Biology subjects

Wu, Y.

Publications and source records attributed to Wu, Y..

At least 721 records · Page 40Linked to original sources

bioRxiv2022

Modulating hierarchical learning by high-definition transcranial alternating current stimulation at theta frequency

Considerable evidence highlights the dorsolateral prefrontal cortex (DLPFC) as a key region for hierarchical (i.e., multi-level) learning. In a previous electroencephalography (EEG) study we found that the low-level prediction errors (PEs) were encoded by frontal theta oscillations (4-7Hz), centered on right DLPFC (rDLPFC). However, the causal relationship between frontal theta oscillations and hierarchical learning remains poorly understood. To investigate this question, in the current study, participants received theta (6Hz) and sham high-definition transcranial alternating current stimulation (HD-tACS) over the rDLPFC, while performing the probabilistic reversal learning task. Behaviorally, theta tACS induced a significant reduction in accuracy for the stable environment, but not for the volatile environment, relative to the sham condition. Computationally, we implemented a combination of a hierarchical Bayesian learning and a decision model. Theta tACS induced a significant increase in low-level (i.e., probability-level) learning rate and uncertainty of low-level estimation relative to sham condition. Instead, the temperature parameter of the decision model, which represents (inverse) decision noise, was not significantly altered due to theta stimulation. These results indicate that theta frequency may modulate the (low-level) learning rate. Furthermore, environmental features (e.g., its stability) may determine whether learning is optimized as a result.

neuroscience↗

bioRxiv2022

In-depth investigation of microRNA-mediated cross-kingdom regulation between Asian honey bee and microsporidian

Asian honey bee Apis cerana is the original host for Nosema ceranae, a unicellular fungal parasite that causes bee nosemosis throughout the world. Currently, interaction between A. cerana and N. ceranae is largely unknown. Here, based on our previously gained high-quality RNA-seq and small RNA-seq data from N. ceranae-infected A. c. cerana workers midguts and clean spores, differentially expressed mRNAs (DEmiRNAs) in N. ceranae targeted by DEmiRNAs in host midguts and A. c. cerana DEmRNAs targeted by microsporidian DEmiRNAs were predicted using bioinformatics, and then target DEmRNAs in microsporidian and host were annotated and investigated, with a focus on targets involved in N. ceranae glycolysis/glyconeogenesis and virulence factors as well as A. c. cerana energy mechanism and immune response. Its found that 97 down-regulated (60 up-regulated) mRNAs in NcCKM vs NcTM1 were potentially targeted by eight up-regulated (six down-regulated) miRNAs in AcCKMI1 vs AcTMI1, 44 down-regulated (15 up-regulated) mRNAs in NcCKM vs NcTM2 were putative targets of seven up-regulated (two down-regulated) miRNAs in AcCKMI2 vs AcTMI2. Additionally, miR-60-y and miR-676-y were found to up-regulate in AcCKMI1 vs AcTMI1 and target genes engaged in spore wall protein and glycolysis/gluconeogenesis, while miR-60-y in AcCKMI1 vs AcTMI1 was up-regulated and potentially targeted a glycolysis/gluconeogenesis-associated gene. Comparatively, 343 down-regulated (138 up-regulated) mRNAs in AcCKM1 vs AcTM1 were putative targets of 121 up-regulated (112 down-regulated) miRNAs in NcCKMI vs NcTMI1, 247 down-regulated (110 up-regulated) mRNAs were putatively targeted by 110 up-regulated (104 down-regulated) miRNAs in NcCKMI vs NcTMI2. Further analysis showed that 31 up-regulated miRNAs in NcCKMI vs NcTMI1 potentially targeted 12 down-regulated mRNAs in AcCKM1 vs AcTM1, which were involved in five immune-related pathways such as phagasome and Jak-STAT signaling pathway, whereas nine up-regulated miRNAs in NcCKMI vs NcTMI2 putatively targeted five down-regulated mRNAs in AcCKM2 vs AcTM2, which were engaged in three immune-related pathways including endocytosis, lysosomes, and regulation of autophagy. In addition, miR-21-x was observed to up-regulate in NcCKMI vs NcTMI1 and target a oxidative phosphorylation-related gene. Finally, potential targeting relationship between two host DEmiRNAs-microsporidian DEmRNAs pairs and two microsporidian DEmiRNAs-host DEmRNAs pairs were verified on basis of RT-qPCR. Our findings not only lay a foundation for exploring the molecular mechanism underlying cross-kingdom regulation between A. c. cerana workers and N. ceranae, but also offer valuable insights into Asian honey bee-microsporidian interaction.

