bluuebunny/crossref_metadata_2025 huggingface dataset:
Systems-wide dissection of organic acid assimilation in Pseudomonas aeruginosa reveals a novel path to underground metabolism
Authors: Dolan Stephen K. , Wijaya Andre , Kohlstedt Michael , Gläser Lars , Brear Paul , Silva-Rocha Rafael , Wittmann Christoph , Welch Martin
Published: August 2022

The human pathogen Pseudomonas aeruginosa (Pa) is one of the most frequent and severe causes of nosocomial infection. This organism is also a major cause of airway infections in people with cystic fibrosis (CF). Pa is known to have a remarkable metabolic plasticity, allowing it to thrive in diverse environmental conditions and ecological niches, yet little is known about the central metabolic pathways which sustain its growth during infection, or precisely how these pathways operate. In this work, we used a combination of ‘omics approaches (transcriptomics, proteomics, metabolomics and 13C-fluxomics) and reverse genetics to provide a systems-level insight into how the infection-relevant organic acids, succinate and propionate, are metabolized by Pa. Moreover, through structural and kinetic analysis of the 2-methylcitrate synthase (PrpC) and its paralogue, citrate synthase (GltA), we show how these two crucial enzymatic steps are interconnected in Pa organic acid assimilation. We found that Pa can rapidly adapt to the loss of GltA function by acquiring mutations in a transcriptional repressor, which then de-represses prpC expression. Our findings provide a clear example of how ‘underground metabolism’, facilitated by enzyme substrate promiscuity, “rewires” Pa metabolism, allowing it to overcome the loss of a crucial enzyme. This pathogen-specific knowledge is critical for the advancement of a model-driven framework to target bacterial central metabolism.
SEC-seq: Association of molecular signatures with antibody secretion in thousands of single human plasma cells
Authors: Cheng Rene Yu-Hong , de Rutte Joseph , Ott Andee R. , Bosler Lucienne , Kuo Wei-Ying , Liang Jesse , Hall Brian E , Rawlings David J , Carlo Dino Di , James Richard G.
Published: August 2022

Protein secretion drives many functionsin vivo; however, methods to link secretions with surface markers and transcriptomes have been lacking. By accumulating secretions close to secreting cells held within cavity-containing hydrogel nanovials, we demonstrate workflows to analyze the amount of IgG secreted from single human antibody-secreting cells and link this information to surface marker expression and transcriptional profiles from the same cells. Measurements using flow cytometry and imaging flow cytometry corroborated an association between levels of IgG secretion and CD138 expression. Using oligonucleotide-labeled antibodies and droplet-based sequencing, we show that pathways encoding protein localization to the endoplasmic reticulum, NADH complex assembly, and mitochondrial respiration were most associated with high IgG secretion. Altogether, this method links secretion information to cell surface and single-cell sequencing information (SEC-seq) and enables exploration of links between genome and secretory function, laying the foundation for numerous discoveries in immunology, stem cell biology, and beyond.
PP4 inhibition sensitizes ovarian cancer to NK cell-mediated cytotoxicity via STAT1 activation and inflammatory signaling
Authors: Raja Remya , Wu Christopher , Rubino Thomas E , Utagawa Emma , Magtibay Paul , Butler Kristina A. , Curtis Marion
Published: August 2022

Background

Increased infiltration of T cells into ovarian tumors has been repeatedly shown to be predictive of enhanced patient survival. However, despite the evidence of an active immune response in ovarian cancer (OC), the frequency of responses to immune checkpoint blockade (ICB) therapy in OC is much lower than other cancer types. Recent studies have highlighted that deficiencies in the DNA damage response (DDR) can drive increased genomic instability and tumor immunogenicity, which leads to enhanced responses to ICB. Protein phosphatase 4 (PP4) is a critical regulator of the DDR; however, its potential role in anti-tumor immunity is currently unknown.

Results

Our results show that the PP4 inhibitor, fostriecin, combined with carboplatin leads to increased carboplatin sensitivity, DNA damage, and micronuclei formation. Using multiple ovarian cancer cell lines, we show that PP4 inhibition or PPP4C knockdown combined with carboplatin triggers inflammatory signaling via NFκB and STAT1 activation. This resulted in increased expression of the pro-inflammatory cytokines and chemokines: CCL5, CXCL10, and IL6. In addition, IFNB1 expression was increased suggesting activation of the type I interferon response. Conditioned media from OC cells treated with the combination of PP4 inhibitor and carboplatin significantly increased migration of both CD8 T cell and NK cells over carboplatin treatment alone. Knockdown of STING in OC cells significantly abrogated the increase in CD8 T cell migration induced by PP4 inhibition. Co-culture of NK-92 cells and OC cells with PPP4C or PPP4R3B knockdown resulted in strong induction of NK cell activation as measured by IFN-γ levels. Further, we also observed increased degranulation and NK cell-mediated cytotoxicity against OC cells with PPP4C or PPP4R3B knockdown.

