title: Typhoid toxin sorting and exocytic transport from Salmonella Typhi infected cells

abstract: Typhoid toxin is an essential virulence factor for Salmonella Typhi, the cause of typhoid fever in humans. This toxin has an unusual biology in that it is produced by Salmonella Typhi only when located within host cells. Once synthesized, the toxin is secreted to the lumen of the Salmonella-containing vacuole from where it is transported to the extracellular space by vesicle carrier intermediates. Here we report the identification of the typhoid toxin sorting receptor and the cellular machinery that packages the toxin into vesicle carriers, and exports it to the extracellular space. We found that the cation- independent mannose-6-phosphate receptor serves as typhoid toxin sorting receptor and that the coat protein COPII and the GTPase Sar1 mediate its packaging into vesicle carriers. Formation of the typhoid toxin carriers requires the specific environment of the Salmonella Typhi-containing vacuole, which is determined by the activities of specific effectors of its type III protein secretion systems. We also found that Rab11B and its interacting protein Rip11 control the intracellular transport of the typhoid toxin carriers, and the SNARE proteins VAMP7, SNAP23, and Syntaxin 4 their fusion to the plasma membrane. Typhoid toxins cooption of specific cellular machinery for its transport to the extracellular space illustrates the remarkable adaptation of an exotoxin to exert its function in the context of an intracellular pathogen.


title: Harnessing DSB repair to promote efficient homology-dependent and -independent prime editing

abstract: Prime editing recently emerged as a next-generation approach for precise genome editing. Here we exploit DNA double-strand break (DSB) repair to develop two novel strategies that install precise genomic insertions using an SpCas9 nuclease-based prime editor (PEn). We first demonstrate that PEn coupled to a regular prime editing guide RNA (pegRNA) efficiently promotes short genomic insertions through a homology-dependent DSB repair mechanism. While PEn editing lead to increased levels of by-products, it rescued pegRNAs that performed poorly with a nickase-based prime editor. We also present a small molecule approach that yielded increased product purity of PEn editing. Next, we developed a homology-independent PEn editing strategy by engineering a single primed insertion gRNA (springRNA) which installs genomic insertions at DSBs through the non-homologous end joining pathway (NHEJ). Lastly, we show that PEn-mediated insertions at DSBs prevent Cas9-induced large chromosomal deletions and provide evidence that continuous Cas9-mediated cutting is one of the mechanisms by which Cas9-induced large deletions arise. Altogether, this work expands the current prime editing toolbox by leveraging distinct DNA repair mechanisms including NHEJ, which represents the primary pathway of DSB repair in mammalian cells.

category: molecular biology


title: Asymmetric Redundancy of Soybean Nodule Inception (NIN) Genes in Root Nodule Symbiosis

abstract: NIN is one of the most important root nodule symbiotic genes as it is required for both infection and nodule organogenesis in legume. Unlike most legumes with a sole NIN gene, there are four putative NIN genes in soybean (Glycine max). Whether and how these orthologs NIN genes contribute to soybean-rhizobia symbiotic interaction remain unknown. In this study, we found that all four GmNIN genes are induced by rhizobia, and that conserved CE and CYC binding motifs in their promoter regions are required for their expression in the nodule formation process. By generation of multiplex Gmnin mutants, we found that Gmnin1a nin2a nin2b triple mutant and Gmnin1a nin1b nin2a nin2b quadruple mutant displayed similar defects in rhizobia infection and root nodule formation, Gmnin2a nin2b produced less nodules but displayed hyper infection phenotype than wild type, while a Gmnin1a nin1b double mutant nodulated as wild type. Overexpression of GmNIN1a, GmNIN1b, GmNIN2a, and GmNIN2b reduced nodule numbers after rhizobia inoculation, with GmNIN1b overexpression having the weakest effect. In addition, overexpression of GmNIN1a, GmNIN2a, or GmNIN2b, but not GmNIN1b, produced malformed pseudo-nodule like structures without rhizobia inoculation. In conclusion, GmNIN1a, GmNIN2a and GmNIN2b play functionally redundant yet complicated roles for soybean nodulation. GmNIN1b, although is expressed at comparable level with other homologs, plays a minor role in root nodule symbiosis. Our work provides insight into the understanding of asymmetrically redundant function of GmNIN genes in soybean.

category: developmental biology


title: Characterization of the composition and functioning of the Crumbs complex in  C. elegans

abstract: The apical domain of epithelial cells can acquire a diverse array of morphologies and functions, which is critical for the function of epithelial tissues. The Crumbs proteins are evolutionary conserved transmembrane proteins with essential roles in promoting apical domain formation in epithelial cells. The short intracellular tail of Crumbs proteins interacts with a variety of proteins, including the scaffolding protein Pals1 (protein associated with LIN7, Stardust in Drosophila). Pals1 in turn binds to a second scaffolding protein termed PATJ (Pals1-associated tight junction protein), to form the core Crumbs/Pals1/PATJ Crumbs complex. While essential roles in epithelial organization have been shown for Crumbs proteins in Drosophila and mammalian systems, the three Caenorhabditis elegans crumbs genes are dispensable for epithelial polarization and animal development. Moreover, the presence and functioning of orthologs of Pals1 and PATJ has not been investigated. Here, we identify MAGU-2 and MPZ-1 as the C. elegans orthologs of Pals1 and PATJ, respectively. We show that MAGU-2 interacts with all three Crumbs proteins as well as MPZ-1, and localizes to the apical membrane domain in a Crumbs-dependent fashion. Similar to crumbs mutants, a magu-2 null mutant shows no developmental or epithelial polarity defects. Finally, we show that overexpression of the Crumbs proteins EAT-20 or CRB-3 in the C. elegans intestine can lead to apical membrane expansion. Our results shed light into the composition of the C. elegans Crumbs complex and indicate that the role of Crumbs proteins in promoting apical domain formation is conserved.


title: Cleavage of histone H2A during embryonic stem cell differentiation destabilizes nucleosomes to counteract gene activation

abstract: Histone proteolysis is a poorly understood phenomenon in which the N-terminal tails of histones are irreversibly cleaved by intracellular proteases. During development, histone post-translational modifications are known to orchestrate gene expression patterns that ultimately drive cell fate decisions. Therefore, deciphering the mechanisms of histone proteolysis is necessary to enhance the understanding of cellular differentiation. Here we show that H2A is cleaved by the lysosomal protease Cathepsin L during ESCs differentiation. Using quantitative mass spectrometry (MS), we identified L23 to be the primary cleavage site that gives rise to the clipped form of H2A (cH2A), which reaches a maximum level of [~]1% of total H2A after four days of differentiation. Using ChIP-seq, we found that preventing proteolysis leads to an increase in acetylated H2A at promoter regions in differentiated ES cells. We also report the identification of novel readers of acetylated H2A in pluripotent ES cells, including members of the PBAF remodeling complex, which can recognize different acetylated forms of H2A. Finally, we show that H2A proteolysis abolishes this recognition. Altogether, our data suggests that proteolysis serves as an efficient mechanism to silence pluripotency genes and destabilize the nucleosome core particle.


title: Rationalizing generation of broad spectrum antibiotics with the addition of a primary amine

abstract: Antibiotic resistance of Gram-negative bacteria is largely attributed to the low permeability of their outer membrane (OM). Recently, we disclosed the eNTRy rules, a key lesson of which is that the introduction of a primary amine enhances OM permeation in certain contexts. To understand the molecular basis for this finding, we perform an extensive set of molecular dynamics (MD) simulations and free energy calculations comparing the permeation of aminated and amine-free antibiotic derivatives through the most abundant OM porin of E. coli, OmpF. To improve sampling of conformationally flexible drugs in MD simulations, we developed a novel, Monte Carlo and graph theory based algorithm to probe more efficiently the rotational and translational degrees of freedom visited during the permeation of the antibiotic molecule through OmpF. The resulting pathways were then used for free-energy calculations, revealing a lower barrier against the permeation of the aminated compound, substantiating its greater OM permeability. Further analysis revealed that the amine facilitates permeation by enabling the antibiotic to align its dipole to the luminal electric field of the porin and while forming favorable electrostatic interactions with specific, highly-conserved charged residues. The importance of these interactions in permeation was further validated with experimental mutagenesis and whole cell accumulation assays. Overall, this study provides insights on the importance of the primary amine for antibiotic permeation into Gram-negative pathogens that could help the design of future antibiotics. We also offer a new computational approach for calculating free-energy of processes where relevant molecular conformations cannot be efficiently captured.

category: biophysics


title: A novel approach to evaluate alpha-synuclein seeding shows a wide heterogeneity in multiple system atrophy

abstract: Several in vitro and in vivo findings have consistently shown that -synuclein derived from multiple system atrophy (MSA) subjects has more seeding capacity than Parkinsons disease-derived -synuclein. However, reliable detection of -synuclein derived from MSA using seeded amplification assays, such as the Real-Time Quaking-induced Conversion, has remained challenging. Here we demonstrate that the interaction of the Thioflavin T dye with -synuclein from MSA and Parkinsons disease patients can be modulated by the type of salt, pH, and ionic strength used to generate strain-specific reaction buffers. Employing this novel approach, we have generated a streamlined Real-Time Quaking-induced Conversion assay capable of categorizing MSA brains according to their -synuclein seeding behavior, and to unravel a previously unrecognized heterogeneity in seeding activity between different brain regions of a given individual that goes beyond immunohistochemical observations and provide a framework for future molecular subtyping of MSA.

category: neuroscience


title: Infectious SARS-CoV-2 is emitted in aerosols

abstract: Respiratory viruses such as SARS-CoV-2 are transmitted in respiratory droplets and aerosols, which are released during talking, breathing, coughing, and sneezing. Non-contact transmission of SARS-CoV-2 has been demonstrated, suggesting transmission in aerosols. Here we demonstrate that golden Syrian hamsters emit infectious SARS-CoV-2 in aerosols, prior to and concurrent with the onset of mild clinical signs of disease. The average emission rate is 25 infectious virions/hour on days 1 and 2 post-inoculation, with average viral RNA levels 200-fold higher than infectious virus in aerosols. Female hamsters have delayed kinetics of viral shedding in aerosols compared to male hamsters, with peak viral emission for females on dpi 2 and for males on dpi 1. The majority of virus is contained within aerosols <8 {micro}m in size. Thus, we provide direct evidence that, in hamsters, SARS-CoV-2 is an airborne virus.