pathology↗

bioRxiv2022

Genome-wide determination of barriers to horizontal gene transfer

Horizontal gene transfer (HGT) is a major contributor to bacterial genome evolution, generating phenotypic diversity, driving the expansion of protein families, and facilitating the evolution of new phenotypes, new metabolic pathways, and new species. Comparative studies of gene gain in bacteria suggest that the frequency with which individual genes successfully undergo HGT varies considerably and may be associated with the number of protein-protein interactions in which the gene participates--its connectivity. Two non-exclusive hypotheses have emerged to explain why transferability should decrease with connectivity: the Complexity Hypothesis (Jain, Rivera, & Lake, 1999) and the Balance Hypothesis (Papp, Pal, & Hurst, 2003). These hypotheses predict that the functional costs of HGT arise from a failure of divergent homologues to make normal protein-protein interactions or from gene mis-expression, respectively. Here we describe genome-wide assessments of these hypotheses in which we used 74 existing prokaryotic whole genome shotgun libraries to estimate rates of horizontal transfer of genes from taxonomically diverse prokaryotic donors into E. coli. We show that transferability declines as connectivity increases, but that this effect is driven primarily by the ribosomal genes. We also show that transferability declines as the divergence (% amino acid difference) between donor and recipient orthologs increases and that this effect of divergence increases with connectivity. We explain how these results, even the stronger effect of connectivity on the transferability of ribosomal compared to non-ribosomal genes, provide strong support for both the Balance and Complexity Hypotheses. Significance StatementComparisons between prokaryotic genomes consistently show that genes with informational functions, e.g. in genome replication, transcription, and translation, have been subject to horizontal gene transfer between species more often than genes with operational functions, e.g. in metabolism and environmental sensing. In this study, we use a genome wide analysis of transferability data obtained from 74 genomes to provide the first experimental evidence that this pattern results from differences between informational and operational genes in the number of other proteins with which they interact, i.e., their connectivity, rather than from their functional differences. The importance of our exceptionally large dataset to the detection of connectivity effects on transferability explains why past experimental studies failed to replicate the consistent finding from comparative genomic studies.

genomics↗

bioRxiv2022

The Optimal Combination of Scanning Speed and Fiber-to-Stone Distance for Effective and Efficient Dusting during Holmium: YAG Laser Lithotripsy

ObjectivesTo investigate mechanistically the effects of fiber scanning speed (vfiber) and fiber tip-to-stone standoff distance (SD) on dusting efficiency during Holmium (Ho): YAG laser lithotripsy (LL) Materials and MethodsPre-soaked BegoStone samples (23 x 23 x 4 mm, W x L x H) were treated in water using a clinical Ho:YAG laser in dusting mode (0.2 J pulse energy delivered at 20 Hz frequency) at three different SDs (0.10, 0.25 and 0.50 mm) with vfiber in the range of 0 to 10 mm/s. Stone damage was quantified by optical coherence tomography, bubble dynamics were captured by high-speed imaging, and associated pressure transients were measured using a needle hydrophone. To compare photothermal ablation vs. cavitation in stone dusting, the experiments were further repeated in air (photothermal ablation only), and in water with the fiber tip advanced at a short (0.25 mm) offset distance (OSD) from a ureteroscope to mitigate the bubble collapse toward the stone surface, thus eliminating cavitation-induced damage. ResultsCompared to the craters produced by a stationary fiber, the damage troughs produced by a scanning fiber after 100 pulses were significantly larger in volume. The optimal vfiber for maximum dusting efficiency was found to be 3.5 mm/s for SD = 0.10 [~] 0.25 mm, resulting in long (17.5 mm), shallow (0.14 - 0.15 mm) and narrow (0.3 - 0.4 mm) troughs. In contrast, the maximum stone damage was produced at an optimal vfiber of 0.5 mm/s for SD = 0.50 mm, which generates much shorter (2.5 mm), yet deeper (0.35 mm) and wider (1.4 mm) troughs. Greater stone damage was produced in water than in air, especially at vfiber = 0 - 3.5 mm/s. With the scope end placed near the fiber tip, stone damage could be significantly reduced in water by 29% - 58% for SD = 0.10 mm, by 51% - 82% for SD = 0.25 mm, and by 66% - 100 % for SD = 0.50 mm, compared to those produced without the scope. Together, these findings suggest that cavitation plays an indispensable role in stone dusting by scanning treatment. Moreover, under clinically relevant vfiber (1 [~] 3 mm/s), dusting at SD = 0.5 mm (i.e., non-contact mode) may leverage higher frequency of the laser (e.g., 40 to 120 Hz) to harvest the full potential of cavitation damage while significantly reducing the procedure time, compared to its counterpart at SD = 0.1 mm (i.e., contact mode) that promotes photothermal ablation. ConclusionDusting efficiency during Ho:YAG LL may be substantially improved by utilizing the optimal combination of vfiber and SD for a given frequency.