Conclusions

Our work has identified a role for PP4 inhibition in promoting inflammatory signaling and enhanced immune cell effector function. These findings support the further investigation of PP4 inhibitors to enhance chemo-immunotherapy for ovarian cancer treatment.
Abiotic niche predictors of long-term trends in body mass and survival of Eastern Himalayan birds
Authors: Bharadwaj Akshay , Chanda Ritobroto , Biswakarma Aman , Tamang Bharath , Munda Binod , Pradhan Dambar K , Rai Mangal K , Rai Shambu , Srinivasan Umesh
Published: August 2022

The synergistic impacts of climate change and habitat degradation threaten tropical species worldwide. However, how species’ abiotic niches affect their demographic vital rates and phenotypic changes under anthropogenic change remains poorly understood. Using an 11-year mark-recapture dataset from primary and selectively logged forest in the Eastern Himalayas, we investigated how temperature-humidity niche characteristics predicted body mass and survival trends in understorey insectivorous birds over time in each habitat. Our results show that logged forest is hotter and drier than primary forest, and the arthropod community shows dramatic shifts in composition upon selective logging. In understorey insectivores, the degree of dissimilarity between species-specific primary and logged forest niches was strongly and negatively correlated with survival and body mass trends in logged forest. Here, we show that temperature-humidity niche shifts in response to anthropogenic habitat modification can impact demographic vital rates and body condition crucial for population persistence. This work has the potential to inform prompt, targeted conservation efforts toward species that are the most threatened in a warmer and more degraded world.
Generic Framework for Quantifying the Influence of the Mitral Valve on Ventricular Blood Flow
Authors: Thiel Jan-Niklas , Steinseifer Ulrich , Neidlin Michael
Published: August 2022

Blood flow within the left ventricle provides important information regarding cardiac function in health and disease. The mitral valve strongly influences the formation of flow structures and there exist various approaches for the representation of the valve in numerical models of left ventricular blood flow. However, a systematic comparison of the various mitral valve models is missing, making a priori decisions considering the overall model’s context of use impossible. Within this study, a benchmark setup to compare the influence of mitral valve modeling strategies on intraventricular flow features was developed. Then, five mitral valve models of increasing complexity: no modeling, static wall, 2D and 3D porous medium with time-dependent porosity, and one-way Fluid-Structure-Interaction (FSI) were compared with each other. The flow features velocity, kinetic energy, transmitral pressure drop, vortex formation, flow asymmetry as well as computational cost and ease-of-implementation were evaluated. The one-way FSI approach provides the highest level of flow detail, which is accompanied by the highest numerical costs and challenges with the implementation. As an alternative, the porous medium approach with our new expansion including time-dependent porosity provides good results with up to 10% deviations in the flow features (except the transmitral pressure drop) in comparison to the FSI model and only a fraction (11%) of numerical costs. Taken together, our benchmark setup allows a quantitative comparison of various mitral valve modeling approaches and is provided to the scientific community for further testing and expansion..
Task matters: individual MEG signatures from naturalistic and neurophysiological brain states
Authors: Colenbier Nigel , Sareen Ekansh , del-Águila Puntas Tamara , Griffa Alessandra , Pellegrino Giovanni , Mantini Dante , Marinazzo Daniele , Arcara Giorgio , Amico Enrico
Published: August 2022

The discovery that human brain connectivity data can be used as a “fingerprint” to identify a given individual from a population, has become a burgeoning research area in the neuroscience field. Recent studies have identified the possibility to extract these brain signatures from the temporal rich dynamics of resting-state magnetoencephalography (MEG) recordings. However, to what extent MEG signatures constitute a marker of human identifiability when engaged in task-related behavior remains an open question. Here, using MEG data from naturalistic and neurophysiological tasks, we show that identification improves in tasks relative to resting-state, providing compelling evidence for a task dependent axis of MEG signatures. Notably, improvements in identifiability were more prominent in strictly controlled tasks. Lastly, the brain regions contributing most towards individual identification were also modified when engaged in task activities. We hope that this investigation advances our understanding of the driving factors behind brain identification from MEG signals.
Non-cyanobacterial diazotrophs from the Rhizobiales order support marine microalgae in nitrogen-depleted environments
Authors: Chandola Udita , Gaudin Marinna , Trottier Camille , Lavier Aydat Louis Josselin , Manirakiza Eric , Menicot Samuel , Fischer Erik Jörg , Louvet Isabelle , Lacour Thomas , Chaumier Timothée , Tanaka Atsuko , Pohnert Georg , Chaffron Samuel , Tirichine Leïla
Published: August 2022

Background

Non-cyanobacteria diazotrophs (NCDs) were shown to dominate in surface waters shifting the long-held paradigm of cyanobacteria dominance and raising fundamental questions on how these putative heterotrophic bacteria thrive in sunlit oceans. The absence of laboratory cultures of these bacteria significantly limits our ability to understand their behavior in natural environments and, consequently, their contribution to the marine nitrogen cycle.

Results

Here, we used a multidisciplinary approach and report an unprecedented finding in the diatomPhaeodactylum tricornutum(Pt) of NCDs in the phycosphere or the pelagic community sustaining its survival in the absence of bioavailable nitrogen. We sequenced the bacterial metacommunity associated with Pt and assembled several bacterial genomes, identifying multiple NCDs from the Rhizobiales order, includingBradyrhizobium,Mesorhizobium,GeorhizobiumandMethylobacterium. We demonstrated the nitrogen-fixing ability of PtNCDs through in silico identification of nitrogen fixation genes, or by using PCR, acetylene reduction, or 15N incorporation. We showed the wide occurrence of this type of interactions with the isolation of NCDs from other microalgae, their identification in the environment, and their predicted associations with photosynthetic microalgae.

