can lead to microcephaly, encephalitis, and neurological abnormalities. Few studies have attempted to explain the impact of T. gondii infection on the physiology of mature nerve cells, and no systematic study concerning the effect of infection of neural progenitor cells by T. gondii in the biology of these progenitors is available. We infected cortical intermediate progenitor cell cultivated as neurospheres obtained from E16.5 Swiss Webster mice with T. gondii (Me49 strain) tachyzoites to mimic the developing mouse cerebral cortex in vitro. Infection decreased cell proliferation as detected by Ki67 staining at 48 and 72 hours post infection (hpi) in floating neurospheres, resulting in reduced cellularity at 96 hpi. Neurogenic and gliogenic potential, assessed in plated neurospheres, was shown to be impaired in infected cultures, as indicated by neurofilament heavy chain (NF-200) and GFAP staining, respectively. To further investigate the impact of infection on neuronal differentiation, Neuro2a neuroblasts were infected and after 24 hpi, neurogenic differentiation was induced with serum withdrawal. We confirmed that infection induces a decrease in neuroblast-neuron differentiation rates in cells stained for NF-200, with reduced neuritogenesis. Migration rates were analyzed in plated neurospheres. At 120 h after plating, infected cultures exhibited decreased overall migration rates and altered the radial migration of nestin-, GFAP- and NF-200-positive cells. These findings indicate that T. gondii infection of neural progenitor cells may lead to reduced neuro/gliogenesis due to an imbalance in cell proliferation alongside an altered migratory profile. If translated to the in vivo situation, these data could explain, in part, the cortical malformations observed in congenitally infected individuals.


title: CamelliA-based simultaneous imaging of Ca2+ dynamics in subcellular compartments

abstract:
As a universal second messenger, calcium (Ca2+) transmits specific cellular signals via a spatiotemporal signature generated from its extracellular source and internal stores. Our knowledge of the mechanisms underlying generation of a Ca2+ signature is hampered by limited tools enabling simultaneous monitoring of the dynamics of Ca2+ levels in multiple subcellular compartments. To overcome the limitation and to further improve spatiotemporal resolutions, here we have assembled a molecular toolset (the CamelliA lines) in Arabidopsis that enables simultaneous and high-resolution monitoring of Ca2+ dynamics in multiple subcellular compartments through imaging analyses of different single-colored GECIs (Genetically Encoded Calcium Indicators). Indeed, the uncovering of the previously unrecognized Ca2+ signatures in three types of Arabidopsis cells in response to internal and external cues is a testimony to the wide applicability of the newly generated toolset for elucidating the subcellular sources contributing to the Ca2+signatures in plants.

One sentence summaryA toolset for simultaneous imaging of Ca2+ dynamics in subcellular compartments has uncovered unrecognized Ca2+ signatures in Arabidopsis cells in response to developmental and external cues.


title: TORC1 and PKA activity towards ribosome biogenesis oscillates in synchrony with the budding yeast cell cycle

abstract: Recent studies have revealed that the growth rate of budding yeast and mammalian cells varies during the cell cycle. By linking a multitude of signals to cell growth, the highly conserved Target of Rapamycin Complex 1 (TORC1) and Protein Kinase A (PKA) pathways are prime candidates for mediating the dynamic coupling between growth and division. However, measurements of TORC1 and PKA activity during the cell cycle are still lacking. Following the localization dynamics of two TORC1 and PKA targets via time-lapse microscopy in hundreds of yeast cells, we found that the activity of these pathways towards ribosome biogenesis fluctuates in synchrony with the cell cycle even under constant external conditions. Mutations of upstream TORC1 and PKA regulators suggested that internal metabolic signals partially mediate these activity changes. Our study reveals a new aspect of TORC1 and PKA signaling, which will be important for understanding growth regulation during the cell cycle.

category: systems biology


title: Reverse engineering of a mechanistic model of gene expression using metastability and temporal dynamics

abstract: Differentiation can be modeled at the single cell level as a stochastic process resulting from the dynamical functioning of an underlying Gene Regulatory Network (GRN), driving stem or progenitor cells to one or many differentiated cell types. Metastability seems inherent to differentiation process as a consequence of the limited number of cell types. Moreover, mRNA is known to be generally produced by bursts, which can give rise to highly variable non-Gaussian behavior, making the estimation of a GRN from transcriptional profiles challenging. In this article, we present CARDAMOM (Cell type Analysis from scRna-seq Data achieved from a Mixture MOdel), a new algorithm for inferring a GRN from timestamped scRNA-seq data, which crucially exploits these notions of metastability and transcriptional bursting. We show that such inference can be seen as the successive resolution of as many regression problem as timepoints, after a preliminary clustering of the whole set of cells with regards to their associated bursts frequency. We demonstrate the ability of CARDAMOM to infer a reliable GRN from in silico expression datasets, with good computational speed. To the best of our knowledge, this is the first description of a method which uses the concept of metastability for performing GRN inference.