category: microbiology


title: Pango lineage designation and assignment using SARS-CoV-2 spike gene nucleotide sequences

abstract: More than 2 million SARS-CoV-2 genome sequences have been generated and shared since the start of the COVID-19 pandemic and constitute a vital information source that informs outbreak control, disease surveillance, and public health policy. The Pango dynamic nomenclature is a popular system for classifying and naming genetically-distinct lineages of SARS-CoV-2, including variants of concern, and is based on the analysis of complete or near-complete virus genomes. However, for several reasons, nucleotide sequences may be generated that cover only the spike gene of SARS-CoV-2. It is therefore important to understand how much information about Pango lineage status is contained in spike-only nucleotide sequences. Here we explore how Pango lineages might be reliably designated and assigned to spike-only nucleotide sequences. We survey the genetic diversity of such sequences, and investigate the information they contain about Pango lineage status. Although many lineages, including the main variants of concern, can be identified clearly using spike-only sequences, some spike-only sequences are shared among tens or hundreds of Pango lineages. To facilitate the classification of SARS-CoV-2 lineages using subgenomic sequences we introduce the notion of designating such sequences to a "lineage set", which represents the range of Pango lineages that are consistent with the observed mutations in a given spike sequence. These data provide a foundation for the development of software tools that can assign newly-generated spike nucleotide sequences to Pango lineage sets.

category: genomics


title: Polysorbate 80 Controls Morphology, Structure and Stability of Human Insulin Amyloid-Like Spherulites

abstract: Amyloid protein aggregates are not only associated with neurodegenerative diseases and may also occur as unwanted by-products in protein-based therapeutics. Surfactants are often employed to stabilize protein formulations and reduce the risk of aggregation. However, surfactants alter protein-protein interactions and may thus modulate the physicochemical characteristics of any aggregates formed. Human insulin aggregation was induced at low pH in the presence of varying concentrations of the surfactant polysorbate 80. Various spectroscopic and imaging methods were used to study the aggregation kinetics, as well as structure and morphology of the formed aggregates. Molecular dynamics simulations were employed to investigate the initial interaction between the surfactant and insulin. Addition of polysorbate 80 slowed down, but did not prevent, aggregation of insulin. Amyloid spherulites formed under all conditions, with a higher content of intermolecular beta-sheets in the presence of the surfactant above its critical micelle concentration. In addition, a denser packing was observed, leading to a more stable aggregate. Molecular dynamics simulations suggested a tendency for insulin to form dimers in the presence of the surfactant, indicating a change in protein-protein interactions. It is thus shown that surfactants not only alter aggregation kinetics, but also affect physicochemical properties of any aggregates formed.


title: GATA4 and GATA6 loss-of-expression is associated with extinction of the classical programme and poor outcome in pancreatic ductal adenocarcinoma

abstract:
ObjectiveGATA6 is a master regulator of pancreatic differentiation and a key regulator of the classical phenotype in pancreatic ductal adenocarcinoma (PDAC). Low GATA6 expression is associated with poor patient outcome. GATA4 is the second most expressed GATA factor in the pancreas. The aim was to assess whether, and how, GATA4 contributes to PDAC phenotype and to analyze the association of expression with clinical outcome.

DesignWe analyzed PDAC transcriptomic data, stratifying cases according to GATA4 and GATA6 expression, and identified differentially expressed genes and pathways. A multicenter TMA study to assess GATA4 and GATA6 expression in PDAC samples (n=745) from patients undergoing tumour resection was performed using immunohistochemistry with antibodies of validated specificity. GATA4 and GATA6 levels were dichotomized into high/low categorical variables; association with outcome was assessed using univariable and multivariable Cox regression models.

ResultsSubtype classification using transcriptomic data revealed that GATA4 mRNA is enriched in classical, compared to basal-like tumours. We classified samples in 4 groups as high/low for GATA4 and GATA6. Reduced expression of GATA4 did not have a major transcriptional impact. However, concomitant low expression of GATA4 enhanced the transcriptomic effects of GATA6 low expression. Reduced expression of both proteins in tumours was associated with the worst patient survival. GATA4 and GATA6 expression significantly decreased in metastases and negatively correlated with basal markers.

ConclusionsOur analyses uncover a cooperative interaction between GATA4 and GATA6 to maintain the classical PDAC phenotype and provide compelling clinical rationale for assessing their expression as biomarkers of poor prognosis.

SUMMARY BOXO_ST_ABSWhat is already known about this subject?C_ST_ABS- Patients with classical-type PDAC have a better outcome
- Retrospective analyses suggest that classical-type PDAC is more sensitive to 5-FU-based chemotherapy
- GATA6 is a surrogate biomarker of classical tumours and its expression is associated with better survival
- GATA4 and GATA6 have overlapping and unique functions during pancreatic and gastrointestinal development

What are the new findings?- Tumours displaying only low GATA4 expression have a transcriptomic profile similar to those with preserved expression of both transcription factors
- Combined low expression of GATA4 and GATA6 has the highest transcriptomic impact
- In a large multicenter tissue microarray study, patients with tumours showing low expression of both GATA4 and GATA6 have the worst overall survival
- Low expression of GATA4 and GATA6 is an independent predictor of survival in patients with resectable PDAC
- GATA4 levels are down-regulated in liver metastases and are negatively correlated with basal markers such as KRT5/6, KRT14, and TP63

How might it impact on clinical practice in the foreseeable future?- The combined assessment of GATA4 and GATA6 expression may improve prognostic stratification of patients with PDAC
- Prospective studies should confirm whether GATA4 and GATA6 expression is also predictive of response to chemotherapy in PDAC patients

category: cancer biology


title: Role Of Efferocytosis In Oral Lichen Planus

abstract:
BackgroundOral lichen planus (OLP) is a chronic, immune mediated interface mucositis of oral mucosa. Though the apoptosis of keratinocytes is a feature of OLP, not much is known about the clearance of cell debris (efferocytosis) resulting from apoptosis. We postulate that there is a defective or delayed efferocytosis in OLP, which may have a role in modulating the immune response in OLP.

MethodsPublished mRNA expression of tissue of 14 patients with OLP and 14 cases of normal tissues were subjected to differential analysis (DE) and a list of DE genes identified. From this list, the genes that involved in efferocytosis were collated, compared and their interactions are typed.

ResultIn all, two studies fulfilled the inclusion and exclusion criteria. On combining the data, 1486 genes were significantly different between OLP and normal tissues. 28 of these 1486 genes are were associated with efferocytosis of which the suppression of LRP1, LDLR, ANAX2, C2, PBX1, PDCD4, S1PR5, CX3CL1, STAT6 and Wnt3A is indicative of defective or delayed efferocytosis in OLP. The role of pathways and associations were analyzed and is presented here.

Discussion and ConclusionThe study revealed that certain key genes mRNAs that are associated with efferocytosis are altered in OLP. They could delay or lead to defective efferocytosis. Studying such genes in detail could provide deeper understanding of the pathogenesis of the disease and the discovery of therapeutic targets.

category: pathology


title: Upregulation of C4 characteristics does not consistently improve photosynthetic performance in intraspecific hybrids of a grass

abstract: C4 photosynthesis is thought to have evolved via intermediate stages, with changes towards the C4 phenotype gradually enhancing photosynthetic performance. This hypothesis is widely supported by modelling studies, but experimental tests are missing. Mixing of C4 components to generate artificial intermediates can be achieved via crossing, and the grass Alloteropsis semialata represents an outstanding system since it includes C4 and non-C4 populations. Here, we analyse F1 hybrids between C3 and C4, and C3+C4 and C4 genotypes to determine whether the acquisition of C4 characteristics increases photosynthetic performance. The hybrids have leaf anatomical characters and C4 gene expression profiles that are largely intermediate between those of their parents. Carbon isotope ratios are similarly intermediate, which suggests that a partial C4 cycle coexists with C3 carbon fixation in the hybrids. This partial C4 phenotype is associated with C4-like photosynthetic efficiency in C3+C4 x C4, but not in C3 x C4 hybrids, which are overall less efficient than both parents. Our results support the hypothesis that the photosynthetic gains from the upregulation of C4 characteristics depend on coordinated changes in anatomy and biochemistry. The order of acquisition of C4 components is thus constrained, with C3+C4 species providing an essential step for C4 evolution.

category: plant biology


title: Aging-related olfactory loss is associated with olfactory stem cell transcriptional alterations in humans

abstract:
Presbyosmia, or aging related olfactory loss, occurs in a majority of humans over age 65 years, yet remains poorly understood, with no specific treatment options. The olfactory epithelium (OE) in the nasal fossa is the peripheral organ for olfaction, and is subject to acquired damage, suggesting a likely site of pathology in aging. OE basal stem cells reconstitute the neuroepithelium in response to cell loss under normal conditions. In aged OE, patches of respiratory-like metaplasia have been observed histologically, consistent with a failure in normal neuroepithelial homeostasis or repair. Accordingly, we have focused on identifying cellular and molecular changes in presbyosmic OE. Combining psychophysical testing with olfactory mucosa biopsy analysis, single cell RNA-sequencing (scRNA-seq), and human olfactory culture studies, we identified evidence for inflammation-associated changes in the OE stem cells of presbyosmic patients. The presbyosmic basal stem cells exhibited increased expression of genes involved in response to cytokines or stress, or the regulation of proliferation and differentiation. To facilitate further study of human OE stem cells, we developed an adult human basal cell culture model. Characterization of cultures using scRNA-seq confirmed maintenance of a reserve stem cell-like phenotype, and brief cytokine exposure in basal cell cultures resulted in increased expression of TP63, a transcription factor acting to prevent OE stem cell differentiation. Our data are consistent with a process by which aging-related inflammatory changes in OE stem cells may contribute to presbyosmia, via the disruption of normal epithelial homeostasis, suggesting that OE stem cells may represent a rational therapeutic target for restoration of olfaction.