Conclusions

Our study underscores the importance of microalgae interactions with NCDs to permit and support nitrogen fixation. This work provides a unique model Pt-NCDs to study the ecology of this interaction advancing our understanding of the key drivers of global marine nitrogen fixation.
Revealing the molecular mechanisms underlying neural transmission of female pheromone signals in the male antennae of Antheraea pernyi by integrative proteomics and metabolomics analysis
Authors: Wang Guobao , Ji Xiang , Nie Lei
Published: August 2022

Detection of sex pheromones of insects relies on the antennae. The female pheromone signal transmission in the male antennae ultimately initiates the courtship and mating behaviors of males. To investigate the proteins and metabolites involved in this neural transduction, the study adopted integrative proteomics and metabolomics analysis including tandem mass tag (TMT) proteomic quantification and liquid chromatography tandem mass spectrometry (LC/MS)-based metabolomics for comparing proteomic and metabolic changes between the antennae of male moths following stimulation by females and the non-stimulated males of A. pernyi. A total of 92 differentially expressed proteins (DEPs) containing 52 up-regulated and 40 down-regulated proteins and 545 differentially expressed metabolites (DEMs) including 218 up- and 327 down-regulated metabolites were identified from the antennae of female-stimulated male moths based on the proteome and metabolome data, respectively. GO enrichment analysis showed that 45 DEPs could be enriched into different GO terms on different levels. COG analysis indicated that 61 DEPs were assigned to 20 functional categories. The 160 DEMs respectively fell into 11 and 44 classes at SuperClass and Class levels based on HMDB annotation. KEGG pathway enrichment analysis indicated that totally 43 DEMs were enriched into 6, 27, and 87 pathways on level 1, 2, and 3, respectively. A number of DEPs and DEMs related to neural transmission of female pheromone signals in the male antennae of A. pernyi were screened, including tyrosine hydroxylase, cryptochrome-1, tachykinin, arylalkylamine N-acetyltransferase, cadherin-23, glutathione S-transferase delta 3, tyramine, tryptamine, n-oleoyl dopamine, n-stearoyl dopamine, and n-stearoyl tyrosine. We concluded that the altered expression levels of those proteins or metabolites were involved in regulating the neuron activity for enhanced transmission of neural impulses and continuous perception, reception, and transduction of female pheromone signals. Our findings yielded novel insights into the potential molecular mechanisms in the antennae of male A. pernyi responding to female attraction.
Deep learning-driven insights into super protein complexes for outer membrane protein biogenesis in bacteria
Authors: Gao Mu , An Davi Nakajima , Skolnick Jeffrey
Published: August 2022

To reach their final destinations, outer membrane proteins (OMPs) of gram-negative bacteria undertake an eventful journey beginning in the cytosol. Multiple molecular machines, chaperones, proteases, and other enzymes facilitate the translocation and assembly of OMPs. These helpers usually associate, often transiently, forming large protein assemblies. They are not well-understood due to experimental challenges in capturing and characterizing protein-protein interactions (PPIs), especially transient ones. Using AF2Complex, we introduce a high-throughput, deep learning pipeline to identify PPIs within the E. coli cell envelope and apply it to several proteins from an OMP biogenesis pathway. Among the top confident hits obtained from screening ~1,500 envelope proteins, we find not only expected interactions but also unexpected ones with profound implications. Subsequently, we predict atomic structures for these protein complexes. These structures, typically of high confidence, explain experimental observations, and lead to mechanistic hypotheses for how a chaperone assists a nascent, precursor OMP emerging from a translocon, how another chaperone prevents it from aggregating and docks to an β-barrel assembly port, and how a protease performs quality control. This work presents a general strategy for investigating biological pathways by using structural insights gained from deep learning-based predictions.
Single-cell RNA-seq reveals stepwise fate conversion of supporting cells to hair cells in the chick auditory epithelium
Authors: Matsunaga Mami , Yamamoto Ryosuke , Kita Tomoko , Ohnishi Hiroe , Yamamoto Norio , Okano Takayuki , Omori Koichi , Nakagawa Takayuki
Published: August 2022

In contrast to mammals, the avian cochlea, specifically the basilar papilla, can regenerate sensory hair cells, which involves fate conversion of supporting cells to hair cells. To determine the mechanisms for converting supporting cells to hair cells, we used single-cell RNA sequencing during hair cell regeneration in explant cultures of chick basilar papillae. We identified dynamic changes in the gene expression of supporting cells, and the pseudotime trajectory analysis demonstrated the stepwise fate conversion from supporting cells to hair cells. Initially, supporting cell identity was erased and transition to the precursor state occurred. A subsequent gain in hair cell identity progressed together with downregulation of precursor-state genes. Transforming growth factor beta receptor 1-mediated signaling was involved in induction of the initial step, and its inhibition resulted in suppression of hair cell regeneration. Our data provide new insights for understanding fate conversion from supporting cells to hair cells in avian basilar papillae.
Genome-wide peptidoglycan profiling of Vibrio cholerae
Authors: Hernandez Sara B. , Alvarez Laura , Ritzl-Rinkenberger Barbara , Schiffthaler Bastian , Serrano Alonso R. , Cava Felipe
Published: August 2022