title: Ciliated protozoa from industrial WWTP activated sludge: a biodiversity survey including trophic interactions and redescription of Bakuella subtropica (Spirotrichea, Hypotrichia) according to Next Generation Taxonomy

abstract: Optimization of wastewater treatment with biological processes is a fundamental challenge of modern society. During past years new technologies have been developed for the purpose and prokaryotic organisms involved in the process extensively investigated. Nevertheless, relatively few studies so far analysed the protozoan community in these systems using modern integrative approaches, despite its obvious role in shaping ecological dynamics and, possibly, process efficiency. In the present study, we characterized the ciliate community in biological reactors of an Italian industrial (tannery) wastewater treatment plant (WWTP) applying modified Ludzack-Ettinger (MLE) process. This plant is characterized by moderate salinity, high solids retention time and high concentration of organic compounds, including a significant recalcitrant fraction. We performed the morphological and 18S rDNA characterizations of almost all the 21 ciliates retrieved along a one-year sampling period, and provided preliminary data on species occurrence, community dynamics, and trophic interactions. Only 16 species were observed on the sample collection day and most of them had an occurrence higher than 50%. The most frequently occurring and highly abundant organisms were Aspidisca cf. cicada, Euplotes spp., Paramecium calkinsi, and Phialina sp. Cyclidium cf. marinum was only found on a single date and its presence was possibly related to a summer break-induced perturbation. All the species showed the capability to survive the short oxic/anoxic cycling typical of the studied WWTP process. Intriguingly, some of them (i.e., Bakuella subtropica and Trochiliopsis australis) turned out to be species isolated from brackish natural environment rich in organic load as well. As for B. subtropica, we provided an emended redescription according to the most recent taxonomy standards that include also mitogenomic sequencing.

category: zoology


title: Enhanced negative emotional processing limits behavioral flexibility in a mouse model of autism spectrum disorder

abstract: Impaired behavioral flexibility might underlie some of the symptoms associated with autism spectrum disorder (ASD). We investigated whether and how behavioral flexibility is impaired in a mouse model of ASD by testing Shank2-knockout (Shank2-KO) mice in reversal learning. Shank2-KO mice were trained in probabilistic classical conditioning with two odor cues paired with water and air puff. Upon the reversal of cue-outcome contingency, Shank2-KO mice were significantly slower than wild-type mice in reversing their anticipatory licking responses. Shank2-KO mice also showed stronger anticipatory eye closure responses than wild-type mice to the air puff, raising a possibility that the impairment might be because of enhanced negative emotional processing. Indeed, Shank2-KO mice showed intact reversal learning when the strong air puff was replaced with a mild air puff. Shank2-KO mice also showed intact reversal learning between two odor cues predicting rewards with different probabilities. These results indicate that enhanced negative emotional processing suppresses reversal learning despite of intact capability to learn cue-outcome contingency changes in Shank2-KO mice in our behavioral settings. Our findings suggest that behavioral flexibility may be seriously limited by abnormal emotional processing in ASD.


title: Optogenetic stimulation of primate V1 reveals local laminar and large-scale cortical networks related to perceptual phosphenes

abstract: Developing optogenetics in non-human primates (NHPs) is essential for translating its successful implementation in rodents to clinical applications in humans. However, information about how optogenetics influences the primate cortex remains limited. Here, we evaluate how optogenetic stimulation of the primate primary visual cortex (V1) affects local and large-scale network activation concerned with visual perception. To this end we injected an optogenetic construct (AAV9-hSyn-ChR2-eYFP) into the V1 cortex of four macaque monkeys (macaca mulatta) and measured the effects of optogenetic V1 stimulation using functional magnetic resonance imaging (fMRI), laminar electrophysiology, and behavioural assessment. In three macaques, blood-oxygen-dependent (BOLD) fMRI activity could be reliably elicited with optogenetic stimulation in V1 and several connected extrastriate brain areas, including V2/V3, motion-sensitive area MT and the frontal-eye-fields (FEF), in particular when pulsed stimulation at 40 Hz was applied. BOLD modulation was associated with consistent neural spiking activity measured in V1 of two macaques. More detailed analysis revealed strongest neuronal activation in layer 4B and infragranular layers, which tightly reflected the histological expression pattern of the optogenetic construct in V1. Driving this visual network proved sufficient to elicit a visual percept ( phosphene) in one macaque during a perceptual choice task. Taken together, our findings reveal the laminar and large-cortical activation pattern related to visual phosphene generation and emphasize the need for further improving optogenetic methods in NHPs as a step towards applications in humans.


