bluuebunny/crossref_metadata_2025 huggingface dataset:
Improving laser standards for three-photon microscopy
Authors: Farinella Deano M. , Roy Arani , Liu Chao J. , Kara Prakash
Published: September 2020

Significance

Three-photon excitation microscopy has double-to-triple the penetration depth in biological tissue over two-photon imaging and thus has the potential to revolutionize the visualization of biological processesin vivo. However, unlike the ‘plug-and-play’ operation and performance of lasers used in two-photon imaging, three-photon microscopy presents new technological challenges that require a closer look at the fidelity of laser pulses.

Aim

We implemented state-of-the-art pulse measurements and developed new techniques for examining the performance of lasers used in three-photon microscopy. We then demonstrated how these techniques can be used to provide precise measurements of pulse shape, pulse energy and pulse-to-pulse intensity variability, all of which ultimately impact imaging.

Approach

We built inexpensive tools, e.g., a second harmonic generation frequency resolved optical gating (SHG-FROG) device, and a deep-memory diode imaging (DMDI) apparatus, to examine laser pulse fidelity.

Results

First, SHG-FROG revealed very large third order dispersion (TOD). This extent of phase distortion prevents the efficient temporal compression of laser pulses to their theoretical limit. Furthermore, TOD cannot be quantified when using a conventional method of obtaining the laser pulse duration, e.g., when using an autocorrelator. Finally, DMDI showed the effectiveness of detecting pulse-to-pulse intensity fluctuations on timescales relevant to three-photon imaging, which were otherwise not captured using conventional instruments and statistics.

Conclusions

The distortion of individual laser pulses caused by TOD poses significant challenges to three-photon imaging by preventing effective compression of laser pulses and decreasing the efficiency of nonlinear excitation. Moreover, an acceptably low pulse-to-pulse amplitude variability should not be assumed. Particularly for low repetition rate laser sources used in three-photon microscopy, pulse-to-pulse variability also degrades image quality. If three-photon imaging is to become mainstream, our diagnostics may be used by laser manufacturers to improve system design and by end-users to validate the performance of their current and future imaging systems.
GiantGALgene clusters for the melibiose-galactose pathway inTorulaspora
Authors: Venkatesh Anjan , Murray Anthony L. , Coughlan Aisling Y. , Wolfe Kenneth H.
Published: September 2020

In many yeast species the three genes at the center of the galactose catabolism pathway,GAL1,GAL10andGAL7, are neighbors in the genome and form a metabolic gene cluster. We report here that some yeast strains in the genusTorulasporahave much largerGALclusters that include genes for melibiase (MEL1), galactose permease (GAL2), glucose transporter (HGT1), phosphoglucomutase (PGM1), and the transcription factorGAL4, in addition toGAL1,GAL10, andGAL7. Together, these 8 genes encode almost all the steps in the pathway for catabolism of extracellular melibiose (a disaccharide of galactose and glucose). We show that a progenitor 5-gene cluster containingGAL 7-1-10-4-2was likely present in the common ancestor ofTorulasporaandZygotorulaspora. It addedPGM1andMEL1in the ancestor of mostTorulasporaspecies. It underwent further expansion in theT. pretoriensisclade, involving the fusion of three progenitor clusters in tandem and the gain ofHGT1. These giantGALclusters are highly polymorphic in structure, and subject to horizontal transfers, pseudogenization and gene losses. We identify recent horizontal transfers of completeGALclusters fromT. franciscaeinto one strain ofT. delbrueckii, and from a relative ofT. maleeaeinto one strain ofT. globosa. The variability and dynamic evolution ofGALclusters inTorulasporaindicates that there is strong natural selection on theGALpathway in this genus.
Migration through a major Andean ecogeographic disruption as a driver of genotypic and phenotypic diversity in a wild tomato species
Authors: Landis Jacob B. , Miller Christopher M. , Broz Amanda K. , Bennett Alexandra A. , Carrasquilla-Garcia Noelia , Cook Douglas R. , Last Robert L. , Bedinger Patricia A. , Moghe Gaurav D.
Published: September 2020

The large number of species on our planet arises from the phenotypic variation and reproductive isolation occurring at the population level. In this study, we sought to understand the origins of such population-level variation in defensive acylsugar chemistry and mating systems inSolanum habrochaites– a wild tomato species found in diverse Andean habitats in Ecuador and Peru. Using Restriction-Associated-Digestion Sequencing (RAD-seq) of 50S. habrochaitesaccessions, we identified eight population clusters generated via isolation and hybridization dynamics of 4-6 ancestral populations. Estimation of heterozygosity, fixation index, isolation by distance, and migration probabilities, allowed identification of multiple barriers to gene flow leading to the establishment of extant populations. One major barrier is the Amotape-Huancabamba Zone (AHZ) – a geographical feature in the Andes with high endemism, where the mountainous range breaks up into isolated microhabitats. The AHZ was associated with emergence of alleles for novel reproductive and acylsugar phenotypes. These alleles led to the evolution of self-compatibility in the northern populations, where alleles for novel defense-related enzyme variants were also found to be fixed. We identified geographical distance as a major force causing population differentiation in the central/southern part of the range, whereS. habrochaiteswas also inferred to have originated. Findings presented here highlight the role of the diverse ecogeography of Peru and Ecuador in generating new, reproductively isolated populations, and enhance our understanding of the microevolutionary processes that lay a path to speciation.
Automatic Traits Extraction and Fitting for Field High-throughput Phenotyping Systems
Authors: Guo Xingche , Qiu Yumou , Nettleton Dan , Yeh Cheng-Ting , Zheng Zihao , Hey Stefan , Schnable Patrick S.
Published: September 2020