Most bacteria cells are protected by a peptidoglycan cell wall. Defining the chemical structure of the peptidoglycan has been instrumental to characterize cell wall associated proteins and to illuminate the mode of action of cell wall-acting antibiotics. However, a major roadblock for a comprehensive understanding of peptidoglycan homeostasis has been the lack of methods to conduct large-scale, systematic studies. Here we have developed and applied an innovative high throughput peptidoglycan analytical pipeline to analyze the entire non-essential, arrayed mutant library of Vibrio cholerae. The unprecedented breadth of these analyses revealed that peptidoglycan homeostasis is preserved by a large percentage of the genome organized in complex networks that functionally link peptidoglycan features with genetic determinants. As an example, we discovered a novel bifunctional penicillin-binding protein in V. cholerae. Collectively, genome-wide peptidoglycan profiling provides a fast, easy, and unbiased method for systematic identification of the genetic determinants of peptidoglycan synthesis and remodeling.
Inhibition of S6K lowers age-related inflammation and immunosenescence and increases lifespan through the endolysosomal system
Authors: Zhang Pingze , Catterson James H. , Grönke Sebastian , Partridge Linda
Published: August 2022

The nutrient-sensing Target of Rapamycin complex 1 (TORC1) is an evolutionarily conserved regulator of longevity and healthspan. S6 kinase (S6K) is an essential downstream mediator for the effect of TORC1 on longevity. However, mechanistic insights on how TORC1-S6K signalling promotes lifespan and healthspan are still limited. Here we show that activity of S6K in theDrosophilafat body is essential for rapamycin-mediated longevity. Fat-body-specific activation of S6K blocked lifespan extension upon rapamycin feeding and induced accumulation of multilamellar lysosomal enlargements. By proteomics analysis we identified Syntaxin 13 (Syx13) as an important downstream mediator of TORC1-S6K signalling involved in regulating lysosomal morphology. Inhibition of TORC1-S6K signalling decreased age-associated hyperactivation of the NF-κB-like IMD pathway in the fat body and promoted bacterial clearance, mediated by Syx13, suggesting that lysosomal and immune function are connected during ageing. Middle-age-onset repression of IMD pathway in the fat body by Relish RNAi promoted bacteria clearance and extended fly lifespan. In mice, chronic rapamycin treatment elevated Syntaxin 12/13 (Stx12) level in liver. We identified alleviated immune processes in the aged liver as a common signature of S6K1-deficient and rapamycin-treated mice. Thus, our results indicate that suppression of the TORC1-S6K-Syx13 axis ameliorates both inflammageing and immunosenescence in hepatic tissues via the endolysosomal system and thereby extends longevity, providing a mechanistic explanation for the effects of rapamycin and suppression of S6K on immune function and lifespan in model organisms and, potentially, humans.
Intrinsic neural diversity quenches the dynamic volatility of neural networks
Authors: Hutt Axel , Rich Scott , Valiante Taufik A , Lefebvre Jérémie
Published: August 2022

Heterogeneity is the norm in biology. The brain is no different: neuronal cell-types are myriad, reflected through their cellular morphology, type, excitability, connectivity motifs and ion channel distributions. While this biophysical diversity enriches neural systems’ dynamical repertoire, it remains challenging to reconcile with the robustness and persistence of brain function over time. To better understand the relationship between heterogeneity and resilience, we analyzed both analytically and numerically a non-linear sparse neural network with balanced excitatory and inhibitory connections evolving over long time scales. We examined how neural diversity expressed as excitability heterogeneity in this network influences its dynamic volatility (i.e., its susceptibility to critical transitions). We exposed this network to slowly-varying modulatory fluctuations, continuously interrogating its stability and resilience. Our results show that excitability heterogeneity implements a homeostatic control mechanism tuning network stability in a context-dependent way. Such diversity was also found to enhance network resilience, quenching the volatility of its dynamics, effectively making the system independent of changes in many control parameters, such as population size, connection probability, strength and variability of synaptic weights as well as modulatory drive. Taken together, these results highlight the fundamental role played by cell-type heterogeneity in the robustness of brain function in the face of change.