One Sentence SummarySingle cell profiling suggests that inflammatory-associated olfactory epithelial stem cell dysfunction is associated with presbyosmia in humans.


title: Intracrine Vascular Endothelial Growth Factor Maintains Hippocampal Neural Stem Cell Quiescence

abstract: In the dentate gyrus (DG) of the adult mouse hippocampus, neural stem cells (NSCs) balance self-renewal and differentiation to produce neurons that support hippocampal function. Vascular endothelial growth factor (VEGF) is a well-known supporting factor for adult neurogenesis, but conflicting studies have left it uncertain how VEGF signals to NSCs. Here, we identified a VEGF-VEGFR2 intracrine signaling mechanism within adult DG NSCs that prevents their exhaustion. We show both in vitro and in vivo that NSC-VEGF loss caused cell-autonomous exhaustion of adult DG NSCs. In contrast, extracellular VEGF was neither necessary nor sufficient to maintain NSC quiescence or to stimulate VEGFR2 signaling, most likely due to sheddase-mediated cleavage of extracellular VEGFR2 ligand binding domains. Our findings support an exclusively intracellular mechanism for VEGF signaling in adult DG NSCs, thereby providing resolution to previously conflicting studies and suggesting cellular source can dictate the functional impact of soluble ligands in DG NSCs.


title: A systematic data-driven approach to analyze sensor-level EEG connectivity: Identifying robust phase-synchronized network components using surface Laplacian with spectral-spatial PCA

abstract: Although conventional averaging across predefined frequency bands reduces the complexity of EEG functional connectivity (FC), it obscures the identification of resting-state brain networks (RSN) and impedes accurate estimation of FC reliability. Extending prior work, we combined scalp current source density (CSD; spherical spline surface Laplacian) and spectral-spatial PCA to identify FC components. Phase-based FC was estimated via debiased weighted phase-locking index from CSD-transformed resting EEGs (71 sensors, 8 min, eyes open/closed, 35 healthy adults, 1-week retest). Spectral PCA extracted 6 robust alpha and theta factors (86.6% variance). Subsequent spatial PCA for each spectral factor revealed seven robust regionally-focused (posterior, central, frontal) and long-range (posterior-anterior) alpha components (peaks at 8, 10 and 13 Hz) and a midfrontal theta (6 Hz) component, accounting for 37.0% of FC variance. These spatial FC components were consistent with well-known networks (e.g., default mode, visual, sensorimotor), and four were sensitive to eyes open/closed conditions. Most FC components had good-to-excellent internal consistency (odd/even epochs, eyes open/closed) and test-retest reliability (ICCs [≥] .8). Moreover, the FC component structure was generally present in subsamples (session x odd/even epoch, or smaller subgroups [n = 7-10]), as indicated by similarity of factor loadings across PCA solutions. Apart from systematically reducing FC dimensionality, our approach avoids arbitrary thresholds and allows quantification of meaningful and reliable network components that may prove to be of high relevance for basic and clinical research applications.


title: Three-level Sleep Stage Classification Based on Wrist-worn Accelerometry Data Alone

abstract:
ObjectiveThe use of wearable sensor devices on daily basis to track real-time movements during wake and sleep has provided opportunities for automatic sleep quantification using such data. Existing algorithms for classifying sleep stages often require large training data and multiple input signals including heart rate and respiratory data. We aimed to examine the capability of classifying sleep stages using sensible features directly from accelerometers only with the aid of advanced recurrent neural networks.

Materials and MethodsWe analyzed a publicly available dataset with accelerometry data in 5s epoch length and polysomnography assessments. We developed long short-term memory (LSTM) models that take the 3-axis accelerations, angles, and temperatures from concurrent and historic observation windows to predict wake, REM and non-REM sleep. Leave-one-subject-out experiments were conducted to compare and evaluate the model performance with conventional nonsequential machine learning models using metrics such as multiclass training and testing accuracy, weighted precision, F1 score and area-under-the-curve (AUC).

ResultsOur sequential analysis framework outperforms traditional non-sequential models in all aspects of model evaluation metrics. We achieved an average of 65% and a maximum of 81% validation accuracy for classifying three sleep labels even with a relatively small training sample of clinical visitors. The presence of two additional derived variables, local variability and range, have shown to strongly improve the model performance.

DiscussionResults indicate that it is crucial to account for deep temporal dependency and assess local variability of the features. The post-hoc analysis of individual model performances on subjects demographic characteristics also suggest the need of including pathological samples in the training data in order to develop robust machine learning models that are capable of capturing normal and anomaly sleep patterns in the population.


title: EEG spectral exponent as a synthetic index for the longitudinal assessment of stroke recovery.

abstract:
BackgroundQuantitative EEG (qEEG) can capture changes in brain activity that follow a stroke. Accordingly, EEG metrics could be used to monitor patients state and recovery. Although qEEG metrics traditionally focus on oscillatory activity, recent findings highlight the importance of aperiodic (power-law) structure in characterizing pathological brain states.

ObjectiveTo assess neurophysiological impairment and recovery from mono-hemispheric stroke by means of the Spectral Exponent (SE), a metric that reflects EEG slowing and quantifies the power-law decay of the EEG Power Spectral Density (PSD). To relate neurophysiological recovery with patients functional outcome.

MethodsEighteen patients (n=18) with Middle Cerebral Artery (MCA) ischaemic stroke were retrospectively enrolled for this study. Patients underwent EEG recording in the sub-acute phase (T0) and after 2 months of physical rehabilitation (T1). Sixteen healthy controls (HC; n=16) matched by age and sex were enrolled as a normative group. SE values and narrow-band PSD were estimated for each recording. We compared SE and band-power between patients and HC, and between the affected (AH) and unaffected hemisphere (UH) at T0 and T1 in patients.

ResultsAt T0, stroke patients showed significantly more negative SE values than HC (p=0.003), reflecting broad-band EEG slowing. Moreover, SE over the AH was consistently more negative compared to the UH and showed a renormalization at T1 in our patient sample. This SE renormalization significantly correlated with NIHSS improvement (R= 0.63, p=0.005).

ConclusionsSE is a reliable readout of the electric changes occurring in the brain after an ischaemic cortical lesion. Moreover, SE holds the promise to be a robust method to assess stroke impairment as well as to monitor and predict functional outcome.


title: Repair of Noise-Induced Damage to Stereocilia F-actin Cores is Facilitated by XIRP2

abstract: Prolonged exposure to loud noise has been shown to affect inner ear sensory hair cells in a variety of deleterious manners, including damaging the stereocilia core. The damaged sites can be visualized as "gaps" in phalloidin staining of F-actin, and the enrichment of monomeric actin at these sites, along with an actin nucleator and crosslinker, suggests that localized remodeling occurs to repair the broken filaments. Herein we show that gaps in mouse auditory hair cells are largely repaired within one week of traumatic noise exposure through the incorporation of newly synthesized actin. Additionally, we report that XIRP2 is required for the repair process and facilitates the enrichment of monomeric{gamma} -actin at gaps through its LIM domain-containing C-terminus. Our study describes a novel process by which hair cells can recover from sub-lethal hair bundle damage and which may contribute to recovery from temporary hearing threshold shifts and the prevention of age-related hearing loss.


title: Localized hypermutation drives the evolution of unstable colistin resistance in small populations of Pseudomonas aeruginosa

abstract: Colistin has emerged as an important last line of defence for the treatment of infections caused by antibiotic resistant Gram-negative pathogens. Here we investigate the responses of {approx}1,000 populations of an MDR strain of P. aeruginosa to a high dose of colistin. Colistin exposure resulted in rapid cell death, but a sub-set of populations eventually recovered due to the outgrowth of heteroresistant cells. Genome sequencing revealed that heteroresistance was primarily driven by mutations in the PmrAB two-component system that occurred at a rate ({approx}2x10-5 per cell division) that was 103-104 fold higher than typical resistance mutation rates. Crucially, this elevated mutation rate was only found in pmrB, demonstrating that hypermutability is localized to this gene. PmrAB provides resistance to antimicrobial peptides that are involved in host immunity, suggesting that this pathogen may have evolved a high mutation rate as an adaption to generate mutants that are resistant to host antimicrobial peptides that are secreted during infection. Interestingly, we found no mutations in 1/3 of populations that recovered from colistin treatment, suggesting that phenotypic plasticity and/or persister cells contribute to the ability of Pseudomonas to adapt to colistin.


title: Dysregulation of Hepatitis B Virus Nucleocapsid Assembly with RNA-directed Small Ligands

abstract: RNA sequences/motifs dispersed across the pre-genomic copy of the Hepatitis B Virus genome regulate formation of nucleocapsids in vitro in an epsilon/polymerase independent fashion. These multiple RNA Packaging Signals (PSs) form stem-loops presenting in each loop a core protein recognition motif, -RGAG-. Small, drug-like molecules binding these motifs were identified by screening an immobilized library with a fluorescently-labelled RNA oligonucleotide encompassing the most conserved of these sites. This identified 66 "hits", with affinities ranging from low nanomolar to high micromolar in SPR assays. High affinity ligand binding is dependent on the presence of the -RGAG-motif, which also appears to be the common element in cross-binding to other PS sites. Some of these compounds are potent inhibitors of in vitro core protein assembly around the HBV pre-genome. Mathematical modelling confirms the potential of these novel anti-viral drug targets for disrupting replication of this major human pathogen. Preliminary structure-activity relationships of the highest affinity compound reveal critical functional groups for PS-binding. PS-regulated assembly is easily adapted to high-throughput screening allowing future development of pharmacologically active compounds.


title: An ultrapotent neutralizing bispecific antibody with broad spectrum against SARS-CoV-2 variants

abstract: In spite of the successful development of effective countermeasures against Covid-19, variants have and will continue to emerge that could compromise the efficacy of currently approved neutralizing antibodies and vaccines. Consequently, novel and more efficacious agents are urgently needed. We have developed a bispecific antibody, 2022, consisting of two antibodies, 2F8 and VHH18. 2F8 was isolated from our proprietary fully synthetic human IDEAL (Intelligently Designed and Engineered Antibody Library)-VH/VL library and VHH18 is a single domain antibody isolated from IDEAL-nanobody library. 2022 was constructed by attaching VHH18 to the C-terminal of Fc of 2F8. 2022 binds two non-overlapping epitopes simultaneously on the RBD of the SARS-CoV-2 spike protein and blocks the binding of RBD to human angiotensin-converting enzyme 2 (ACE2). 2022 potently neutralizes SARS-CoV-2 and all of the variants tested in both pseudovirus and live virus assays, including variants carrying mutations known to resist neutralizing antibodies approved under EUA and that reduce the protection efficiency of current effective vaccines. The half-maximum inhibitory concentration (IC50) of 2022 is 270 pM, 30 pM, 20 pM, and 1 pM, for wild-type, alpha, beta, and delta pseudovirus, respectively. In the live virus assay, 2022 has an IC50 of 26.4 pM, 13.3 pM, and 88.6 pM, for wild-type, beta, and delta live virus, respectively. In a mouse model of SARS-CoV-2, 2022 showed strong prophylactic and therapeutic effects. A single administration of 2022 intranasal (i.n.) or intraperitoneal (i.p.) 24 hours before virus challenge completely protected all mice from bodyweight loss, as compared with up to 20% loss of bodyweight in placebo treated mice. In addition, the lung viral titers were undetectable (FRNT assay) in all mice treated with 2022 either prophylactically or therapeutically, as compared with around 1x105 pfu/g lung tissue in placebo treated mice. In summary, bispecific antibody 2022 showed potent binding and neutralizing activity across a variety of SARS-CoV-2 variants and could be an attractive weapon to combat the ongoing waves of the COVID-19 pandemic propagated mainly by variants, especially, the much more contagious delta variant.