bioengineering↗

bioRxiv2022

The mechanosensitive ion channel Piezo1 modulates the migration and immune response of microglia

Microglia are the brains resident immune cells, performing surveillance to promote homeostasis and healthy functioning. While chemical signaling in microglia is well-studied, the mechanical cues regulating their function are less well-understood. Here, we investigate the role of the mechanosensitive ion channel Piezo1 in microglia migration, pro-inflammatory cytokine production and stiffness sensing. We demonstrated the functional expression of Piezo1 in microglia and identified genes whose expression was affected by conditional Piezo1 knockout in transgenic mice. Functional assays revealed that Piezo1-deficiency in microglia enhanced migration towards amyloid {beta}-protein, and decreased levels of pro-inflammatory cytokines produced upon stimulation by lipopolysaccharide, both in vitro and in vivo. The phenomenon could be mimicked or reversed using a Piezo1-specific agonist or antagonist. Finally, we also showed that Piezo1 mediated the effect of substrate stiffness-induced migration and cytokine expression. Altogether, we show that Piezo1 is an important molecular mediator for microglia, its activation modulating their migration and immune responses.

neuroscience↗

bioRxiv2022

A pangenome analysis pipeline (PSVCP) provides insights into rice functional gene identification

BackgroundA pangenome aims to capture the complete genetic diversity within a species and reduce bias in genetic analysis inherent in using a single reference genome. However, the current linear format of most plant pangenomes limits the presentation of position information for novel sequences. Graph pangenomes have been developed to overcome this limitation. However, there is a lack of bioinformatics analysis tools for graph format genomes. ResultsTo overcome this problem, we have developed a novel pangenome construction strategy and a downstream pangenome analysis pipeline that captures position information while maintaining a linearized layout. We applied this strategy to construct a high-quality rice pangenome using 12 representative rice genomes and analyze an international rice panel with 413 diverse accessions using the pangenome reference. Our results provide insights into rice population structure and genomic diversity. Applying the pangenome for PAV-based GWAS analysis can identify causal structural variations for rice grain weight and plant height, while SNP-based GWAS can only identify approximate genomic locations. Additionally, a new locus (qPH8-1) was found to be associated with plant height on chromosome 8 that could not be detected using the SNP-based GWAS. ConclusionsOur results demonstrate that the pangenome constructed by our pipeline combined with PAV-based GWAS can provide additional power for genomic and genetic analysis. The pangenome constructed in this study and associated genome sequence data provide valuable genomic resources for future rice crop improvement.

genetics↗

bioRxiv2022

Synaptic vesicle proteins and ATG9A self-organize in distinct vesicle phases within synapsin condensates

Ectopic expression in fibroblasts of synapsin 1 and synaptophysin is sufficient to generate condensates of vesicles highly reminiscent of synaptic vesicle (SV) clusters and with liquid-like properties. We show that unlike synaptophysin, other major integral SV membrane proteins fail to form condensates with synapsin, but coassemble into the clusters formed by synaptophysin and synapsin in this ectopic expression system. Another vesicle membrane protein, ATG9A, undergoes activity-dependent exo-endocytosis at synapses, raising questions about the relation of ATG9A traffic to the traffic of SVs. We have found that both in fibroblasts and in nerve terminals ATG9A does not coassemble into synaptophysin-positive vesicle condensates but localizes on a distinct class of vesicles that also assembles with synapsin but into a distinct phase. Our findings suggest that ATG9A undergoes differential sorting relative to SV proteins and also point to a dual role of synapsin in controlling clustering at synapses of SVs and ATG9A vesicles.

neuroscience↗

bioRxiv2022

Genome-wide prediction of pathogenic gain- and loss-of-function variants from ensemble learning of diverse feature set