Significance Statement

Contemporary research has identified widespread cell-to-cell intrinsic diversity in the brain, manifest through variations in biophysical features such as neuronal excitability. A natural question that arises from this phenomenon is what functional role, if any, this heterogeneity might serve. Combining computational and mathematical techniques, this interdisciplinary research shows that intrinsic cell-to-cell diversity, far from mere developmental noise, represents a homeostatic control mechanism, promoting the resilience of neuronal circuits. These results highlight the importance of diversity in the robustness and persistence of brain function over time and in the face of change.
Changes in social position predict survival in bottlenose dolphins
Authors: Rankin Robert William , Foroughirad Vivienne Jilla , Krzyszczyk Ewa Beata , Frère Céline H , Mann Janet
Published: August 2022

1

Social bonds and social structure are important features of animal systems that impact individual fitness. Few studies have examined how temporal dynamics in individual social bonds predict fitness outcomes. This is critical to understand given the high variation in types of social structures and strategies within populations of social mammals, both across time and among individuals. If individually-differentiated social bonds are important, it might be the change in bonds which has fitness consequences, not the absolute number of bonds nor their strength. We investigated how network dynamics predict survival in a wild population of bottlenose dolphins using a 35-year longitudinal study. In particular, we were interested in two sex-specific measures of the “widowhood effect”, as well as a more general investigation into the predictability of mortality from changes in higher-order social network metrics. We used two inferential frameworks to provide complimentary evidence for or against hypotheses; namely, a gradient-boosting predictivist approach with relative importance measures, and an inclusion probabilities approach based on stability-selection. We found evidence of a widowhood effect among males but not females. Surprisingly, the most robust predictor of survival was closeness-centrality, whereby the loss of closeness centrality preceded death. This finding, that absolute network position may not be as relevant to survival as changes in network status, is consistent with a large social-psychological literature in humans on the impacts of loss of social capital, dissolution of close friendships, and loss through death of partners (widowhood effect). This study highlights the critical nature of social connections and how its disruption can be a matter of life and death.
Pathogen transmission modes determine contact network structure, altering other pathogen characteristics
Authors: Collier Melissa , Albery Gregory F , McDonald Grant C. , Bansal Shweta
Published: August 2022

Pathogen traits can vary greatly and impact the ability of a pathogen to persist in a population. Although this variation is fundamental to disease ecology, little is known about the evolutionary pressures that drive these differences, particularly where they interact with host behavior. We hypothesized that host behaviors relevant to different transmission routes give rise to differences in contact network structure, constraining the space over which pathogen traits can evolve to maximize fitness. Our analysis of 232 contact networks across mammals, birds, reptiles, amphibians, arthropods, fish, and mollusks found that contact network topology varies by contact events, most notably in networks that are representative of fluid-exchange transmission. Using infectious disease model simulations, we showed that these differences in network structure suggest pathogens transmitted through fluid-exchange contact events will need traits associated with high transmissibility to successfully proliferate, compared to pathogens that transmit through other types of contact. These findings were supported through a review of known traits of pathogens that transmit in humans. Our work demonstrates that contact network structure may drive the evolution of compensatory pathogen traits according to transmission strategy, providing essential context for understanding pathogen evolution and ecology.
Reprogramming of fish somatic cells for nuclear transfer is primed by Xenopus egg extract
Authors: Chênais Nathalie , Le Cam Aurelie , Guillet Brigitte , Lareyre Jean-Jacques , Labbé Catherine
Published: August 2022

Somatic cell reprogramming in vitro prior to nuclear transfer is one strategy expected to improve clone survival during development. In this study, we investigated the reprogramming extent of fish fin somatic cells after in vitro exposure to Xenopus egg extract and subsequent culture. Using a cDNA microarray approach, we observed drastic changes in the gene expression profile of the treated cells. Several actors of the TGFβ and Wnt/β-catenin signaling pathways, as well as some mesenchymal markers, were inhibited in treated cells, while several epithelial markers were upregulated. This was associated with morphological changes of the cells in culture, suggesting that egg extract drove somatic cells towards a mesenchymal-epithelial transition (MET), the hallmark of somatic reprogramming in induced pluripotent stem cells (iPSCs). However, treated cells were also characterized by a strong decrease in de novo lipid biosynthesis metabolism, the lack of re-expression of pou2 and nanog pluripotency markers, and absence of DNA methylation remodeling of their promoter region. In all, this study showed that Xenopus egg extract treatment initiated an in vitro reprogramming of fin somatic cells in culture. Although not thorough, the induced changes have primed the somatic chromatin for a better embryonic reprogramming upon nuclear transfer.
Syntaxin-6 delays prion protein fibril formation and prolongs presence of toxic aggregation intermediates
Authors: Sangar Daljit , Hill Elizabeth , Jack Kezia , Batchelor Mark , Mistry Beenaben , Ribes Juan M. , Jackson Graham S. , Mead Simon , Bieschke Jan
Published: August 2022