title: TSC2-extracellular matrix crosstalk controls pulmonary vascular proliferation and pulmonary hypertension

abstract:
Increased proliferation and survival of resident cells in small pulmonary arteries (PA) are important drivers of pulmonary hypertension (PH). Tuberous sclerosis complex 2 (TSC2) is a negative regulator of mTOR complex 1 and cell growth. Here we show that TSC2 is deficient in small remodeled PA/PA vascular smooth muscle cells (PAVSMC) from human PAH and experimental PH lungs. TSC2 deficiency was reproduced in vitro by maintaining PAVSMC on pathologically stiff substrates and was required for stiffness-induced proliferation, accumulation of transcriptional co-activators YAP/TAZ and up-regulation of mTOR. Depletion of TSC2 reproduced PH features in vitro in human PAVSMC and in vivo in SM22-Tsc2+/- mice. TSC2 loss in PAVSMC was supported by YAP and led to the up-regulation of YAP/TAZ and mTOR via modulating the extracellular matrix (ECM) composition. ECM, produced by TSC2-deficient PAVSMC, promoted growth of non-diseased PA adventitial fibroblasts and PAVSMC, which, in turn, was prevented by 5{beta}1 integrin receptor antagonist ATN161. In vitro, molecular and pharmacological (SRT2104) restoration of TSC2 down-regulated YAP/TAZ, mTOR, and ECM production, inhibited proliferation and induced apoptosis in human PAH PAVSMC. In vivo, orally administrated SRT2104 restored TSC2, resolved pulmonary vascular remodeling, PH, and improved right heart in two rodent models of PH. Thus, PAVSMC TSC2 is a critical integrator of ECM composition and stiffness with pro-proliferative signaling and PH, and the restoration of functional TSC2 could be an attractive therapeutic option to treat PH.

One Sentence SummaryTSC2 acts as mechanosensor and mechanotransducer, integrating ECM composition and stiffness with pro-proliferative signaling in pulmonary vasculature; its deficiency in PA vascular smooth muscle cells results in ECM remodeling, hyper-proliferation and pulmonary hypertension, which could be reversed by pharmacological restoration of functional TSC2.


title: Modeling myosin Va liposome transport through actin filament networks reveals a percolation threshold that modulates transport properties

abstract: Myosin Va (myoVa) motors transport membrane-bound cargo through three-dimensional, intracellular actin filament networks. We developed a coarse-grained, in silico model to predict how actin filament density (3-800 filaments) within a randomly oriented actin network affects fluid-like liposome (350nm vs. 1,750nm) transport by myoVa motors. 5,000 simulated liposomes transported within each network adopted one of three states: transport, tug of war, or diffusion. Diffusion due to liposome detachment from actin rarely occurred given at least 10 motors on the liposome surface. However, with increased actin density, liposomes transitioned from primarily directed transport on single actin filaments to an apparent random walk, resulting from a mixture of transport and tug of wars as the probability of encountering additional actin filaments increased. This phase transition arises from a percolation phase transition at a critical number of accessible actin filaments, Nc. Nc, is a geometric property of the actin network that depends only on the position and polarity of the actin filaments and the liposome diameter, as evidenced by a five-fold increase in liposome diameter resulting in a five-fold decrease in Nc. Thus, in cells, actin network density and cargo size may be regulated to match cargo delivery to the cells physiological demands.


title: Concentration and quantification of Tilapia tilapinevirus from water using a simple iron flocculation coupled with probe-based RT-qPCR

abstract: Tilapia tilapinevirus (also known as tilapia lake virus, TiLV) is an important virus responsible for die-off of farmed tilapia globally. Detection and quantification of the virus from environmental DNA/RNA (eDNA/eRNA) using pond water represents a potential, noninvasive routine approach for pathogen monitoring and early disease forecasting in aquaculture systems. Here, we report a simple iron flocculation method for viral concentration from water combined with a newly developed hydrolysis probe quantitative RT-qPCR method for detection and quantification of TiLV. The RT-qPCR method targeting a conserved region of TiLV genome segment 9 has a detection limit of 10 viral copies per {micro}L of template. The method had a 100% analytical specificity and sensitivity for TiLV. The optimized iron flocculation method was able to recover 16.11 {+/-} 3.3% of virus from water samples spiked with viral cultures. During disease outbreak cases from an open-caged system and a closed hatchery system, both tilapia and water samples were collected for detection and quantification of TiLV. The results revealed that TiLV was detected from both clinically sick fish and asymptomatic fish. Most importantly, the virus was successfully detected from water samples collected from different locations in the affected farms e.g. river water samples from affected cages (8.50 x 102 to 2.79 x 104 copies/L) and fish-rearing water samples, sewage, and reservoir (4.29 x 102 to 3.53 x 103 copies/L) from affected and unaffected ponds of the hatchery. In summary, this study suggests that the eRNA detection system using iron flocculation coupled with probe based-RT-qPCR is feasible for concentration and quantification of TiLV from water. This approach might be useful for noninvasive monitoring of TiLV in tilapia aquaculture systems and facilitating appropriate decisions on biosecurity interventions needed.


title: RNA X-ray footprinting reveals the consequences of an in vivo acquired determinant of viral infectivity

abstract: The secondary structures of the bacteriophage MS2 ssRNA genome, frozen in defined states, were determined with minimal perturbation using constraints from X-ray synchrotron footprinting (XRF). The footprints of the gRNA in the virion and as transcript are consistent with single, dominant but distinct conformations, and reveal the presence of multiple Packaging Signals potentially involved in assembly regulation that have not been detected by other techniques. XRF also reveals the dramatic effect of the unique Maturation Protein (MP) on both the capsid lattice, and the gRNA conformation inside the phage compared with a virus-like-particle composed only of coat protein subunits. Aspects of genome organisation in the phage, their impacts on the capsid shell, and the distortion of lattice geometry by MP, are hallmarks of molecular frustration. Phage assembly therefore appears to prepare the particle for the next step of the infectious cycle.


title: Stress-dependent inhibition of cell polarity through unbalancing the GEF/GAP regulation of Cdc42

abstract: Cdc42 rules cell polarity and growth in fission yeast. It is negatively and positively regulated by GTPase-activating proteins (GAPs) and by Guanine nucleotide Exchange factors (GEFs), respectively. Active Cdc42-GTP localizes to the poles, where it associates with numerous proteins constituting the polarity module. However, little is known about its down-regulation. We describe here that oxidative stress causes Sty1 kinase-dependent Cdc42 inactivation at cell poles. Both the amount of active Cdc42 at poles and cell length inversely correlate with Sty1 activity, explaining the elongated morphology of{Delta} sty1 cells. We have created stress-blinded cell poles by either eliminating two Cdc42 GAPs or through the constitutive tethering of a GEF to the cell tips, and biochemically demonstrate that Rga3 is a direct substrate of Sty1. We propose that stress-activated Sty1 promotes GTP hydrolysis and prevents GEF activity at the cell tips, thus leading to the inhibition of Cdc42 and polarized growth cessation.

category: cell biology


title: Lung injury induces alveolar type 2 cell hypertrophy and polyploidy with implications for repair and regeneration

abstract: Epithelial polyploidization post-injury is a conserved phenomenon, recently shown to improve barrier restoration during wound healing. Whether lung injury can induce alveolar epithelial polyploidy is not known. We show that bleomycin injury induces AT2 cell hypertrophy and polyploidy. AT2 polyploidization is also seen in short term ex vivo cultures, where AT2-to-AT1 trans-differentiation is associated with substantial binucleation due to failed cytokinesis. Both hypertrophic and polyploid features of AT2 cells can be attenuated by inhibiting the integrated stress response (ISR) using the small molecule ISRIB. These data suggest that AT2 hypertrophic growth and polyploidization may be a feature of alveolar epithelial injury. As AT2 cells serve as facultative progenitors for the distal lung epithelium, a propensity for injury-induced binucleation has implications for AT2 self-renewal and regenerative potential upon reinjury, which may benefit from targeting the ISR.


title: High Leaf Respiration Rates May Limit the Success of White Spruce Saplings growing in The Kampfzone at the Arctic Treeline

abstract:
Arctic Treeline is the transition from the boreal forest to the treeless tundra and may be determined by growing season temperatures. The physiological mechanisms involved in determining the relationship between the physical and biological environment and the location of treeline are not fully understood. In Northern Alaska we studied the relationship between temperature and leaf respiration in 36 white spruce (Picea glauca) trees, sampling both the upper and lower canopy, to test two research hypotheses (H0). The first H01 is that canopy position will not influence leaf respiration. The associated alternative hypothesis (HA) is that the upper canopy leaves which are more directly coupled to the atmosphere will experience more challenging environmental conditions and thus have higher respiration rates to facilitate metabolic function. The second H02 is that tree size will not influence leaf respiration. The associated HA is that saplings (stems that are 5-10 cm DBH (diameter at breast height)) will have higher respiration rates than trees (stems [≥] 10 cm DBH) since saplings represent the transition from seedlings growing in the more favorable aerodynamic boundary layer, to trees which are fully coupled to the atmosphere but of sufficient size to persist. Respiration did not change with canopy position, however respiration at 25{degrees}C was 42% higher in saplings compared to trees (3.43 {+/-} 0.19 vs. 2.41 {+/-} 0.14 mol m-2 s-1). Furthermore, there were significant differences in the temperature response of respiration, and seedlings reached their maximum respiration rates at 59{degrees}C, more than two degrees higher than trees. Our results demonstrate that the respiratory characteristics of white spruce saplings at treeline are extreme, imposing a significant carbon cost that may contribute to their lack of perseverance beyond treeline. In the absence of thermal acclimation, the rate of leaf respiration could increase by 57% by the end of the century, posing further challenges to the ecology of this massive ecotone.