High-throughput phenotyping is a modern technology to measure plant traits efficiently and in large scale by imaging systems over the whole growth season. Those images provide rich data for statistical analysis of plant phenotypes. We propose a pipeline to extract and analyze the plant traits for field phenotyping systems. The proposed pipeline include the following main steps: plant segmentation from field images, automatic calculation of plant traits from the segmented images, and functional curve fitting for the extracted traits. To deal with the challenging problem of plant segmentation for field images, we propose a novel approach on image pixel classification by transform domain neural network models, which utilizes plant pixels from greenhouse images to train a segmentation model for field images. Our results show the proposed procedure is able to accurately extract plant heights and is more stable than results from Amazon Turks, who manually measure plant heights from original images.
ARL15 modulates magnesium homeostasis through N-glycosylation of CNNMs
Authors: Zolotarov Yevgen , Ma Chao , González-Recio Irene , Hardy Serge , Franken Gijs , Uetani Noriko , Latta Femke , Kostantin Elie , Boulais Jonathan , Thibault Marie-Pier , Côté Jean-François , Moreno Irene Díaz , Quintana Antonio Díaz , Hoenderop Joost G.J. , Martínez-Cruz Luis Alfonso , Tremblay Michel L. , de Baaij Jeroen H.F.
Published: September 2020

Cyclin M (CNNM1-4) proteins maintain cellular and body magnesium (Mg2+) homeostasis. Using various biochemical approaches, we have identified members of the CNNM family as direct interacting partners of ADP-ribosylation factor-like protein 15 (ARL15), a small GTP-binding protein. ARL15 interacts with CNNMs at their carboxyl-terminal conserved cystathionine-β-synthase (CBS) domains. In silico modeling of the interaction using the reported structures of both CNNM2 and ARL15 supports that the small GTPase specifically binds the CBS1 domain. Immunocytochemical experiments demonstrate that CNNM2 and ARL15 co-localize in the kidney, with both proteins showing subcellular localization in the Golgi-apparatus. Most importantly, we found that ARL15 is required for forming complex N-glycosylation of CNNMs. Overexpression of ARL15 promotes complex N-glycosylation of CNNM3. Mg2+ uptake experiments with a stable isotope demonstrate that there is a significant increase of 25Mg2+ uptake upon knockdown of ARL15 in multiple kidney cancer cell lines. Altogether, our results establish ARL15 as a novel negative regulator of Mg2+ transport by promoting the complex N-glycosylation of CNNMs.
Allele-specific gene regulation by KDM6A
Authors: Ma Wenxiu , Fang He , Pease Nicolas , Filippova Galina N. , Disteche Christine M. , Berletch Joel B.
Published: September 2020

KDM6A demethylates the repressive histone mark H3K27me3 and thus plays an important role in developmental gene regulation. KDM6A expression is female-biased due to escape from X inactivation, suggesting that this protein may play a role in sex differences. Here, we report that maternal and paternal alleles of a subset of mouse genes are differentially regulated by KDM6A. Knockouts ofKdm6ain male and female embryonic stem cells derived from F1 hybrid mice from reciprocal interspecific crosses resulted in preferential downregulation of maternal alleles of a number of genes implicated in development. Moreover, the majority of these genes exhibited a maternal allele expression bias, which was observed in both reciprocal crosses. Promoters of genes downregulated on maternal but not paternal alleles demonstrated a loss of chromatin accessibility, while the expected increase in H3K27me3 levels occurred only at promoters of genes downregulated on paternal but not maternal alleles. These results illustrate parent-of-origin mechanisms of gene regulation by KDM6A, consistent with histone demethylation-dependent and -independent activities.
American mammals susceptibility to dengue according to geographical, environmental and phylogenetic distances
Authors: Robles-Fernández Ángel L. , Santiago-Alarcon Diego , Lira-Noriega Andrés
Published: September 2020

Many human emergent and re-emergent diseases have a sylvatic cycle. Yet, little effort has been put into discovering and modeling the wild mammal reservoirs of dengue (DENV), particularly in the Americas. Here, we show a species-level susceptibility prediction to dengue of wild mammals in the Americas as a function of the three most important biodiversity dimensions (ecological, geographical, and phylogenetic spaces), using machine learning protocols. Model predictions showed that different species of bats would be highly susceptible to DENV infections, where susceptibility mostly depended on phylogenetic relationships among hosts and their environmental requirement. Mammal species predicted as highly susceptible coincide with sets of species that have been reported infected in field studies, but it also suggests other species that have not been previously considered or that have been captured in low numbers. Also, the environment (i.e., the distance between the species’ optima in bioclimatic dimensions) in combination with geographic and phylogenetic distance is highly relevant in predicting susceptibility to DENV in wild mammals. Our results agree with previous modeling efforts indicating that temperature is an important factor determining DENV transmission, and provide novel insights regarding other relevant factors and the importance of considering wild reservoirs. This modeling framework will aid in the identification of potential DENV reservoirs for future surveillance efforts.
Assembly of a double-stranded RNA synthesizing complex: RNA-DEPENDENT RNA POLYMERASE 2 docks with NUCLEAR RNA POLYMERASE IV at the clamp domain
Authors: Mishra Vibhor , Singh Jasleen , Fukudome Akihito , Wang Feng , Zhang Yixiang , Takagi Yuichiro , Trinidad Jonathan C , Pikaard Craig S
Published: September 2020