title: Decoding continuous variables from EEG data using linear support vector regression (SVR) analysis with the Decision Decoding Toolbox (DDTBOX)

abstract: Multivariate classification analysis for non-invasively acquired neuroimaging data is a powerful tool in cognitive neuroscience research. However, an important constraint of such pattern classifiers is that they are restricted to predicting categorical variables (i.e. assigning trials to classes). Here, we present an alternative approach, Support Vector Regression (SVR), which uses single-trial neuroimaging (e.g., EEG or MEG) data to predict a continuous variable of interest such as response time, response force, or any kind of subjective rating (e.g., emotional state, confidence, etc.). We describe how SVR can be used, how it is implemented in the Decision Decoding Toolbox (DDTBOX), and how it has been used in previous research. We then report results from two simulation studies, designed to closely resemble real EEG data, in which we predicted a continuous variable of interest across a range of analysis parameters. In Simulation Study 1, we observed that SVR was effective for analysis windows ranging from 2 ms - 100 ms, and that it was relatively unaffected by temporal averaging. In Simulation Study 2, we showed that prediction was still successful when only a small number of channels encoded information about the output variable, and that it was robust to temporal jitter regarding when that information was present in the EEG. Finally, we reanalysed a previously published dataset of similar size and observed highly comparable results in real EEG data. We conclude that linear SVR is a powerful tool for the investigation of single-trial EEG data in relation to continuous and more nuanced variables, which are not well-captured using classification approaches requiring distinct classes.


title: PGfinder, a novel analysis pipeline for the consistent, reproducible and high- resolution structural analysis of bacterial peptidoglycans

abstract: Many software solutions are available for proteomics and glycomics studies, but none are ideal for the structural analysis of peptidoglycan, the essential and major component of bacterial cell envelopes. It is comprised of glycan chains and peptide stems, both containing unusual amino acids and sugars. This has forced the field to rely on manual analysis approaches, which are time-consuming, labour-intensive, and prone to error. The lack of automated tools has hampered the ability to perform high-throughput analyses and prevented the adoption of a standard methodology. Here, we describe a novel tool called PGfinder for the analysis of peptidoglycan structure and demonstrate that it represents a powerful tool to quantify PG fragments and discover novel structural features. Our analysis workflow, which relies on open-access tools, is a breakthrough towards a consistent and reproducible analysis of bacterial peptidoglycans. It represents a significant advance towards peptidoglycomics as a full-fledged discipline.


title: More than a stick in the mud: Eelgrass leaf and root bacterial communities are distinct from those on physical mimics

abstract:
We examine the role of physical structure vs. biotic interactions in structuring host-associated microbial communities on a marine angiosperm, Zostera marina, eelgrass. Across several months and sites, we compared microbiomes on physical mimics of eelgrass roots and leaves to those on intact plants. We find large, consistent differences in the microbiome of mimics and plants, especially on roots, but also on leaves. Key taxa that are more abundant on leaves have been associated with microalgal and macroalgal disease and merit further investigation to determine their role in mediating plant-microalgal-pathogen interactions. Root associated taxa were associated with sulfur and nitrogen cycling, potentially ameliorating environmental stresses for the plant. Our work identifies targets for future work on the functional role of the seagrass microbiome in promoting the success of these angiosperms in the sea.

Originality-Significance StatementWe show that eelgrass establishes a distinct microbial community from a physical mimic on both its leaves and roots. This is, to our knowledge, the first comparison of seagrass to a mimicked physical environment. Insights from our study establish bacterial targets for future functional studies of seagrass-microbiome interactions.


title: Genome characterization of 'Candidatus Phytoplasma meliae' (isolate ChTYXIII)

abstract: Candidatus Phytoplasma meliae (subgroups 16SrXIII-G and XIII-C) has been reported in association to chinaberry yellowing disease in Argentina, Bolivia and Paraguay. In Argentina, this disease constitutes a major phytosanitary problem for chinaberry forestry production. To date, no genome information of this phytoplasma and others from 16SrXIII-group has been published, hindered its characterization at genomic level. Here we analyze the draft genome of  Candidatus Phytoplasma meliae strain ChTYXIII obtained from a chinaberry-infected plant using a metagenomics approach. The draft assembly consists of twenty-one contigs with a total length of 751.949 bp. The annotation contains 669 CDSs, 34tRNA and one set of rRNA operons. Metabolic pathways analysis indicated that the ChTYXIII contains the complete core genes for glycolysis and functional sec system for translocation of proteins. The phylogenetic relationships inferred 132 single copy genes (orthologues core) analysis revealed that  Ca. P. meliae constitutes a clade closely related to the  Ca. australiense and  Ca. P. solani. Thirty-one putative effectors were identified, among which a homologue to SAP11 was found and others that have only been described in this pathogen. This work provides relevant genomic information for  Ca. P. meliae and constitutes the first genome described for the group 16SrXIII (MPV).