This paper is dedicated to the memory of James N. Siedow, Professor of Botany at Duke University. I am honored to have learned from, and to have been inspired by Jim. I will always be grateful for the time he spent helping me, the depth of education he gave me, and his steady mentoring as part of my thesis committee. His devotion to science and push for deeper knowledge of plant respiration set an example for all of us, particularly those who were lucky enough to study with him. Jim was also a memorable Father Christmas at the departmental holiday parties, always keeping us laughing with his particular brand of quick wit and sarcasm. - Kevin L. Griffin

category: ecology


title: Nucleated Fish Erythrocyte Extracellular Traps (FEETs) release is an evolutionary conserved immune defence process

abstract: Fish erythrocytes remain nucleated for their life-span, unlike mammalian erythrocytes which undergo enucleation. Asides transportation of oxygen, fish erythrocytes are capable of several immune defence processes. Nucleated fish erythrocytes represent prime candidates for carrying out ETotic responses. ETosis is an evolutionary conserved innate immune defence process found in both vertebrates and invertebrates, which involves the extrusion of DNA studded with antimicrobial proteins into the extracellular space serving to trap and kill microorganisms. In this report, we demonstrate that fish erythrocytes isolated from Danio rerio (zebrafish) produce ETotic-like responses when exposed to chemical and physiological stimuli. Furthermore, we found Salmo salar (Atlantic salmon) erythrocytes produce similar ETotic responses. We have termed these ET-like formations Fish Erythrocyte Extracellular Traps (FEETs). Interestingly, we discovered that mammalian inducers of NETosis, such as the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate and the calcium ionophore ionomycin, induced FEETs. Moreover, we found that FEETs are dependent upon activation of PKC and generation of mitochondrial reactive oxygen species. Thus, this brief report represents the first demonstration that fish erythrocytes can exhibit ETotic-like responses, unveiling a previously unknown function of nucleated erythrocytes, which sheds new light on the innate immune arsenal of erythrocytes.

category: evolutionary biology


title: LRcell: detecting the source of differential expression at the sub-cell type level from bulk RNA-seq data

abstract: Given most tissues are consist of abundant and diverse sub cell-types, an important yet unaddressed problem in bulk RNA-seq analysis is to identify at which sub cell-type(s) the differential expression occur. Single-cell RNA-sequencing (scRNA-seq) technologies can answer the question, but they are often labor-intensive and cost-prohibitive. Here, we present LRcell, a computational method aiming to identify specific sub-cell type(s) that drives the changes observed in a bulk RNA-seq experiment. In addition, LRcell provides pre-embedded marker genes computed from putative single-cell RNA-seq experiments as options to execute the analyses. Using three different real datasets, we show that LRcell successfully identifies known cell types involved in psychiatric disorders and LRcell is more sensitive than even the leading deconvolution methods.

category: bioinformatics


title: Structural basis for context-specific inhibition of translation by oxazolidinone antibiotics

abstract: The antibiotic linezolid, the first clinically approved member of the oxazolidinone class, inhibits translation of bacterial ribosomes by binding to the peptidyl transferase center. Recent work has demonstrated that linezolid does not inhibit peptide bond formation at all sequences but rather acts in a context-specific manner, namely when alanine occupies the penultimate position of the nascent chain. In this study, we determined that the second-generation oxazolidinone radezolid also induces stalling with alanine at the penultimate position. However, the molecular basis for context-specificity of these inhibitors has not been elucidated. In this study, we determined high-resolution cryo-EM structures of both linezolid and radezolid-stalled ribosome complexes. These structures reveal that the alanine side chain fits within a small hydrophobic crevice created by oxazolidinone, resulting in improved ribosome binding. Modification of the ribosome by the antibiotic resistance enzyme Cfr disrupts stalling by forcing the antibiotic to adopt a conformation that narrows the hydrophobic alanine pocket. Together, the structural and biochemical findings presented in this work provide molecular understanding of context-specific inhibition of translation by clinically important oxazolidinone antibiotics.


title: Light-sheet photonic force optical coherence elastography for high-throughput quantitative 3D micromechanical imaging

abstract: Microscale mechanical properties of the extracellular matrix (ECM) and dynamic cell-ECM interactions play an important role in physiological processes and disease. However, it remains a challenge for current mechanical characterization methods to combine quantitative 3D imaging of ECM mechanics with cellular-scale resolution and dynamic monitoring of cell-mediated changes to pericellular viscoelasticity. Here, we present light-sheet photonic force optical coherence elastography (LS-pfOCE) to address these challenges by leveraging a light-sheet for parallelized, non-invasive, and localized mechanical loading. We demonstrate the capabilities of LS-pfOCE by imaging the micromechanical heterogeneity of fibrous 3D collagen matrices and perform a live-cell study to image micromechanical heterogeneity induced by NIH-3T3 cells seeded in 3D fibrin constructs. We also show that LS-pfOCE is able to quantify temporal variations in pericellular viscoelasticity in response to altered cellular activity. By providing access to the spatiotemporal variations in the micromechanical properties of 3D complex biopolymer constructs and engineered cellular systems, LS-pfOCE has the potential to drive new discoveries in the rapidly growing field of mechanobiology.


title: Molecular Understanding of Calorimetric Protein Unfolding Experiments

abstract: Protein unfolding is a dynamic cooperative equilibrium between short lived protein conformations. The Zimm-Bragg theory is an ideal algorithm to handle cooperative processes. Here, we extend the analytical capabilities of the Zimm-Bragg theory in two directions. First, we combine the Zimm-Bragg partition function Z(T) with statistical-mechanical thermodynamics, explaining the thermodynamic system properties enthalpy, entropy and free energy with molecular parameters only. Second, the molecular enthalpy h0 to unfold a single amino acid residue is made temperature-dependent. The addition of a heat capacity term cv allows predicting not only heat denaturation, but also cold denaturation. Moreover, it predicts the heat capacity increase [Formula] in protein unfolding. The theory is successfully applied to differential scanning calorimetry experiments of proteins of different size and structure, that is, gpW62 (62aa), ubiquitin (74aa), lysozyme (129aa), metmyoglobin (153aa) and mAb monoclonal antibody (1290aa). Particular attention was given to the free energy, which can easily be obtained from the heat capacity Cp(T). The DSC experiments reveal a zero free energy for the native protein with an immediate decrease to negative free energies upon cold and heat denaturation. This trapezoidal shape is precisely reproduced by the Zimm-Bragg theory, whereas the so far applied non-cooperative 2-state model predicts a parabolic shape with a positive free energy maximum of the native protein. We demonstrate that the molecular parameters of the Zimm-Bragg theory have a well-defined physical meaning. In addition to predicting protein stability, independent of protein size, they yield estimates of unfolding kinetics and can be connected to molecular dynamics calculations.


title: Unusually slow spike frequency adaptation in deep cerebellar nuclei neurons preserves linear transformations on the sub-second time scale

abstract: Purkinje cells (PCs) are spontaneously active neurons of the cerebellar cortex that inhibit glutamatergic projection neurons within the deep cerebellar nuclei (DCN) that in turn provide the primary cerebellar output. Brief reductions in PC firing rapidly increase DCN neuron firing. However, prolonged reductions in PC inhibition, as seen in some disease states, certain types of transgenic mice, and in acute slices of the cerebellum, do not evoke large sustained increases in DCN firing. Here we test whether there is a mechanism of spike-frequency adaptation in DCN neurons that could account for these properties. We find that prolonged optogenetic suppression of PC synapses in vivo transiently elevates PC firing that strongly adapts within ten seconds. We perform current-clamp recordings at near physiological temperature in acute brain slices to examine how DCN neurons respond to prolonged depolarizations. Adaptation in DCN neurons is exceptionally slow and bidirectional. A depolarizing current step evokes large initial increases in firing that decay to less than 20% of the initial increase within approximately ten seconds. Such slow adaptation could allow DCN neurons to adapt to prolonged changes in PC firing while maintaining their linear firing frequency-current relationship on subsecond time scales.


title: Neural network-based predictions of antimicrobial resistance in Salmonella spp. using k-mers counting from whole-genome sequences

abstract: AO_SCPLOWBSTRACTC_SCPLOWArtificial intelligence-based predictions have emerged as a friendly and reliable tool for the surveillance of the antimicrobial resistance (AMR) worldwide. In this regard, genome databases typically include whole-genome sequencing (WGS) data containing AMR meta-data that can be used to train machine learning (ML) models, in order to predict phenotype features from genome samples. In this study, using a Neural Network (NN) architecture and the SGD-ADAM algorithm, we build ML antibiotic resistance models that can predict Minimum Inhibitory Concentrations (MICs) and antimicrobial susceptibility profiles of Salmonella spp. Data analysis was based on 7,268 genomes publicly available in PATRIC database, containing about 75,000 AMR annotations. ML models were built using reference-free k-mer analysis of whole-genome sequences, MIC measurements and susceptibility categories, obtaining robust and accurate results for 9 antibiotics belonging to beta-lactam, fluoroquinolone, phenicol, aminoglycoside, tetracycline and sulphonamide classes. Al-though the accuracy of predicting the actual MIC reaches modest levels, the within {+/-} 1 2-fold dilution accuracy per antibiotic reaches significant levels with values that varies from 85% to 95%, with narrow 95% CIs of about 5% and individual accuracies per MIC {gtrsim} 80%. For differentiation between "susceptible" and "resistant" values, by measuring the accuracy and error of models susceptibility predictions to different antibiotics, the accuracy is the same as before and ranges from 85% to 95%, with 95% CIs of about 5%, the recall extends from 75% to 85%, the precision from 60% to 90%, whereas the very major error is [lsim] 20%. In summary, these results show that NN-based models are able to learn and predict the AMR phenotype from bacterial genomes based on a gene-free k-mer analysis.


title: Quality-preserving low-cost probabilistic 3D denoising with applications to Computed Tomography

abstract:
We propose a pipeline for a synthetic generation of personalized Computer Tomography (CT) images, with a radiation exposure evaluation and a lifetime attributable risk (LAR) assessment. We perform a patient-specific performance evaluation for a broad range of denoising algorithms (including the most popular Deep Learning denoising approaches, wavelets-based methods, methods based on Mumford-Shah denoising etc.), focusing both on accessing the capability to reduce the patient-specific CT-induced LAR and on computational cost scalability. We introduce a parallel probabilistic Mumford-Shah denoising model (PMS), showing that it markedly-outperforms the compared common denoising methods in denoising quality and cost scaling. In particular, we show that it allows an approximately 22-fold robust patient-specific LAR reduction for infants and a 10-fold LAR reduction for adults. Using a normal laptop the proposed algorithm for PMS allows a cheap and robust (with the Multiscale Structural Similartity index > 90%) denoising of very large 2D videos and 3D images (with over 107 voxels) that are subject to ultra-strong Gaussian and various non-Gaussian noises, also for Signal-to-Noise Ratios much below 1.0. The code is provided for open access.