In plants, transcription of selfish genetic elements such as transposons and DNA viruses is suppressed by RNA-directed DNA methylation. This process is guided by 24 nt short-interfering RNAs (siRNAs) whose double-stranded precursors are synthesized by DNA-dependent NUCLEAR RNA POLYMERASE IV (Pol IV) and RNA-DEPENDENT RNA POLYMERASE 2 (RDR2). Pol IV and RDR2 co-immunoprecipitate, and their activities are tightly coupled, yet the basis for their association is unknown. Here, we show that RDR2 stably associates with Pol IV’s largest catalytic subunit, NRPD1 at three sites, all within the clamp module. The clamp is a ubiquitous feature of DNA-dependent RNA polymerases that opens to allow DNA template entry and closes to encase the DNA-RNA hybrid adjacent to the RNA exit channel. The clamp also provides binding sites for polymerase-specific subunits or regulatory proteins, thus RDR2 binding to the Pol IV clamp is consistent with this theme. Within RDR2, the site of interaction with NRPD1 is very near the catalytic center. The locations of the NRPD1-RDR2 contact sites suggest a model in which transcripts emanating from Pol IV’s RNA exit channel align with the template cleft of RDR2, facilitating rapid conversion of terminated Pol IV transcripts into double-stranded RNAs.

Significance Statement

Short interfering RNAs (siRNAs) play important roles in gene regulation by inhibiting mRNA translation into proteins or by guiding chromatin modifications that inhibit gene transcription. In plants, transcriptional gene silencing is guided by siRNAs derived from double-stranded (ds) RNAs generated by coupling the activities of DNA-dependent NUCLEAR RNA POLYMERASE IV and RNA-DEPENDENT RNA POLYMERASE 2. We show that the physical basis for Pol IV-RDR2 coupling is RDR2 binding to the clamp domain of Pol IV’s largest subunit. The positions of the protein docking sites suggest that nascent Pol IV transcripts are generated in close proximity to RDR2’s catalytic site, enabling rapid conversion of Pol IV transcripts into dsRNAs.
Different force fields give rise to different amyloid aggregation pathways in molecular dynamics simulations
Authors: Samantray Suman , Yin Feng , Kav Batuhan , Strodel Birgit
Published: September 2020

The progress towards understanding the molecular basis of Alzheimers’s disease is strongly connected to elucidating the early aggregation events of the amyloid-β(Aβ) peptide. Molecular dynamics (MD) simulations provide a viable technique to study the aggregation of Aβinto oligomers with high spatial and temporal resolution. However, the results of an MD simulation can only be as good as the underlying force field. A recent study by our group showed that none of the force fields tested can distinguish between aggregation-prone and non-aggregating peptide sequences, producing the same and in most cases too fast aggregation kinetics for all peptides. Since then, new force fields specially designed for intrinsically disordered proteins such as Aβwere developed. Here, we assess the applicability of these new force fields to studying peptide aggregation using the Aβ16−22peptide and mutations of it as test case. We investigate their performance in modeling the monomeric state, the aggregation into oligomers, and the stability of the aggregation end product, i.e., the fibrillar state. A main finding is that changing the force field has a stronger effect on the simulated aggregation pathway than changing the peptide sequence. Also the new force fields are not able to reproduce the experimental aggregation propensity order of the peptides. Dissecting the various energy contributions shows that AMBER99SB-dispoverestimates the interactions between the peptides and water, thereby inhibiting peptide aggregation. More promising results are obtained with CHARMM36m and especially its version with increased protein–water interactions. It is thus recommended to use this force field for peptide aggregation simulations and base future reparameterizations on it.
Identifying protein subcellular localisation in scientific literature using bidirectional deep recurrent neural network
Authors: David Rakesh , Menezes Rhys-Joshua D. , De Klerk Jan , Castleden Ian R. , Hooper Cornelia M. , Carneiro Gustavo , Gilliham Matthew
Published: September 2020

With the advent of increased diversity and scale of molecular data, there has been a growing appreciation for the applications of machine learning and statistical methodologies to gain new biological insights. An important step in achieving this aim is the Relation Extraction process which specifies if an interaction exists between two or more biological entities in a published study. Here, we employed natural-language processing (CBOW) and deep Recurrent Neural Network (bi-directional LSTM) to predict relations between biological entities that describe protein subcellular localisation in plants. We applied our system to 1700 publishedArabidopsisprotein subcellular studies from the SUBA manually curated dataset. The system was able to extract relevant text and the classifier predicted interactions between protein name, subcellular localisation and experimental methodology. It obtained a final precision, recall rate, accuracy and F1 scores of 0.951, 0.828, 0.893 and 0.884 respectively. The classifier was subsequently tested on a similar problem in crop species (CropPAL) and demonstrated a comparable accuracy measure (0.897). Consequently, our approach can be used to extract protein functional features from unstructured text in the literature with high accuracy. The developed system will improve dissemination or protein functional data to the scientific community and unlock the potential of big data text analytics for generating new hypotheses from diverse datasets.
Disease Network Delineates the Disease Progression Profile of Cardiovascular Diseases
Authors: Tang Zefang , Yu Yiqin , Ng Kenney , Sow Daby , Hu Jianying , Mei Jing
Published: September 2020