Prions replicate via the autocatalytic conversion of cellular prion protein (PrPC) into fibrillar assemblies of misfolded PrP. While this process has been extensively studied in vivo and in vitro, non-physiological reaction conditions of fibril formation in vitro have precluded the identification and mechanistic analysis of cellular proteins, which may alter PrP self-assembly and prion replication. Here, we have developed a fibril formation assay for recombinant murine and human PrP (23-231) under near-native conditions (NAA) to study the effect of cellular proteins, which may be risk factors or potential therapeutic targets in prion disease. Genetic screening suggests that variants that increase syntaxin-6 expression in the brain (gene: STX6) are risk factors for sporadic Creutzfeldt-Jakob disease (CJD). Analysis of the protein in NAA revealed counterintuitively that syntaxin-6 is a potent inhibitor of PrP fibril formation. It significantly delayed the lag phase of fibril formation at highly sub-stoichiometric molar ratios. However, when assessing toxicity of different aggregation time points to primary neurons, syntaxin-6 prolonged the presence of neurotoxic PrP species. Electron microscopy and super-resolution fluorescence microscopy revealed that, instead of highly ordered fibrils, in the presence of syntaxin-6 PrP formed less-ordered aggregates containing syntaxin-6. These data strongly suggest that the protein can directly alter the initial phase of PrP self-assembly and, uniquely, can act as an ‘anti-chaperone’, which promotes toxic aggregation intermediates by inhibiting fibril formation.
Vimentin intermediate filaments provide structural stability to the mammalian Golgi apparatus
Authors: Vitali Teresa , Witkos Tomasz M. , Cutiongco Marie F.A. , Yan Guanhua , Mironov Alexander A. , Swift Joe , Lowe Martin
Published: August 2022

The Golgi apparatus comprises a connected ribbon of stacked cisternal membranes localized to the perinuclear region of most vertebrate cells. The position and morphology of this organelle depends upon interactions with microtubules and the actin cytoskeleton. In contrast, we know relatively little about the relationship of the Golgi apparatus with intermediate filaments. In this study we show that the Golgi is in close physical proximity to vimentin intermediate filaments (IFs) in cultured mouse and human cells. We also show that the trans-Golgi network coiled-coil protein GORAB can physically associate with IFs. Although loss of vimentin and/or GORAB does not have major effects upon Golgi morphology at steady-state, the Golgi undergoes more rapid disassembly upon chemical disruption with the drug brefeldin A, and slower reassembly upon drug washout, in vimentin knockout cells. Moreover, loss of vimentin causes reduced Golgi ribbon integrity when cells are cultured on high stiffness hydrogels, which is exacerbated by loss of GORAB. These results indicate that vimentin IFs contribute to the structural stability of the Golgi apparatus, and suggest a role for GORAB in this process.
Elasmobranchs’ metabarcoding requires a pragmatic approach to reach its promises
Authors: Cruz Marcelo Merten , Sauvage Thomas , Chariton Anthony , de Freitas Thales Renato Ochotorena
Published: August 2022

Human impacts have been eroding marine ecosystems in such a way that biodiversity patterns are changing. Therefore, policies and science-based solutions are indispensable for monitoring threats to the most impacted species. In such effort, the analysis of elasmobranchs’ environmental traces via eDNA metabarcoding represent a candidate tool for effective monitoring and conservation that is often advocated to be cost-effective and easily replicated. Here, we tested a realistic approach to monitor future changes through elasmobranchs’ metabarcoding with published primers, in which, elasmobranch diversity from the coastal waters of the Fernando de Noronha Archipelago (Brazil) was studied here. We detected a total of three elasmobranch species, namelyHypanus berthalutzae, Ginglymostoma cirratum, andPrionace glaucaamong numerous other fish species. Even though the technique proved to be a useful tool, some practical constraints were identified, and primarily caused by currently published environmental primers. In order to ensure the broad application of the method, we pointed out feasible adjustments to the problematic parameters based on our survey and other elasmobranch metabarcoding studies. The current drawbacks of the approach need to be considered by managers, conservation actors, and researchers, who are considering this methodology in order to avoid unrealistic promises for the cost incurred.
Reprograming human fibroblasts into Sertoli cells: a tool for personalized medicine
Authors: Parivesh Abhinav , Délot Emmanuèle , Reyes Alejandra , Ryan Janelle , Bhattacharya Surajit , Harley Vincent , Vilain Eric
Published: August 2022

Disorders/Differences of Sex Development (DSD) are congenital conditions in which the development of chromosomal, gonadal, or anatomical sex is atypical. With overlapping phenotypes and multiple genes involved, poor diagnostic yields are achieved for many of these conditions. The current DSD diagnostic regimen can be augmented by investigating transcriptome/proteome in vivo, but it is hampered by the unavailability of affected gonadal tissue at the relevant developmental stage. We try to mitigate this limitation by reprogramming readily available skin tissue-derived dermal fibroblasts into Sertoli cells (SC), which could then be deployed for different diagnostic strategies. SCs form the target cell type of choice because they act like an organizing center of embryonic gonadal development and many DSD arise when these developmental processes go awry.

We employed a computational predictive algorithm for cell conversions called Mogrify to predict the transcription factors (TFs) required for direct reprogramming of human dermal fibroblasts into SCs. We established trans-differentiation culture conditions where stable transgenic expression of these TFs was achieved in 46, XY adult dermal fibroblasts using lentiviral vectors. The resulting Sertoli like cells (SLCs) were validated for SC phenotype using several approaches. SLCs exhibited Sertoli-like morphological and cellular properties as revealed by morphometry and xCelligence cell behavior assays. They also showed Sertoli-specific expression of molecular markers such as SOX9, PTGDS, BMP4, or DMRT1 as revealed by IF imaging, RNAseq and qPCR. The SLC transcriptome shared about two thirds of its differentially expressed genes with a human adult SC transcriptome and expressed markers typical of embryonic SCs. Notably, SLCs lacked expression of markers of other gonadal cell types such as Leydig, germ, peritubular myoid or granulosa cells.