title: Population and Individual Firing Behaviors in Sparsely Synchronized Rhythms in The Hippocampal Dentate Gyrus

abstract: We investigate population and individual firing behaviors in sparsely synchronized rhythms (SSRs) in a spiking neural network of the hippocampal dentate gyrus (DG). The main encoding granule cells (GCs) are grouped into lamellar clusters. In each GC cluster, there is one inhibitory (I) basket cell (BC) along with excitatory (E) GCs, and they form the E-I loop. Winner-take-all competition, leading to sparse activation of the GCs, occurs in each GC cluster. Such sparsity has been thought to enhance pattern separation performed in the DG. During the winner-take-all competition, SSRs are found to appear in each population of the GCs and the BCs through interaction of excitation of the GCs with inhibition of the BCs. Sparsely synchronized spiking stripes appear successively with the population frequency fp (= 13 Hz) in the raster plots of spikes. We also note that excitatory hilar mossy cells (MCs) control the firing activity of the GC-BC loop by providing excitation to both the GCs and the BCs. SSR also appears in the population of MCs via interaction with the GCs (i.e., GC-MC loop). Population behaviors in the SSRs are quantitatively characterized in terms of the synchronization measures. In addition, we investigate individual firing activity of GCs, BCs, and MCs in the SSRs. Individual GCs exhibit random spike skipping, leading to a multi-peaked inter-spike-interval histogram, which is well characterized in terms of the random phase-locking degree. In this case, population-averaged mean-firing-rate [Formula] is less than the population frequency fp. On the other hand, both BCs and MCs show "intrastripe" burstings within stripes, together with "interstripe" random spike skipping. Thus, the population-averaged [Formula] (X = MC and BC) is larger than fp, in contrast to the case of the GCs. MC loss may occur during epileptogenesis. With decreasing the fraction of the MCs, changes in the population and individual firings in the SSRs are also studied. Finally, quantitative association between the population/individual firing behaviors in the SSRs and the winner-take-all competition is discussed.


title: The metabolome and lipidome of colorectal adenomas and cancers

abstract:
IntroductionIn-depth knowledge of metabolic dysregulations in colorectal cancer (CRC) (and other cancers as well) is essential for developing treatments that specifically kill neoplastic cells. It may also allow us to pinpoint metabolites or lipids with potential for development as tumor biomarkers for use in body-fluid or breath assays. CRC onset is preceded by an interval of [~]10 years characterized by the presence of precancerous lesions, and our previous studies have revealed epigenomic, transcriptomic, and proteomic evidence in these lesions of certain metabolic changes typical of CRC. These findings prompted us to conduct untargeted metabolomic and lipidomic analyses of CRCs and colorectal adenomas (the most common precancerous lesions of the gut).

MethodsWe analyzed 29 endoscopically collected tumor tissue samples (29 adenomas [ADNs], 10 CRCs, each with a colon segment-matched sample of normal mucosa [i.e., 29 NM-ADN, 10 NM-CRC]). The freshly collected samples were promptly frozen in liquid nitrogen and later processed to obtain metabolite and lipid extracts. Each of the 78 samples was analyzed with nano-flow LC-MS/MS (liquid chromatography with mass spectrometry) to characterize its metabolome (using HILIC, Hydrophilic Interaction Liquid Chromatography) and lipidome (using RP, Reversed Phase chromatography). The data acquired were processed using Progenesis QI. For statistical and multivariate analysis of the resulting peak tables, we used basic R packages and the R package made4.