As Electronic Health Records (EHR) data accumulated explosively in recent years, the tremendous amount of patient clinical data provided opportunities to discover real world evidence. In this study, a graphical disease network, named progressive cardiovascular disease network (progCDN), was built based on EHR data from 14.3 million patients1to delineate the progression profiles of cardiovascular diseases (CVD). The network depicted the dominant diseases in CVD development, such as the heart failure and coronary arteriosclerosis. Novel progression relationships were also discovered, such as the progression path from long QT syndrome to major depression. In addition, three age-group progCDNs identified a series of age-associated disease progression paths and important successor diseases with age bias. Furthermore, we extracted a list of salient features to build a series of disease risk models based on the progression pairs in the disease network. The progCDN network can be further used to validate or explore novel disease relationships in real world data. Features with sufficient abundance and high correlation can be widely applied to train disease risk models when using EHR data.
A detailed mathematical theory of thalamic and cortical microcircuits based on inference in a generative vision model
Authors: George Dileep , Lázaro-Gredilla Miguel , Lehrach Wolfgang , Dedieu Antoine , Zhou Guangyao
Published: September 2020

Understanding the information processing roles of cortical circuits is an outstanding problem in neuroscience and artificial intelligence. Theory-driven efforts will be required to tease apart the functional logic of cortical circuits from the vast amounts of experimental data on cortical connectivity and physiology. Although the theoretical setting of Bayesian inference has been suggested as a framework for understanding cortical computation, making precise and falsifiable biological mappings need models that tackle the challenge of real world tasks. Based on a recent generative model, Recursive Cortical Networks, that demonstrated excellent performance on visual task benchmarks, we derive a family of anatomically instantiated and functional cortical circuit models. Efficient inference and generalization guided the representational choices in the original computational model. The cortical circuit model is derived by systematically comparing the computational requirements of this model with known anatomical constraints. The derived model suggests precise functional roles for the feed-forward, feedback, and lateral connections observed in different laminae and columns, assigns a computational role for the path through the thalamus, predicts the interactions between blobs and inter-blobs, and offers an algorithmic explanation for the innate inter-laminar connectivity between clonal neurons within a cortical column. The model also explains several visual phenomena, including the subjective contour effect, and neon-color spreading effect, with circuit-level precision. Our work paves a new path forward in understanding the logic of cortical and thalamic circuits.
T cell competition profoundly reduces the effect of initial precursor frequency on the generation of CD4 T cell memory
Authors: Smith Adrian L. , de St. Groth Barbara Fazekas
Published: September 2020

Protective immune responses are accompanied by increases in the frequency of high affinity T cells, which contribute to subsequent immunological memory. There is evidence that the fold-change in T cell number during the immune response is inversely related to initial precursor frequency, but the size of this effect remains poorly defined. Indeed, in many reports precursor frequency has been considered as directly proportional to the magnitude of the response. We have determined the effect of initial precursor frequency over the course of an in vivo antigen-specific response, in an experimental setting in which the other variables, TCR affinity and antigen dose, are kept constant. A major effect of precursor frequency was apparent in both the expansion and contraction phases; low initial precursor frequency in the physiological range was associated with greater initial expansion in T cell numbers, and also with preferential retention of memory cells. The effect was seen continuously across a 1000-fold naïve cell frequency range, leading to memory cell frequencies that differed by only 3-fold. These results are consistent with the existence of ongoing competition for antigen throughout the course of the immune response and explain the paradoxical ability of populations of genetically diverse individuals to make appropriate protective immune responses despite the large differences in initial repertoire that result from semi-random thymic TCR repertoire generation and selection.
The effects of seasonal anoxia on the microbial community structure in demosponges in a marine lake (Lough Hyne, Ireland)
Authors: Schuster Astrid , Strehlow Brian William , Eckford-Soper Lisa , McAllen Rob , Canfield Donald Eugene
Published: September 2020

Climate change is expanding marine oxygen minimum zones (OMZs), while anthropogenic nutrient input depletes oxygen concentrations locally. The effects of deoxygenation on animals are generally detrimental; however, some sponges (Porifera) exhibit hypoxic and anoxic tolerance through currently unknown mechanisms. Sponges harbor highly specific microbiomes, which can include microbes with anaerobic capabilities. Sponge-microbe symbioses must also have persisted through multiple anoxic/hypoxic periods throughout Earth history. Since sponges lack key components of the hypoxia-inducible factor (HIF) pathway responsible for hypoxic responses in other animals, it was hypothesized that sponge tolerance to deoxygenation may be facilitated by its microbiome. To test this hypothesis, we determined the microbial composition of sponge species tolerating seasonal anoxia and hypoxiain situin a semi-enclosed marine lake, using 16S rRNA amplicon sequencing. We discovered a high degree of cryptic diversity among sponge species tolerating seasonal deoxygenation, including at least nine encrusting species of the orders Axinellida and Poecilosclerida. Despite significant changes in microbial community structure in the water, sponge microbiomes were species specific and remarkably stable under varied oxygen conditions, though some symbiont sharing occurred under anoxia. At least three symbiont combinations, all including large populations ofThaumarchaeota, corresponded with deoxygenation tolerance, and some combinations were shared between distantly related hosts. We propose hypothetical host-symbiont interactions following deoxygenation that could confer deoxygenation tolerance.