One-sentence summaryProbabilisitc formulation of Mumford-Shah principle (PMS) allows a cheap quality-preserving denoising of ultra-noisy 3D images and 2D videos.


title: Natural reassortment of a segmented RNA arbovirus illustrates plasticity of phenotype in the arthropod vector and mammalian host in vivo

abstract:
Segmented RNA viruses are a taxonomically diverse group of 11 families that can infect plant, wildlife, livestock and human hosts. A shared feature of these viruses is the ability to exchange genome segments during co-infection of a host by a process termed  reassortment. Reassortment enables rapid evolutionary change, but in the case of segmented RNA viruses utilising an arthropod vector is set against the constraint of purifying selection and genetic bottlenecks imposed by replication in two evolutionarily distant hosts. In this study, we use an in vivo host: arbovirus: vector model to investigate the impact of reassortment on two phenotypic traits: vector competence and virulence in the host. Bluetongue virus (BTV) (Reoviridae) is the causative agent of bluetongue (BT), an economically important disease of domestic and wild ruminants and deer. The genome of BTV is comprised of 10 linear segments of dsRNA and the virus is transmitted between ruminants by Culicoides biting midges (Diptera: Ceratopogonidae). Five strains of BTV representing three serotypes (BTV-1, BTV-4 and BTV-8) were isolated from naturally infected ruminants in Europe and parental/reassortant lineage status assigned through full genome sequencing. Each strain was then assessed in parallel for the ability to infect Culicoides and to cause BT in sheep. Our results demonstrate that two reassortment strains, which themselves became established in the field, had obtained high replication ability in C. sonorensis from one of the parental virus strains which allowed inferences of the genome segments conferring this phenotypic trait.

IMPORTANCEReassortment between strains can lead to major shifts in the transmission parameters and virulence of segmented RNA viruses with consequences for spread, persistence and impact. The ability of these pathogens to change their phenotypes rapidly in response to selection pressure in new environments presents a major challenge in understanding factors driving emergence. Utilising a natural mammalian host-insect vector infection and transmission model, we demonstrated for the first time the genetic basis for a phenotypic trait of BTV within strains directly isolated from the field and, hence, selected and relevant for natural transmission.


title: Artemisinin-independent inhibitory activity of Artemisia sp. infusions against different Plasmodium stages including relapse-causing hypnozoites

abstract: Artemisinin-based combination therapies (ACT) are the frontline treatments against malaria worldwide. Recently the use of traditional infusions from Artemisia annua (from which artemisinin is obtained) or A. afra (lacking artemisinin) has been controversially advocated. Such unregulated plant-based remedies are strongly discouraged as they might constitute sub-optimal therapies and promote drug resistance. Here, we conducted the first comparative study of the anti-malarial effects of both plant infusions in vitro against the asexual erythrocytic stages of P. falciparum and the pre-erythrocytic (i. e., liver) stages of various Plasmodium species. Low concentrations of either infusion accounted for significant inhibitory activities across every parasite species and stage studied. We show that these antiplasmodial effects were essentially artemisinin-independent and were additionally monitored by observations of the parasite apicoplast and mitochondrion. In particular, the infusions significantly incapacitated sporozoites, and for P. vivax and P. cynomolgi, disrupted the hypnozoites. This provides the first indication that compounds other than 8-aminoquinolines could be effective antimalarials against relapsing parasites. These observations advocate for further screening to uncover urgently needed novel antimalarial lead compounds.


title: MAP4K4 determines cancer cell phenotype by controlling the plasma membrane-associated proteome

abstract:
The composition of the plasma membrane (PM)-associated proteome of tumor cells determines cell-cell and cell-matrix interactions and the response to environmental cues. Whether the PM-associated proteome impacts the phenotype of Medulloblastoma (MB) tumor cells and how it adapts in response to growth factor cues is poorly understood. Using a spatial proteomics approach, we observed that hepatocyte growth factor (HGF)-induced activation of the receptor tyrosine kinase c-MET in MB cells changes the abundance of transmembrane and membrane-associated proteins. The depletion of MAP4K4, a pro-migratory effector kinase downstream of c-MET, leads to a specific decrease of the adhesion and immunomodulatory receptor CD155 and of components of the fast-endophilin-mediated endocytosis (FEME) machinery in the PM-associated proteome of HGF-activated MB cells. The decreased surface expression of CD155 or of the FEME effector Endophilin A1 reduces growth and invasiveness of MB tumor cells in the tissue context. These data thus describe a novel function of MAP4K4 in the control of the PM-associated proteome of tumor cells and identified two downstream effector mechanisms controlling proliferation and invasiveness of MB cells.

Graphical abstractc-MET activation upon HGF stimulation induces c-MET internalization and induces downstream MAP4K4 activity. (1) MAP4K4 is required downstream of activated c-MET for the maintenance of surface presentation of CD155 in activated cells. CD155 expression is required for MB cell migration, invasion and proliferation in the tissue context. (2) MAP4K4 is required downstream of activated c-MET to maintain membrane depolarization, possibly by regulating the surface localization of several ion channels and transporters. (3) MAP4K4 is required downstream of activated c-MET cause PM-proximal localization of FEME effector CIP4, FBP17 and CIN85. The FEME effector endophilin A is necessary for MB cell migration, invasion and dissemination.


title: CD155 and EndoA1 mediate growth and tissue invasion downstream of MAP4K4 in medulloblastoma cells


title: Evolutionary divergence of anaphase spindle mechanics in nematode embryos constrained by antagnonistic pulling and viscous forces

abstract: Cells are the basic unit of biological organization, and their division is remarkably conserved across phyla. However from an evolutionary perspective, it remains unclear how much cellular parameters can diverge, without altering the basic function they sustain. We address the mechanics of asymmetric mitotic spindle positioning during the first embryonic division of six nematode species. We propose a viscoelastic model of spindle positioning and mobility that can provide a physical explanation of why in C. elegans it undergoes oscillations during elongation, whereas most others lack oscillations. To test this model, we measured the pulling forces and opposing cytoplasmic drag by a combination of laser ablation of the anaphase spindle and tracking of intracellular granules. While centrosomes of all species recoil on spindle cutting, quantitative differences correlate with the cytoplasmic viscosity. In fact, increased viscosity correlates with decreased oscillation speeds of intact spindles across species. However, the absence of oscillations despite low viscosity in some species, can only be explained by smaller pulling forces. Consequently, we find that spindle mobility across the species analyzed here is characterized by a tradeoff between cytoplasmic viscosity and pulling forces. Our work provides a framework for understanding mechanical constraints on evolutionary diversification of spindle mobility.


title: Caveats and nuances of model-based and model-free representational connectivity analysis

abstract: Brain connectivity analyses have conventionally relied on statistical relationship between one-dimensional summaries of activation in different brain areas. However, summarising activation patterns within each area to a single dimension ignores the potential statistical dependencies between their multi-dimensional activity patterns. Representational Connectivity Analyses (RCA) is a method that quantifies the relationship between multi-dimensional patterns of activity without reducing the dimensionality of the data. We consider two variants of RCA. In model-free RCA, the goal is to quantify the shared information for two brain regions. In model-based RCA, one tests whether two regions have shared information about a specific aspect of the stimuli/task, as defined by a model. However, this is a new approach and the potential caveats of model-free and model-based RCA are still understudied. We first explain how model-based RCA detects connectivity through the lens of models, and then present three scenarios where model-based and model-free RCA give discrepant results. These conflicting results complicate the interpretation of functional connectivity. We highlight the challenges in three scenarios: complex intermediate models, common patterns across regions and transformation of representational structure across brain regions. The paper is accompanied by scripts that reproduce the results. In each case, we suggest potential ways to mitigate the difficulties caused by inconsistent results. The results of this study shed light on some understudied aspects of RCA, and allow researchers to use the method more effectively.


title: Anterior Cingulate Cortex Signals the Need to Control Intrusive Thoughts During Motivated Forgetting

abstract: How do people limit awareness of unwanted memories? Evidence suggests that when unwelcome memories intrude, a retrieval stopping process engages the right dorsolateral prefrontal cortex (rDLPFC; Anderson et al., 2004) to inhibit hippocampal activity (Benoit and Anderson, 2012; Benoit et al., 2015; Gagnepain et al., 2017) and disrupt retrieval. It remains unknown how and when the need to engage prefrontal control is detected, and whether control operates proactively to prevent an unwelcome memory from being retrieved, or must respond reactively, to counteract its intrusion. We hypothesized that dorsal anterior cingulate cortex (dACC) achieves this function by detecting signals indicating that an unwanted trace is emerging in awareness, and transmitting the need for inhibitory control to right DLPFC (Alexander and Brown, 2011; Botvinick et al., 2001). During a memory suppression task, we measured trial-by-trial variations in dACCs theta power and N2 amplitude, two electroencephalographic (EEG) markers of the need for enhanced control (Cavanagh and Frank, 2014). With simultaneous EEG-fMRI recordings, we tracked dynamic interactions between the dACC, rDLPFC and hippocampus during suppression. EEG analyses revealed a clear role of dACC in detecting the need for memory control, and in upregulating prefrontal inhibition. Importantly, we identified dACC contributions before episodic retrieval could have occurred (500 ms) and afterwards, indicating distinct proactive and reactive control signalling. Stronger proactive control by the dACC led to reduced hippocampal activity and diminished overall blood-oxygen-level-dependent (BOLD) signal in dACC and rDLPFC, suggesting that pre-empting retrieval early reduced overall control demands. However, when dACC activity followed the likely onset of recollection, retrieval was cancelled reactively: effective connectivity analyses revealed robust communication from dACC to rDLPFC and from rDLPFC to hippocampus, tied to successful forgetting. Together, our findings support a model in which dACC detects the emergence of unwanted content, triggering top-down inhibitory control, and in which rDLPFC countermands intruding thoughts that penetrate awareness.