The trans-differentiation method was applied to a variety of commercially available 46, XY fibroblasts derived from patients with DSD and to a 46, XX cell line. The DSD SLCs displayed altered levels of trans-differentiation in comparison to normal 46, XY-derived SLCs, thus showcasing the robustness of this new trans-differentiation model.
Assessment of anti-inflammatory bioactivity of extracellular vesicles is susceptible to error via media component contamination
Authors: Kronstadt Stephanie M. , Van Heyningen Lauren Hoorens , Aranda Amaya , Jay Steven M.
Published: August 2022

Extracellular vesicles (EVs) are widely implicated as novel diagnostic and therapeutic modalities for a wide range of diseases. Thus, optimization of EV biomanufacturing is of high interest. In the course of developing parameters for a HEK293T EV production platform, we examined the combinatorial effects of cell culture conditions (i.e., static vs dynamic) and isolation techniques (i.e., ultracentrifugation vs tangential flow filtration vs size-exclusion chromatography) on functional characteristics of HEK293T EVs, including anti-inflammatory bioactivity using a well-established LPS-stimulated mouse macrophage model. We unexpectedly found that, depending on culture condition and isolation strategy, HEK293T EVs appeared to significantly suppress the secretion of pro-inflammatory cytokines (i.e., IL-6, RANTES) in the stimulated mouse macrophages. Further examination revealed that these results were most likely due to fetal bovine serum (FBS) EV contamination in HEK293T EV preparations. Thus, future research assessing the anti-inflammatory effects of EVs should be designed to account for this phenomenon.
Learning-dependent evolution of odor mixture representations in piriform cortex
Authors: Berners-Lee Alice , Shtrahman Elizabeth , Grimaud Julien , Murthy Venkatesh N.
Published: August 2022

Rodents can learn from exposure to rewarding odors to make better and quicker decisions. The piriform cortex is thought to be important for learning complex odor associations, however it is not understood exactly how it learns to remember discriminations between many, sometimes overlapping, odor mixtures. We investigated how odor mixtures are represented in the posterior piriform cortex (pPC) of mice while they learn to discriminate a unique target odor mixture against hundreds of nontarget mixtures. We find that a significant proportion of pPC neurons discriminate between the target and all other nontarget odor mixtures. Neurons that prefer the target odor mixture tend to respond with brief increases in firing rate at odor onset compared to other neurons, which exhibit sustained and/or decreased firing. We allowed mice to continue training after they had reached high levels of performance and find that pPC neurons become more selective for target odor mixtures as well as for randomly chosen repeated nontarget odor mixtures that mice did not have to discriminate from other nontargets. These single unit changes during overtraining are accompanied by better categorization decoding at the population level, even though behavioral metrics of mice such as reward rate and latency to respond do not change. However, when difficult ambiguous trial types are introduced, the robustness of the target selectivity is correlated with better performance on the difficult trials. Taken together, these data reveal pPC as a dynamic and robust system that can optimize for both current and possible future task demands at once.
Stable kinetochore-microtubule attachment requires loop-dependent Ndc80-Ndc80 binding
Authors: Polley Soumitra , Müschenborn Helen , Terbeck Melina , De Antoni Anna , Vetter Ingrid R. , Dogterom Marileen , Musacchio Andrea , Volkov Vladimir A. , Huis in ‘t Veld Pim J.
Published: August 2022

During cell division, kinetochores link chromosomes to spindle microtubules. The Ndc80 complex, a crucial microtubule binder, populates each kinetochore with dozens of copies. Whether adjacent Ndc80 complexes cooperate to promote microtubule binding remains unclear. Here we demonstrate that the Ndc80 loop, a short sequence identified across eukaryotes and predicted to interrupt the Ndc80 coiled-coil, promotes direct interactions between full-length Ndc80 complexes. Both in dividing cells and in a fully reconstituted system, these interactions are essential for the formation of force-resistant kinetochore-microtubule attachments, explaining why deletion of the loop ablates chromosome congression. The loop may fold into a more rigid structure than previously assumed and we identify point mutations that impair Ndc80-Ndc80 interactions and recapitulate the effects of loop depletion. The loop also promotes spindle checkpoint signaling, suggesting that the organisation of adjacent Ndc80 complexes is crucial to couple kinetochore-microtubule binding to cell cycle control.
Genome-wide prediction of disease variants with a deep protein language model
Authors: Brandes Nadav , Goldman Grant , Wang Charlotte H. , Ye Chun Jimmie , Ntranos Vasilis
Published: August 2022