Importance

The oceans have an uncertain future due to anthropogenic stressors and an uncertain past that is becoming clearer with advances in biogeochemistry. Both past and future oceans were, or will be, deoxygenated compared to present conditions. Studying how sponges and their associated microbes tolerate deoxygenation provides insights into future marine ecosystems. Moreover, sponges form the earliest branch of the animal evolutionary tree and they likely resemble some of the first animals. We determined the effects of variable environmental oxygen concentrations on the microbial communities of several demosponge species during seasonal anoxia in the field. Our results indicate that anoxic tolerance in some sponges may depend on their symbionts, but anoxic tolerance was not universal in sponges. Therefore, some sponge species could likely outcompete benthic organisms like corals in future, reduced-oxygen ecosystems. Our results support the molecular evidence that sponges and other animals have a Neoproterozoic origin, and that animal evolution was not limited by low-oxygen conditions.
Transcriptomic, protein-DNA interaction, and metabolomic studies of VosA, VelB, and WetA inAspergillus nidulansasexual spores
Authors: Wu Ming-Yueh , Mead Matthew E. , Lee Mi-Kyung , Neuhaus George F. , Adpressa Donovon A. , Martien Julia I. , Son Ye-Eun , Moon Heungyun , Amador-Noguez Daniel , Han Kap-Hoon , Rokas Antonis , Loesgen Sandra , Yu Jae-Hyuk , Park Hee-Soo
Published: September 2020

In filamentous fungi, asexual development involves morphological differentiation and metabolic changes leading to the formation of asexual spores. The process of asexual spore formation inAspergillusis precisely regulated by multiple transcription factors (TFs), including VosA, VelB, and WetA, and these three TFs are key regulators of the formation and maturation of asexual spores (conidia) inAspergillusincluding the model fungusAspergillus nidulans. To gain a mechanistic insight on the complex regulatory roles of these TFs in asexual spores, we conducted genome-wide studies on the expression, protein-DNA interactions, and primary and secondary metabolism employingA. nidulansconidia. RNA sequencing and chromatin immunoprecipitation-sequencing data have revealed that the three TFs directly or indirectly regulate the expression of genes associated with spore-wall formation/integrity, asexual development, and secondary metabolism. In addition, metabolomics analyses of wild-type and mutant conidia indicate that these three TFs regulate a diverse array of primary and secondary metabolism. In summary, WetA, VosA, and VelB play inter-dependent and distinct roles governing morphological development and primary/secondary metabolic remodeling inAspergillusconidia.

Importance

Filamentous fungi produce a vast number of asexual spores that act as reproductive and propagator cells. These spores affect humans, due to the infectious or allergenic nature of the propagule.Aspergillusspecies produce asexual spores called conidia and their formation involves morphological development and metabolic changes, and the associated regulatory systems are coordinated by spore-specific transcription factors. To understand the underlying global regulatory programs and cellular outcomes associated with conidia formation, functional genomic and metabolomic analyses were performed in the model fungusAspergillus nidulans. Our results show that the fungus specific WetA/VosA/VelB transcription factors govern the coordination of morphological and chemical developments during sporogenesis. The results of this study provide insights into the genetic regulatory networks about how morphological developments and metabolic changes are coordinated in fungi. The findings are relevant for otherAspergillusspecies such as the major human pathogenAspergillus fumigatusand the aflatoxin-producerAspergillus flavus.
Sequencing and analysis ofArabidopsis thalianaNOR2 reveal its distinct organization and tissue-specific expression of rRNA ribosomal variants
Authors: Sims Jason , Sestini Giovanni , Elgert Christiane , von Haeseler Arndt , Schlögelhofer Peter
Published: September 2020

Despite vast differences between organisms, some characteristics of their genomes are conserved, such as the nucleolus organizing region (NOR). The NOR is constituted of multiple, highly repetitive rDNA genes, encoding the catalytic ribosomal core RNAs which are transcribed from 45S rDNA units. Their precise sequence information and organization remained uncharacterized.

We used a combination of long- and short-read sequencing technologies to assemble contigs of theArabidopsisNOR2 rDNA domain providing a first map. We identified several expressed rRNA gene variants which are integrated into translating ribosomes in a tissue-specific manner. These findings support the concept of tissue specific ribosome subpopulations that differ in their rRNA composition and provide the higher order organization of NOR2.
Network and systems based re-engineering of dendritic cells with non-coding RNAs for cancer immunotherapy
Authors: Lai Xin , Dreyer Florian S. , Cantone Martina , Eberhardt Martin , Gerer Kerstin F. , Jaitly Tanushree , Uebe Steffen , Lischer Christopher , Ekici Arif , Wittmann Jürgen , Jäck Hans-Martin , Schaft Niels , Dörrie Jan , Vera Julio
Published: September 2020

Dendritic cells (DCs) are professional antigen-presenting cells that induce and regulate adaptive immunity by presenting antigens to T cells. Due to their coordinative role in adaptive immune responses, DCs have been used as cell-based therapeutic vaccination against cancer. The capacity of DCs to induce a therapeutic immune response can be enhanced by re-wiring of cellular signalling pathways with microRNAs (miRNAs). Since the activation and maturation of DCs is controlled by an interconnected signalling network, we deploy an approach that combines RNA sequencing data and systems biology methods to delineate miRNA-based strategies that enhance DC-elicited immune responses.