Gain-of-function (GOF) variants give rise to increased or novel protein functions whereas loss-of-function (LOF) variants lead to diminished protein function. GOF and LOF variants can result in markedly varying phenotypes, even when occurring in the same gene. However, experimental approaches for identifying GOF and LOF are generally slow and costly, whilst currently available computational methods have not been optimized to discriminate between GOF and LOF variants. We have developed LoGoFunc, an ensemble machine learning method for predicting pathogenic GOF, pathogenic LOF, and neutral genetic variants. LoGoFunc was trained on a broad range of gene-, protein-, and variant-level features describing diverse biological characteristics, as well as network features summarizing the protein-protein interactome and structural features calculated from AlphaFold2 protein models. We analyzed GOF, LOF, and neutral variants in terms of local protein structure and function, splicing disruption, and phenotypic associations, thereby revealing previously unreported relationships between various biological phenomena and variant functional outcomes. For example, GOF and LOF variants exhibit contrasting enrichments in protein structural and functional regions, whilst LOF variants are more likely to disrupt canonical splicing as indicated by splicing-related features employed by the model. Further, by performing phenome-wide association studies (PheWAS), we identified strong associations between relevant phenotypes and high-confidence predicted GOF and LOF variants. LoGoFunc outperforms other tools trained solely to predict pathogenicity or general variant impact for the identification of pathogenic GOF and LOF variants.

genetics↗

bioRxiv2022

Starvation Induces Diverse Autophagic Response in Cardiomyocytes and Noncardiomyocytes.

Cardiac autophagy serves as a potential mechanism to be genetically, epigenetically and pharmaceutically manipulated to prevent or alleviate pathological conditions in heart diseases. One of the complexities of this manipulation comes from the cellular heterogeneity of the heart. In this study, we assessed the dynamics of nutrient deprivation-induced autophagy activation in four major cardiac cell types in vitro and in vivo and found a major difference between cardiomyocyte and other three types of noncardiomyocyte. In comparison to the constant increase and maintenance of autophagic activity in noncardiomyocytes, cardiomyocyte undergoes rapid gain and loss of autophagic activity. This difference contributes to at least two aspects of cardiomyocyte pathophysiology: high vulnerability to nutrient deprivation-induced cell death and unresponsiveness to pharmaceutical regulation of autophagy. Our present findings support a cell type-specific targeting of autophagy for therapeutic intervention of cardiac diseases.

cell biology↗

bioRxiv2022

Redefining and subtyping of major depression based on brain functional connectivity signatures with high generalization: an ensemble hybrid framework

Current clinical diagnosis of major depression is solely based on subjective symptoms and signs rather than the underlying biological mechanisms, which has hindered the improvement in treatment effectiveness and outcomes. Redefining and subtyping major depressive disorder (MDD) based on neural circuits has been an emerging consensus. Here, we proposed a novel ensemble hybrid deep-learning framework based on graph neural network for redefining and subtyping of MDD with the brains "fingerprinting", i.e. nuanced functional connectivity profiles. A graph neural network (GNN) based supervised model was first employed to discriminate MDD patients from healthy controls. The graph embedding features obtained from the supervised classifier was then employed for subtyping of MDD with an unsupervised clustering by fast search and find of density peaks (CFDP) approach. The generalization accuracy and reproducibility of the proposed framework were further validated with one of the largest resting-state fMRI data set for MDD which included 2428 participants recruited from 25 independent sites. Our classifier showed a high generalization of 75% for leave-one-site-out cross validation. In addition, we obtained three biologically homogenous MDD subtypes characterized by impairments in subcortical-limbic, DMN-FPN, VN-SMN circuits, respectively. Notably, we demonstrated that subtyping of MDD according to graph embedding features showed high robustness and reproducibility across different sites. The proposed framework is not only essential for redefining and subtyping of MDD, but also paves the way towards precision medicine of mental disorders.

neuroscience↗

bioRxiv2022

Single cell RNA sequencing reveals mechanisms underlying a senescence-like phenotype of Alveolar Macrophages during Aging

Alveolar Macrophages (AMs) are unique innate immune cells that reside in the alveolar space and accommodate the ever-changing needs of the lungs against internal and external challenges. During homeostasis, AMs maintain themselves through self-renewal without input from adult hematopoietic stem cells. Currently, little is known regarding the influence of aging on AM dynamics, heterogeneity and transcriptional profiles. Here, we identified CBF{beta} as an indispensable transcription factor that ensures AM self-renewal. Deficiency in CBF{beta} led to decreased proliferation and self-renewal ability of AMs. Moreover, with single cell RNA sequencing analysis of AMs from young and aged mice, we discovered that despite similarities in the transcriptome of proliferating cells, AMs from the aged mice exhibited reduced embryotic stem cell-like features. Aged AMs also showed diminished capacity for DNA repair, potentially contributing to impaired cell cycle progression and elevation of senescence markers. In accordance with the analysis, we observed reduced number of AMs in aged mice, which had defective self-renewal ability and were more sensitive to the reduction of GM-CSF. Interestingly, decreased CBF{beta} was observed in the cytosol of AMs from aged mice. A similar senescence-like phenotype was also found in human AMs. Taken together, we conclude that AMs in the aged host harbor a senescence-like phenotype, potentially mediated by the activity of CBF{beta}. HighlightsO_LIscRNAseq revealed Alveolar Macrophage (AM) heterogeneity and self-renewal C_LIO_LICBF{beta} is associated with AM cell cycle and facilitate AM self-renewal C_LIO_LIAMs displayed a senescence-like phenotype during physiological Aging C_LIO_LIAging impairs CBF{beta} expression in mouse and human AMs C_LI