ResultsUnsupervised between-group analysis based on the full set of detected metabolites (n=1830) and lipids (n=2365) clearly discriminated ADNs and CRCs from their matched samples of normal mucosa at both the metabolome and lipidome levels. Compared with the NM-ADN, the ADNs contained significantly different levels of 14.6% of the metabolites and 10.8% of the lipids. Fewer compounds (9.1% of metabolites, 6.2% of lipids) displayed differential abundance in CRCs (vs. NM-CRC). The metabolome and lipidome of the NM-ADN also differed from those of the NM-CRC, probably reflecting the presence of a field cancerization effect exerted by the invasive tumors. A substantial number of metabolites (n=340) and lipids (n=201) also displayed abundance differentials across the sequential tumorigenic stages represented by the NM-ADN (considered more representative of NM from a lesion-free colon) [->] ADN [->] CRC. In most cases, the trend consisted of progressive increases or progressive decreases in abundance as the tumorigenesis advanced.

ConclusionsOur findings provide a preliminary picture of the progressive metabolomic and lipidomic changes occurring during the adenomatous phase of colorectal tumorigenesis. Once definitively annotated, the numerous differentially abundant compounds detected in this study may well shed valuable light on the metabolic dysregulations occurring during this process and provide useful clues for the development of novel tools for the diagnosis and treatment of colorectal tumors.

category: cancer biology


title: A multi-tiered map of EMT defines major transition points and identifies vulnerabilities

abstract: Epithelial to mesenchymal transition (EMT) is a complex cellular program proceeding through a hybrid E/M state linked to cancer-associated stemness, migration and chemoresistance. Deeper molecular understanding of this dynamic physiological landscape is needed to define events which regulate the transition and entry into and exit from the E/M state. Here, we quantified >60,000 molecules across ten time points and twelve omic layers in human mammary epithelial cells undergoing TGF{beta}-induced EMT. Deep proteomic profiles of whole cells, nuclei, extracellular vesicles, secretome, membrane and phosphoproteome defined state-specific signatures and major transition points. Parallel metabolomics showed metabolic reprogramming preceded changes in other layers, while single-cell RNA sequencing identified transcription factors controlling entry into E/M. Covariance analysis exposed unexpected discordance between the molecular layers. Integrative causal modeling revealed co-dependencies governing entry into E/M that were verified experimentally using combinatorial inhibition. Overall, this dataset provides an unprecedented resource on TGF{beta} signaling, EMT and cancer.


title: Cryo-EM structures of human RNA polymerase I

abstract: RNA polymerase I (Pol I) specifically synthesizes ribosomal RNA. Pol I upregulation is linked to cancer, while mutations in the Pol I machinery lead to developmental disorders. Here, we report the cryo-EM structure of elongating human Pol I at 2.7 [A] resolution. In the exit tunnel, we observe a double-stranded RNA helix that may support Pol I processivity. Our structure confirms that human Pol I consists of 13 subunits with only one subunit forming the Pol I stalk. Additionally, the structure of human Pol I in complex with the initiation factor RRN3 at 3.1 [A] resolution reveals stalk flipping upon RRN3 binding. We also observe an inactivated state of human Pol I bound to an open DNA scaffold at 3.3 [A] resolution. Lastly, the high-resolution structure of human Pol I allows mapping of disease-related mutations that can aid understanding of disease etiology.

category: molecular biology


title: VIQoR: a web service for Visually supervised protein Inference and protein Quantification

abstract:
MotivationIn quantitative bottom-up mass spectrometry (MS)-based proteomics the reliable estimation of protein concentration changes from peptide quantifications between different biological samples is essential. This estimation is not a single task but comprises the two processes of protein inference and protein abundance summarization. Furthermore, due to the high complexity of proteomics data and associated uncertainty about the performance of these processes, there is a demand for comprehensive visualization methods able to integrate protein with peptide quantitative data including their post-translational modifications. Hence, there is a lack of a suitable tool that provides post-identification quantitative analysis of proteins with simultaneous interactive visualization.

ResultsIn this article, we present VIQoR, a user-friendly web service that accepts peptide quantitative data of both labeled and label-free experiments and accomplishes the processes for relative protein quantification, along with interactive visualization modules, including the novel VIQoR plot. We implemented two parsimonious algorithms to solve the protein inference problem, while protein summarization is facilitated by a well established factor analysis algorithm called fast-FARMS followed by a weighted average summarization function that minimizes the effect of missing values. In addition, summarization is optimized by the so-called Global Correlation Indicator (GCI). We test the tool on three publicly available ground truth datasets and demonstrate the ability of the protein inference algorithms to handle degenerate peptides. We furthermore show that GCI increases the accuracy of the quantitative analysis in data sets with replicated design.

Availability and implementationVIQoR is accessible at: http://computproteomics.bmb.sdu.dk:8192/app_direct/VIQoR/

The source code is available at: https://bitbucket.org/vtsiamis/viqor/

Contactveits@bmb.sdu.dk

Supplementary informationSupplementary data are available at Bioinformatics online.