Through RNA sequencing of IKKβ-matured DCs that are currently being tested in a clinical trial on therapeutic anti-cancer vaccination, we identified 44 differentially expressed miRNAs. According to a network analysis, most of these miRNAs regulate targets that are linked to immune pathways, such as cytokine and interleukin signalling. We employed a network topology-oriented scoring model to rank the miRNAs, analysed their impact on immunogenic potency of DCs, and identified dozens of promising miRNA candidates with miR-15a and miR-16 as the top ones. The results of our analysis are incorporated in a database which constitutes a tool to identify DC-relevant miRNA-gene interactions with therapeutic potential (www.synmirapy.net/dc-optimization).
Inflammatory response in hematopoietic stem and progenitor cells triggered by activating SHP2 mutations evokes blood defects
Authors: Solman Maja , Blokzijl-Franke Sasja , Piques Florian , Yan Chuan , Yang Qiqi , Strullu Marion , Kamel Sarah M. , Ak Pakize , Bakkers Jeroen , Langenau David M. , Cavé Hélène , den Hertog Jeroen
Published: September 2020

Gain-of-function mutations in the protein-tyrosine phosphatase SHP2 are the most frequently occurring mutations in sporadic juvenile myelomonocytic leukemia (JMML) and JMML-like myeloproliferative neoplasm (MPN) associated with Noonan syndrome (NS). Hematopoietic stem and progenitor cells (HSPCs) are the disease propagating cells of JMML. Here, we explored transcriptomes of HSPCs with Shp2 mutations derived from JMML patients and a novel NS zebrafish model. In addition to major NS traits, CRISPR/Cas9 knock-in Shp2D61Gmutant zebrafish recapitulated a JMML-like MPN phenotype, including myeloid lineage hyperproliferation,ex vivogrowth of myeloid colonies andin vivotransplantability of HSPCs. Single cell mRNA sequencing of HSPCs from Shp2D61Gzebrafish embryos and bulk sequencing of HSPCs from JMML patients revealed an overlapping inflammatory gene expression pattern. Strikingly, an anti-inflammatory agent rescued JMML-like MPN in Shp2D61Gzebrafish embryos. Our results indicate that a common inflammatory response was triggered in the HSPCs from sporadic JMML patients and syndromic NS zebrafish, which potentiated MPN and may represent a future target for JMML therapies.
Single-Cell RNAseq of Out-of-Thaw Mesenchymal Stromal Cells Shows Striking Tissue-of-Origin Differences and Inter-donor Cell-Cycle Variations
Authors: Medrano-Trochez Camila , Chatterjee Paramita , Pradhan Pallab , Ogle Molly E , Botchwey Edward A , Kurtzberg Joanne , Yeago Carolyn , Gibson Greg , Roy Krishnendu
Published: September 2020

Mesenchymal stromal cells (MSCs) from a variety of tissue sources are widely investigated in clinical trials, and the MSCs are often administered immediately after thawing the cryopreserved product. While previous reports have examined the transcriptome of freshly-cultured MSCs from some tissues, little is known about the single-cell transcriptomic profiles of out-of-thaw MSCs from different tissue sources. Such understanding could help determine which tissue origins and delivery methods are best suited for specific indications. Here, we characterized cryopreserved MSCs, immediately post-thaw, from bone marrow (BM) and cord tissue (CT), using single-cell RNA sequencing (scRNA-seq). We show that out-of-thaw BM-vs. CT-MSCs have significant differences in gene expression. Gene-set enrichment analyses implied divergent functional potential. In addition, we show that MSC-batches can vary significantly in cell cycle status, suggesting different proliferative vs. immunomodulatory potentials. Our results provide a comprehensive single-cell transcriptomic landscape of clinically and industrially relevant MSC products.

Highlights

Single cell gene expression comparison between Bone-marrow derived MSCs and Cord-tissue derived MSCs

Donor effects and cell heterogeneity on tissue-specific MSC gene expression

Single Cell Pooling Enhances Differential Expression Analysis for Bone marrow and Cord tissue MSC samples

Gene ontology reveals tissue specific unique molecular function and pathways
Stress Assessment of Vestibular Endurance Training for Civil Aviation Flight Students Based on EEG
Authors: Hu Haixu , Fang Zhou , Qian Zhiyu , Yao Liuye , Tao Ling , Qin Bing
Published: September 2020

Objective

The main goal of our study is to clarify the EEG characteristics of the stress response caused by vestibular endurance training under the real conditions.

Methods

Ten pilot trainees received a series of acute anti-vertigo training stimulations on the rotary ladder while recording electroencephalographic data (64 electrodes). Afterwards, the subject’s anti-vertigo ability was tested for the best performance after 1 month of training, and verifying whether it is relating to the EEG signals we collected before.

Results

(1) The absolute power ofαwaves in the C3 and C4 regions is same as the difference between 1 min before and 2 min after stimulation, and their activity is enhanced by stimulation. Otherwise, the activation of the C3 region after 5min of stimulation is still significant changed. (2) Discover a spearman rank correlation, theαwaves in the C3 and C4 the greater the power change, the better the performance of the subject in the proficient stage.