title: Liver Organoid and T Cell Coculture Models Cytotoxic T Cell Responses against Hepatitis C Virus

abstract: Hepatitis C virus (HCV) remains a global public health challenge with an estimated 71 million people chronically infected, with surges in new cases and no effective vaccine. New methods are needed to study the human immune response to HCV since in vivo animal models are limited and in vitro cancer cell models often show dysregulated immune and proliferative responses. Here we developed a CD8+ T cell and adult stem cell liver organoid system using a microfluidic chip to coculture 3D human liver organoids embedded in extracellular matrix with HLA-matched primary human T cells in suspension. We then employed automated phase contrast and immunofluorescence imaging to monitor T cell invasion and morphological changes in the liver organoids. This microfluidic coculture system supports targeted killing of liver organoids when pulsed with a peptide specific for HCV nonstructural protein 3 (NS3) (KLVALGINAV) in the presence of patient-derived CD8+ T cells specific for KLVALGINAV. This demonstrates the novel potential of the coculture system to molecularly study adaptive immune responses to HCV in an in vitro setting using primary human cells.

category: immunology


title: The Role of Low Molecular Weight Fungal Metabolites in Grapevine Trunk Disease Pathogenesis: Eutypa Dieback and Esca

abstract:
Eutypa dieback and Esca are serious grapevine trunk diseases (GTDs) caused by fungal consortia causing large economic losses in vineyards. Depending on the disease the species involved include Eutypa lata, Phaeoacremonium minimum, and Phaeomoniella chlamydospora. There is a need to understand the complex pathogenesis mechanisms used by these causative fungi to develop treatments for the diseases they cause. Low molecular weight metabolites (LMW) are known to be involved in non-enzymatic oxygen radical generation in fungal degradation of wood by some Basidiomycota species, and as part of our work to explore the basis for fungal consortia pathogenesis, LMW metabolite involvement by the causal GTD fungi was explored. The GTD fungal pathogens examined, Eutypa lata, Phaeoacremonium minimum and Phaeomoniella chlamydospora, were found to produce low molecular weight iron binding metabolites that preferentially reduced iron or redox cycled to produce hydrogen peroxide. Uniquely, different LMW metabolites isolated from the GTD fungi promoted distinct chemistries that are important in a type of non-enzymatic catalysis known as chelator-mediated Fenton (CMF) reactions. CMF chemistry promoted by LMW metabolites from these fungi allowed for the generation of highly reactive hydroxyl radicals under conditions promoted by the fungi. We hypothesize that this new reported mechanism may help to explain the necrosis of woody grapevine tissue as a causal mechanism important in pathogenesis in these two grapevine trunk diseases.

IMPORTANCEUnderstanding the pathogenesis of grape trunk diseases (GTDs) is the key to the development of disease control and treatment. While fungal extracellular enzyme systems are typically cited relative to their fungal mechanisms in pathogenesis, non-enzymatic mechanisms have been less studied in this regard and the role of low molecular weight (LMW) fungal metabolites in GTD development is quite limited. In this article, we demonstrate that GTD-causative fungal pathogens Eutypa lata, Phaeoacremonium minimum and Phaeomoniella chlamydospore produce LMW phenolic metabolites under iron-restricted conditions. These metabolites undergo a series of redox reactions, with different fungi producing metabolites that preferentially either reduce iron, or generate hydrogen peroxide, under conditions simulating grapevine woody tissue. These conditions have the potential to promote generation of highly damaging hydroxyl radicals through a mechanism that appears to be similar to non-enzymatic chelator-mediated Fenton (CMF) chemistry which is involved in fungal degradation of wood by non-related fungal orders. This is the first report of CMF chemistry promoted by GTD-causative fungi under laboratory conditions and the research suggests an alternate pathway that may contribute to pathogenesis in GTDs, and a potential target for vine protection.


title: Bacterial growth, communication and guided chemotaxis in 3D bioprinted hydrogel environments

abstract: Bioprinting of engineered bacteria is of great interest for applications of synthetic biology in the context of living biomaterials, but so far only few viable approaches are available for the printing of gels hosting live Escherichia coli bacteria. Here we develop a gentle bioprinting method based on an alginate/agarose bioink that enables precise printing of E.coli into three-dimensional hydrogel structures up to 10 mm in height. Addition of a calcium peroxide-based oxygen generation system enables maturation of fluorescent proteins deep within the printed structures. We utilize spatial patterning with the bioprinter to control different types of chemical interaction between bacteria. We first show quorum sensing-based chemical communication between engineered sender and receiver bacteria placed at different positions inside the bioprint, and then demonstrate the fabrication of barrier structures defined by non-motile bacteria that can guide the movement of chemotactic bacteria inside a gel.

category: bioengineering


title: Using ultraviolet absorption spectroscopy to study nanoswitches based on non-canonical DNA structures

abstract:
Non-canonical forms of DNA are attracting increasing interest for applications in nanotechnology. It is frequently convenient to characterize DNA molecules using a label-free approach such as ultraviolet absorption spectroscopy. In this paper we present the results of our investigation into the use of this technique to probe the folding of quadruplex and triplex nanoswitches. We confirmed that four G-quartets were necessary for folding at sub-mM concentrations of potassium and found that the wrong choice of sequence for the linker between G-tracts could dramatically disrupt folding, presumably due to the presence of kinetic traps in the folding landscape. In the case of the triplex nanoswitch we examined, we found that the UV spectrum showed a small change in absorbance when a triplex was formed. We anticipate that our results will be of interest to researchers seeking to design DNA nanoswitches based on quadruplexes and triplexes.

HighlightsO_LIUltraviolet absorption spectroscopy can probe non-canonical DNA structures.
C_LIO_LIAbsorbance at 295nm tends to increase as G-quadruplexes form.
C_LIO_LIFour G-quartets are needed to form a quadruplex with less than 1mM potassium.
C_LIO_LIFormation of DNA triplexes can also yield a small change in UV spectra.
C_LIO_LIUV absorption is a cheap label-free method for studying DNA nanoswitches.
C_LI


title: Cryo-EM structure of disease-related prion fibrils provides insights into seeding barriers

abstract: One of the least understood aspects of prion diseases is the structure of infectious prion protein aggregates. Here we report a high-resolution cryo-EM structure of amyloid fibrils formed by human prion protein with Y145Stop mutation that is associated with a familial prion disease. This structural insight allows us not only to explain previous biochemical findings, but also provides direct support for the conformational adaptability model of prion transmissibility barriers.


title: Incompatibility of DFHBI based fluorescent RNA aptamers with particular commercial cell-free expression systems

abstract: Fluorescent RNA aptamers are an increasingly used tool for quantifying transcription and for visualising RNA interactions, both in vitro and in vivo. However when tested in the commercially available, E. coli extract based Expressway cell-free expression system, no fluorescence is detected. The same experimental setup is shown to successfully produce fluorescent RNA aptamers when tested in another buffer designed for in vitro transcription, and RNA purification of the Expressway reaction products show that transcription does occur, but does not result in a fluorescent product. In this paper we demonstrate the incompatibility of a narrow selection of RNA aptamers in one particular cell-free expression system, and consider that similar issues may arise with other cell-free expression systems, RNA aptamers, and their corresponding fluorophores.

category: synthetic biology


title: No need to choose: independent regulation of cognitive stability and flexibility challenges the stability-flexibility tradeoff

abstract: Adaptive behavior requires the ability to focus on a current task and protect it from distraction (cognitive stability) as well as the ability to rapidly switch to another task in light of changing circumstances (cognitive flexibility). Cognitive stability and flexibility have been conceptualized as opposite endpoints on a stability-flexibility tradeoff continuum, implying an obligatory reciprocity between the two: greater flexibility necessitates less stability, and vice versa. Surprisingly, rigorous empirical tests of this critical assumption are lacking. Here, we acquired simultaneous measurements of cognitive stability (congruency effects) and flexibility (switch costs) on the same stimuli within the same task, while independently varying contextual demands on these functions with block-wise manipulations of the proportion of incongruent trials and task switches, respectively. If cognitive stability and flexibility are reciprocal, increases in flexibility in response to higher switch rates should lead to commensurate decreases in stability, and increases in stability in response to more frequent incongruent trials should result in decreased flexibility. Across three experiments, using classic cued task switching (Experiments 1 and 3) and attentional set shifting (Experiment 2) protocols, we found robust evidence against an obligatory stability-flexibility tradeoff. Although we observed the expected contextual adaptation of stability and flexibility to changing demands, strategic adjustments in stability had little influence on flexibility, and vice versa. These results refute the long-held assumption of a stability-flexibility tradeoff, documenting instead that the cognitive processes mediating these functions can be regulated independently - it is possible to be both stable and flexible at the same time.


title: Improved blind demixing methods for recovering dense neuronal morphology from barcode imaging data

abstract: Cellular barcoding methods offer the exciting possibility of  infinite-pseudocolor anatomical reconstruction -- i.e., assigning each neuron its own random unique barcoded  pseudocolor, and then using these pseudocolors to trace the microanatomy of each neuron. Here we use simulations, based on densely-reconstructed electron microscopy microanatomy, with signal structure matched to real barcoding data, to quantify the feasibility of this procedure. We develop a new blind demixing approach to recover the barcodes that label each neuron. We also develop a neural network which uses these barcodes to reconstruct the neuronal morphology from the observed fluorescence imaging data,  connecting the dots between discontiguous amplicon signals. We find that accurate recovery should be feasible, provided that the barcode signal density is sufficiently high. This study suggests the possibility of mapping the morphology and projection pattern of many individual neurons simultaneously, at high resolution and at large scale, via conventional light microscopy.


title: Structure-function analysis of distant diphtheria toxin homologs reveals insights into host adaptation

abstract: Diphtheria toxin (DT) is the archetype of bacterial exotoxins implicated in human diseases and has played a central role in defining the field of toxinology since its discovery in 1888. Despite being one of the most extensively characterized bacterial toxins, the origins and molecular evolution of DT host specialization remain unknown. Here, we determined high-resolution structures of two recently discovered distant homologs of DT. These DT-like proteins from non-human associated Streptomyces albireticuli (17% identity to DT) and Seinonella peptonophila (20% identity to DT) display remarkable structural similarity to DT enabling a comparative investigation into DTs unique toxicity toward mammalian cells. We find that the individual domains of DT-like toxins retain two critical features of DTs activity: full catalytic function and ability to translocate across mammalian cell membranes. However, we show that receptor-binding, pH-dependent pore-formation and proteolytic release of the cytotoxic enzyme into the cytosol are not optimized for human cell physiology and thus unable to efficiently deliver the cytotoxic cargo into human hosts. Our work provides structural insights into DTs evolutionary history, and implies key transitions required for the emergence of human-specificity of a major bacterial exotoxin with an important history in human disease.