Distinguishing between damaging and neutral missense variants is an ongoing challenge in human genetics, with profound implications for clinical diagnosis, genetic studies and protein engineering. Recently, deep-learning models have achieved state-of-the-art performance in classifying variants as pathogenic or benign. However, these models are currently unable to provide predictions over all missense variants, either because of dependency on close protein homologs or due to software limitations. Here we leveraged ESM1b, a 650M-parameter protein language model, to predict the functional impact of human coding variation at scale. To overcome existing technical limitations, we developed a modified ESM1b workflow and functionalized, for the first time, all proteins in the human genome, resulting in predictions for all ∼450M possible missense variant effects. ESM1b was able to distinguish between pathogenic and benign variants across ∼150K variants annotated in ClinVar and HGMD, outperforming existing state-of-the-art methods. ESM1b also exceeded the state of the art at predicting the experimental results of deep mutational scans. We further annotated ∼2M variants across ∼9K alternatively-spliced genes as damaging in certain protein isoforms while neutral in others, demonstrating the importance of considering all isoforms when functionalizing variant effects. The complete catalog of variant effect predictions is available at: https://huggingface.co/spaces/ntranoslab/esm_variants.
Fgf8 promotes survival of nephron progenitors by regulating BAX/BAK-mediated apoptosis
Authors: Anderson Matthew J. , Misaghian Salvia , Sharma Nirmala , Perantoni Alan O. , Lewandoski Mark
Published: August 2022

Fibroblast growth factors (Fgfs) have long been implicated in processes critical to embryonic development, such as cell survival, migration, and differentiation. Several mouse models of organ development ascribe a prosurvival requirement specifically to FGF8. Here, we explore the potential role of prosurvival FGF8 signaling in kidney development. We have previously demonstrated that conditional deletion of Fgf8 in the mesodermal progenitors that give rise to the kidney leads to renal aplasia in the mutant neonate. Deleterious consequences caused by loss of FGF8 begin to manifest by E14.5 when massive aberrant cell death occurs in the cortical nephrogenic zone in the rudimentary kidney as well as in the renal vesicles that give rise to the nephrons. To rescue cell death in the Fgf8 mutant kidney, we inactivate the genes encoding the pro-apoptotic factors BAK and BAX. In a wild-type background, the loss of Bak and Bax abrogates normal cell death and has minimal effect on renal development. However, in Fgf8 mutants, the combined loss of Bak and Bax rescues aberrant cell death in the kidneys and restores some measure of kidney development: 1) the nephron progenitor population is greatly increased; 2) some glomeruli form, which are rarely observed in Fgf8 mutants; and 3) kidney size is rescued by about 50% at E18.5. The development of functional nephrons, however, is not rescued. Thus, FGF8 signaling is required for nephron progenitor survival by regulating BAK / BAX and for subsequent steps involving, as yet, undefined roles in kidney development.
Inventa: a computational tool to discover chemical novelty in natural extracts libraries
Authors: Quiros-Guerrero Luis-Manuel , Nothias Louis-Félix , Gaudry Arnaud , Marcourt Laurence , Allard Pierre-Marie , Rutz Adriano , David Bruno , Queiroz Emerson Ferreira , Wolfender Jean-Luc
Published: August 2022

Collections of natural extracts hold potential for the discovery of novel natural products with original modes of action. The prioritization of extracts from collections remains challenging due to the lack of workflow that combines multiple-source information to facilitate the data interpretation. Results from different analysis techniques and literature reports need to be organized, processed, and interpreted to enable optimal decision-making for extracts prioritization. Here, we introduce Inventa, a computational tool that highlights the chemical novelty potential within extracts, considering untargeted mass spectrometry data, spectral annotation, and literature reports. Based on this information, Inventa calculates multiple scores that inform their chemical potential. Thus, Inventa has the potential to accelerate new natural products discovery. Inventa was applied to a set of plants from the Celastraceae family as a proof of concept. The Pristimera indica (Willd.) A.C.Sm roots extract was highlighted as a promising source of potentially novel compounds. Its phytochemical investigation resulted in the isolation and de novo characterization of thirteen new dihydro-β-agarofuran sesquiterpenes, five of them presenting a new 9-oxodihydro-β-agarofuran base scaffold.
Dispersal inference from population genetic variation using a convolutional neural network
Authors: Smith Chris C. R. , Tittes Silas , Ralph Peter L. , Kern Andrew D.
Published: August 2022

The geographic nature of biological dispersal shapes patterns of genetic variation over landscapes, making it possible to infer properties of dispersal from genetic variation data. Here we present an inference tool that uses geographically distributed genotype data in combination with a convolutional neural network to estimate a critical population parameter: the mean per-generation dispersal distance. Using extensive simulation, we show that our deep learning approach is competitive with or outperforms state-of-the-art methods, particularly at small sample sizes. In addition, we evaluate varying nuisance parameters during training—including population density, demographic history, habitat size, and sampling area—and show that this strategy is effective for estimating dispersal distance when other model parameters are unknown. Whereas competing methods depend on information about local population density or accurate identification of identity-by-descent tracts, our method uses only single-nucleotide-polymorphism data and the spatial scale of sampling as input. Strikingly, and unlike other methods, our method does not use the geographic coordinates of the genotyped individuals. These features make our method, which we call “disperseNN”, a potentially valuable new tool for estimating dispersal distance in non-model systems with whole genome data or reduced representation data. We applydisperseNNto 12 different species with publicly available data, yielding reasonable estimates for most species. Importantly, our method estimated consistently larger dispersal distances than mark-recapture calculations in the same species, which may be due to the limited geographic sampling area covered by some mark-recapture studies. Thus genetic tools like ours complement direct methods for improving our understanding of dispersal.