Conclusions

C3 and C4 areas are specific brain regions of the stress response of anti-vertigo endurance training, and the absolute power of theαwave can be used as a parameter for identifying the degree of motion sickness (MS). The absolute power changes ofαwaves in the C3 and C4 areas are positively correlated with their anti-vertigo potential.

Significance

The increasing of the absolute power ofαwave in the C3 and C4 is a manifestation of MS stress adaptability.
Keratin5-BMP4 mechanosignaling regulates reciprocal acetylation and methylation at H3K9 to define blastema cell remodeling during zebrafish heart regeneration
Authors: Wang Xuelong , Guo Huiping , Yu Feifei , Zheng Wei , Zhang Hui , Peng Ying , Wang Chenghui , Wei Gang , Yan Jizhou
Published: September 2020

Heart regeneration after myocardial infarction remains challenging due to scar and ischemia-reperfusion injury. Here, we show that zebrafish blastema regeneration can effectively resalvage the wound myocardium and blood clot through cytoplasmic exocytosis and nuclear reorganization. The cell remodeling process are also visualized by spatiotemporal expression of three core blastema genes: alpha-SMA- which marks for fibrogenesis, Flk1for angiogenesis/hematopoiesis, Pax3a for remusculogensis, and by characteristic chromatin depositions of H3K9Ac/H3K9Me3. Genome-wide enhancer identification links the depositions of the two histone marks to the chromatin state and these three core blastema cell phenotypes. When the blastema subcellular fractions are introduced into the cultured zebrafish embryonic fibroblasts the altered transcription profile is comparable to the blastema transcription in terms of extracellular matrix structural constituent, vasculature development/angiogenesis, and cardiac muscle regeneration. From the subcellular fractions we identify 15 extracellular components and intermediate filaments, and show that introduction of human Krt5 and noggin peptides conversely regulate PAC2 cells F-actin reorganization, chromatin depositions of H3K9Ac/ H3K9Me3 and phosphorylation of Smad, which are accompanied by characteristic transcriptions ofbmp, bmp4, three core blastema genes as well as specific histone acetylation/methylation-related genes. Collectively, this study establishes a new Krt5-BMP4 mechanosignaling cascade that links extracellular molecules to chromatin modifications and regulates blastema cell remodeling, thus providing mechanistic insights into the mesoderm-derived blastema regeneration and underlining a therapy strategy for myocardial infarction.
DTFLOW: Inference and Visualization of Single-cell Pseudo-temporal Trajectories Using Diffusion Propagation
Authors: Wei Jiangyong , Zhou Tianshou , Zhang Xinan , Tian Tianhai
Published: September 2020

One of the major challenges in single-cell data analysis is the determination of cellular developmental trajectories using single-cell data. Although substantial studies have been conducted in recent years, more effective methods are still strongly needed to infer the developmental processes accurately. In this work we devise a new method, named DTFLOW, for determining the pseudo-temporal trajectories with multiple branches. This method consists of two major steps: namely a new dimension reduction method (i.e. Bhattacharyya kernel feature decomposition (BKFD)) and a novel approach, named Reverse Searching on kNN Graph (RSKG), to identify the underlying multi-branching processes of cellular differentiations. In BKFD we first establish a stationary distribution for each cell to represent the transition of cellular developmental states based on the random walk with restart algorithm and then propose a new distance metric for calculating pseudo-times of single-cells by introducing the Bhattacharyya kernel matrix. The effectiveness of DTFLOW is rigorously examined by using four single-cell datasets. We compare the efficiency of the new method with two state-of-the-art methods. Simulation results suggest that our proposed method has superior accuracy and strong robustness properties for constructing pseudo-time trajectories. Availability: DTFLOW is implemented in Python and available athttps://github.com/statway/DTFLOW.
FMRP sustains presynaptic function via control of activity-dependent bulk endocytosis
Authors: Bonnycastle Katherine , Kind Peter C. , Cousin Michael A.
Published: September 2020

Synaptic vesicle (SV) recycling defects are linked to neurodevelopmental disorders, including fragile X syndrome (FXS), which results from loss of fragile X mental retardation protein (FMRP) encoded by the FMR1 gene. Hyperexcitability of neuronal circuits is a key feature of FXS, therefore we investigated whether SV recycling was affected by the absence of FMRP during increased neuronal activity. We revealed that primary neuronal cultures from a Fmr1 knockout rat model display a specific defect in activity-dependent bulk endocytosis (ADBE). This defect resulted in an inability of Fmr1 knockout neurons to sustain SV recycling during high frequency stimulation. Using a molecular replacement strategy, we also revealed that a human FMRP mutant that cannot bind BK channels failed to correct ADBE dysfunction in knockout neurons, however this dysfunction was corrected by BK channel agonists. Therefore, FMRP performs a key role in sustaining neurotransmitter release via selective control of ADBE, suggesting intervention via this endocytosis mode may correct hyperexcitabiltiy observed in FXS.