title: HDAC9 structural variants disrupting TWIST1 transcriptional regulation lead to craniofacial and limb malformations

abstract: Structural variants (SVs) can affect protein-coding sequences as well as gene regulatory elements. However, SVs disrupting protein-coding sequences that also function as cis-regulatory elements remain largely uncharacterized. Here, we show that craniosynostosis patients with SVs containing the Histone deacetylase 9 (HDAC9) protein-coding sequence are associated with disruption of TWIST1 regulatory elements that reside within HDAC9 sequence. Based on SVs within the HDAC9-TWIST1 locus, we defined the 3 HDAC9 sequence (~500Kb) as a critical TWIST1 regulatory region, encompassing craniofacial TWIST1 enhancers and CTCF sites. Deletions of either Twist1 enhancers (eTw5-7{Delta}/{Delta}) or Ctcf site (Ctcf{Delta}/{Delta}) within the Hdac9 protein-coding sequence in mice led to decreased Twist1 expression and altered anterior\posterior limb expression patterns of Shh pathway genes. This decreased Twist1 expression results in a smaller sized and asymmetric skull and polydactyly that resembles Twist1+/- mouse phenotype. Chromatin conformation analysis revealed that the Twist1 promoter region interacts with Hdac9 sequences that encompass Twist1 enhancers and a Ctcf site and that interactions depended on the presence of both regulatory regions. Finally, a large inversion of the entire Hdac9 sequence (Hdac9INV/+) in mice that does not disrupt Hdac9 expression but repositions Twist1 regulatory elements showed decreased Twist1 expression and led to a craniosynostosis-like phenotype and polydactyly. Thus, our study elucidated essential components of TWIST1 transcriptional machinery that reside within the HDAC9 sequence, suggesting that SVs, encompassing protein-coding sequence, such as HDAC9, could lead to a phenotype that is not attributed to its protein function but rather to a disruption of the transcriptional regulation of a nearby gene, such as TWIST1.


title: Parent-of-origin effects propagate through networks to shape metabolic traits

abstract: Parent-of-origin effects are unexpectedly common in complex traits, including metabolic and neurological diseases. Parent-of-origin effects can be modified by the environment, but the architecture of these gene-by-environmental effects on phenotypes remains to be unraveled. Previously, quantitative trait loci (QTL) showing context-specific parent-of-origin effects on metabolic traits were mapped in the F16 generation of an advanced intercross between LG/J and SM/J inbred mice. However, these QTL were not enriched for known imprinted genes, suggesting another mechanism is needed to explain these parent-of-origin effects phenomena. We propose that non-imprinted genes can generate complex parent-of-origin effects on metabolic traits through interactions with imprinted genes. Here, we employ data from mouse populations at different levels of intercrossing (F0, F1, F2, F16) of the LG/J and SM/J inbred mouse lines to test this hypothesis. Using multiple populations and incorporating genetic, genomic, and physiological data, we leverage orthogonal evidence to identify networks of genes through which parent-of-origin effects propagate. We identify a network comprised of 3 imprinted and 6 non-imprinted genes that show parent-of-origin effects. This epistatic network forms a nutritional responsive pathway and the genes comprising it jointly serve cellular functions associated with growth. We focus on 2 genes, Nnat and F2r, whose interaction associates with serum glucose levels across generations in high fat-fed females. Single-cell RNAseq reveals that Nnat and F2r are negatively correlated in pre-adipocytes along an adipogenic trajectory, a result that is consistent with our observations in bulk white adipose tissue.


title: QSAR Model Based Gradient Boosting Regression of N-Arylsulfonyl-Indole-2-Carboxamide Derivatives as Inhibitors for Fructose-1,6-Bisphosphatase

abstract: As is known to all, diabetes metellius is a global health threaten and it has caused worldwide attention of scientists. To get a better investigation of the drug design of diabetes, we used heuristic method to established the linear model and used Gradient Boosting Regression to establish the nonlinear model of Fructose-1,6-Bisphosphatse inhibitor successively. In this study, 84 derivatives of N-Arylsulfonyl-Indole-2-Carboxamide were introduced into the models, two outstanding QSAR models with 2 molecule descriptors were established successfully. Grandient Boosting Regression rendered a good correlation with R2 of 0.943 and MSE of 0.135 for the training set, 0.916 and 0.213 for test set, which also proves the feasibility of the implementation of the new method GBR in the field of QSAR. Meanwhile, the optimal model displayed wonderful statistical significance. This study shows unlimited potential for design of new drugs for diabetes.

category: biochemistry


title: SCL28 promotes cell expansion and endoreplication in Arabidopsis by activating SIAMESE-RELATED cyclin-dependent kinase inhibitors.

abstract: The processes that contribute to plant organ morphogenesis are spatial-temporally organized. Within the meristem the mitotic cell cycle produces new cells that subsequently engage in specific cell expansion and differentiation programs once they exit the division competent zone. The latter is frequently accompanied by endoreplication, being an alternative cell cycle that replicates the DNA without nuclear division, causing a stepwise increase in somatic ploidy. We have previously shown that the Arabidopsis SCL28 transcription factor promotes progression through G2/M and modulates division plane orientation. Here, we demonstrate that SCL28 co-express and regulates genes specific to cell elongation and differentiation, including genes related to cell wall and cytoskeleton assembly. Consistently, this correlates with defects in post-mitotic cell expansion in a scl28 mutant. Strikingly, SCL28 controls expression of 6 members of the SIAMESE/SIAMESE-RELATED (SIM/SMR) family, encoding cyclin-dependent kinase inhibitors with a role in promoting mitotic cell cycle exit and endoreplication onset, both in response to developmental and environmental cues. Consistent with this role, scl28 mutants displayed reduced endoreplication, both in roots and leaves. Altogether, these results suggest that SCL28 controls cell expansion and differentiation by promoting endoreplication onset and by modulating aspects of the biogenesis, assembly and remodeling of the cytoskeleton and cell wall.


title: Crosstalk between Ethylene and ABA during changes in soil water content reveals a role of ACC in coffee anthesis regulation

abstract:
O_LICoffee (Coffea arabica L.) presents an asynchronous flowering regulated by endogenous and environmental stimulus, and anthesis occurs once plants are rehydrated after a period of water deficit.
C_LIO_LIWe evaluated the evolution of Abscisic Acid (ABA), ethylene, 1-aminocyclopropane-1-carboxylate (ACC) content, ACC oxidase (ACO) activity, and expression analysis of the Lysine Histidine Transporter 1 (LHT1) transporter, in roots, leaves and, flower buds from three coffee genotypes (Coffea arabica L. cv Oeiras, Acaua, and Semperflorens) cultivated under field conditions with two experiments. In a third field experiment, the effect of exogenous supply of ACC in coffee anthesis was evaluated.
C_LIO_LIWe found an increased ACC level in all tissues from the three coffee genotypes in the re-watering period just before anthesis for all tissues and high expression of the LHT1 gene in flower buds and leaves. Ethylene content and ACO activity decreased from rainy to dry period whereas ABA content increased. Higher number of opened and G6 stage flower buds were observed in the treatment with exogenous ACC.
C_LIO_LIThe results showed that the interaction of ABA-ACO-ethylene and intercellular ACC transport among leaves, buds, and roots in coffee favors an increased level of ACC that is most likely, involved as a modulator in coffee anthesis.
C_LI


title: Cortical efficient coding dynamics shape behavioral performance.

abstract: The efficient coding hypothesis postulates that neurons shape their response properties to match their dynamic range to the statistics of incoming signals. However, whether and how the dynamics of efficient neuronal adaptation inform behavior has not been directly shown. Here, we trained mice to detect a target presented in background noise shortly after a change in the background contrast. The observed changes in cortical gain and detection behavior followed the predictions of a normative model of efficient cortical sound processing; specifically, target detection and sensitivity to target volume improved in low contrast backgrounds relative to high contrast backgrounds. Additionally, the time course of target detectability adapted asymmetrically depending on contrast, decreasing rapidly after a transition to high contrast, and increasing more slowly after a transition to low contrast. Auditory cortex was required for detection of targets in background noise and cortical neuronal responses exhibited the patterns of target detectability observed during behavior and in the normative model. Furthermore, variability in cortical gain predicted behavioral performance beyond the effect of stimulus-driven gain control. Combined, our results demonstrate that efficient neural codes in auditory cortex directly influence perceptual behavior.


title: Dopamine D3 Receptor Ligand Suppresses the Expression of Levodopa-Induced Dyskinesia in Nonhuman Primate Model of Parkinson's Disease

abstract: Parkinsons disease (PD) is a complex multisystem, chronic and so far incurable disease with significant unmet medical needs. The incidence of PD increases with aging and the expected burden will continue to escalate with our aging population. Since its discovery in the 1961 levodopa remains the gold standard pharmacotherapy for PD. However, the progressive nature of the neurodegenerative process in and beyond the nigrostriatal system causes a multitude of side effects, including levodopa-induced dyskinesia within 5 years of therapy. Attenuating dyskinesia has been a significant challenge in the clinical management of PD. We report on a small molecule that eliminates the expression of levodopa-induced dyskinesia and significantly improves PD-like symptoms. The lead compound PD13R we discovered is a dopamine D3 receptor partial agonist with high affinity and selectivity, orally active and with desirable drug-like properties. Future studies are aimed at developing this lead compound for treating PD patients with dyskinesia.


title: Hierarchical control of visually-guided movements in a 3D-printed robot arm

abstract: The control architecture guiding simple movements such as reaching toward a visual target remains an open problem. The nervous system needs to integrate different sensory modalities and coordinate multiple degrees of freedom in the human arm to achieve that goal. The challenge increases due to noise and transport delays in neural signals, nonlinear and fatigable muscles as actuators, and unpredictable environmental disturbances. Here we examined the capabilities of a previously proposed hierarchical feedback control model (Powers 1999, 2008), so far only tested in silico. We built a robot arm system with four degrees of freedom, including a visual system for locating the planar position of the hand, joint angle proprioception, and pressure sensing in one point of contact. We subjected the robot to various human-inspired reaching and tracking tasks and found features of biological movement, such as isochrony and bell-shaped velocity profiles in straight-line movements, and the speed-curvature power law in curved movements. These behavioral properties emerge without trajectory planning or explicit optimization algorithms. We then applied static structural perturbations to the robot: we blocked the wrist joint, tilted the writing surface, extended the hand with a tool, and rotated the visual system. For all of them, we found that the arm in machina adapts its behavior without being reprogrammed. In sum, while limited in speed and precision (by the nature of the do-it-yourself inexpensive components we used to build the robot from scratch), when faced with the noise, delays, nonlinearities, and unpredictable disturbances of the real world, the embodied control architecture shown here balances biological realism with design simplicity.