Fragile X syndrome (FXS) is caused by loss of fragile X mental retardation protein (FMRP). Bonnycastle et al show that FMRP is specifically required for activity-dependent bulk endocytosis (ADBE), revealing 1) FMRP sustains neurotransmitter release and 2) intervention via ADBE may correct circuit hyperexcitabilty in FXS.
Vanilloids HamperCaenorhabditis elegansResponse to Noxious Heat
Authors: Nkambeu Bruno , Salem Jennifer Ben , Beaudry Francis
Published: September 2020

Eugenol, a known vanilloid, was frequently used in dentistry as a local analgesic in addition, antibacterial and neuroprotective effects were also reported. Eugenol, capsaicin and many vanilloids are interacting with the transient receptor potential vanilloid 1 (TRPV1) in mammals and are activated by noxious heat. The pharmacological manipulation of the TRPV1 has been shown to have therapeutic value.Caenorhabditis elegans(C. elegans) express TRPV orthologs (e.g. OCR-2, OSM-9) and it is a commonly used animal model system to study nociception as it displays a well-defined and reproducible nocifensive behavior. After exposure to vanilloid solutions,C. eleganswild type (N2) and mutants were placed on petri dishes divided in quadrants for heat stimulation. Thermal avoidance index was used to phenotype each testedC. elegansexperimental groups. The results showed that eugenol, vanillin and zingerone can hamper nocifensive response ofC. elegansto noxious heat (32°C – 35°C) following a sustained exposition. Also, the effect was reversed 6h post exposition. Furthermore, eugenol and vanillin did not target specifically the OCR-2 or OSM-9 but zingerone did specifically target the OCR-2 similarly to capsaicin. Further structural and physicochemical analyses were performed. Key parameters for quantitative structure-property relationships (QSPR), quantitative structure-activity relationships (QSAR) and frontier orbital analyses suggest similarities and dissimilarities amongst the tested vanilloids and capsaicin in accordance with the relative anti-nociceptive effects observed.
Inhibition of USP28 overcomes Cisplatin-Resistance of Squamous Tumors by Suppression of the Fanconi Anemia Pathway
Authors: Prieto-Garcia Cristian , Hartmann Oliver , Reissland Michaela , Fischer Thomas , Maier Carina R. , Rosenfeldt Mathias , Schülein-Völk Christina , Klann Kevin , Kalb Reinhard , Dikic Ivan , Münch Christian , Diefenbacher Markus E.
Published: September 2020

Squamous cell carcinomas (SCC) frequently have a limited response to or develop resistance to platinum-based chemotherapy, and have an exceptionally high tumor mutational burden. As a consequence, overall survival is limited and novel therapeutic strategies are urgently required, especially in light of a rising incidences. SCC tumors express ΔNp63, a potent regulator of the Fanconi Anemia (FA) DNA-damage response pathway during chemotherapy, thereby directly contributing to chemotherapy-resistance. Here we report that the deubiquitylase USP28 affects the FA DNA repair pathway during cisplatin treatment in SCC, thereby influencing therapy outcome. In an ATR-dependent fashion, USP28 is phosphorylated and activated to positively regulate the DNA damage response. Inhibition of USP28 reduces recombinational repair via an ΔNp63-Fanconi Anemia pathway axis, and weakens the ability of tumor cells to accurately repair DNA. Our study presents a novel mechanism by which tumor cells, and in particular ΔNp63 expressing SCC, can be targeted to overcome chemotherapy resistance.

Significance

Limited treatment options and low response rates to chemotherapy are particularly common in patients with squamous cancer. The SCC specific transcription factor ΔNp63 enhances the expression of Fanconi Anemia genes, thereby contributing to recombinational DNA repair and Cisplatin resistance. Targeting the USP28-ΔNp63 axis in SCC tones down this DNA damage response pathways, thereby sensitizing SCC cells to cisplatin treatment.
Multistage feedback driven compartmental dynamics of hematopoiesis
Authors: Mon Père Nathaniel V. , Lenaerts Tom , Pacheco Jorge M. , Dingli David
Published: September 2020

Human hematopoiesis is surprisingly resilient to disruptions, providing suitable responses to severe bleeding, long lasting immune activation, and even bone marrow transplants. Still, many blood disorders exist which push the system past its natural plasticity, resulting in abnormalities in the circulating blood. While proper treatment of such diseases can benefit from understanding the underlying cell dynamics, these are non-trivial to predict due to the hematopoietic system’s hierarchical nature and complex feedback networks. To characterize the dynamics following different types of perturbations we investigate a model representing hematopoiesis as a sequence of compartments covering all maturation stages – from stem to mature cells – where feedback regulates cell production to ongoing necessities. We find that a stable response to perturbations requires the simultaneous adaptation of cell differentiation and self-renewal rates, and show that under conditions of continuous disruption – as found in chronic hemolytic states – compartment cell numbers evolve to novel stable states.
Predictive encoding of motion begins in the primate retina
Authors: Liu Belle , Hong Arthur , Rieke Fred , Manookin Michael B.
Published: September 2020

Survival in the natural environment often relies on an animal’s ability to quickly and accurately predict the trajectories of moving objects. Motion prediction is primarily understood in the context of translational motion, but the environment contains other types of behaviorally salient motion, such as that produced by approaching or receding objects. However, the neural mechanisms that detect and predictively encode these motion types remain unclear. Here, we address these questions in the macaque monkey retina. We report that four of the parallel output pathways in the primate retina encode predictive information about the future trajectory of moving objects. Predictive encoding occurs both for translational motion and for higher-order motion patterns found in natural vision. Further, predictive encoding of these motion types is nearly optimal with transmitted information approaching the theoretical limit imposed by the stimulus itself. These findings argue that natural selection has emphasized encoding of information that is relevant for anticipating future properties of the environment.