immunology↗

bioRxiv2022

Hippo pathway-mediated YAP1/TAZ inhibition is essential for proper pancreatic endocrine specification and differentiation

The Hippo pathway plays a central role in tissue development and homeostasis. However, the function of Hippo in pancreatic endocrine development remains obscure. Here, we examined the roles of Hippo pathway mediated YAP1/TAZ inhibition in the development stages of endocrine specification and differentiation. While YAP1 protein was localized to the nuclei in bipotent progenitor cells, Ngn3 expressing endocrine progenitors completely lost YAP1. Using genetic mouse models, we found that this inactivation of YAP1 requires both intact Hippo pathway and NGN3 protein. Deleting the Lats1 and 2 kinases (Lats1&2) in endocrine progenitor cells of developing mouse pancreas with Ngn3-Cre blocked endocrine progenitor cell differentiation and specification, resulting in reduced islets size and disorganized pancreas at birth. Loss of Lats1&2 in NGN3 expressing cells activated YAP1/TAZ transcriptional activity and recruited macrophages to the developing pancreas. These defects were rescued by deletion of YAP1/TAZ genes, suggesting that tight regulation of YAP1/TAZ by Hippo signaling is crucial for pancreatic endocrine specification. In contrast, deletion of Lats1&2 using beta-cell specific MIP1CreER resulted in a phenotypically normal pancreas, indicating that Lasts1&2 are dispensable for pancreatic beta-cell function. Our results demonstrate that YAP1/TAZ inhibition in the pancreatic endocrine compartment is not a passive consequence of endocrine specification. Rather, Hippo pathway-mediated YAP1/TAZ inhibition in endocrine progenitors is a prerequisite for endocrine specification and differentiation.

developmental biology↗

bioRxiv2022

Spatiotemporal heterogeneity and the long-term impact of meteorological , environmental and socio-economic factors of scrub typhus in China from 2012 to 2018

Large-scale outbreaks of scrub typhus combined with the emergence of this vector-borne rickettsiosis in new areas indicate that this disease remains seriously neglected. This study aimed to explore the long-term changes and regional leading factors of scrub typhus in China, so as to provide fresh insights for the prevention and control of this disease. In this study, a Bayesian space-time hierarchical model (BSTHM) was used to identify the long-term spatiotemporal heterogeneity of scrub typhus and quantify the association between meteorological factors and scrub typhus in southern and northern China from 2012 to 2018. GeoDetector model was used to quantify the dominant forces of environmental and socioeconomic factors in the Northern and the Southern China. Scrub typhus often appeared in summer and autumn (June to November), and epidemically peaked in October, with obvious temporal seasonality. Spatially, the hot spots (high-risk regions) were concentrated in the south, on the contrary the cold spots (low-risk regions) in the north. In addition, the main meteorological factor, average temperature, gave a significant impact in both areas. The average temperature increased by 1 {degrees}C, resulting in a decrease of 1.10% in southern China and an increase of 0.96% in northern China in the risk of scrub typhus. The determinant environmental and socio-economic factors of scrub typhus in the two areas were altitude and per capita GDP, with q-values of 0.91 and 0.87, respectively. Meteorological, environmental and socio-economic factors had a significant impact on the distribution of scrub typhus, with obvious seasonality and spatial heterogeneity. This study provides helpful suggestions and basis for reasonably allocating resources and controlling the occurrence of scrub typhus. Author summaryScrub typhus is a natural-focus disease caused by the bite of chigger mite larval. In this study, we use BSTHM to capture the overall temporal trend and spatial hot spots of scrub typhus, and quantify the relationship between the disease and major meteorological factors. Meanwhile, Geodetector model was used to quantify the influence of other potential risk factors and estimate the spatio-temporal heterogeneity of scrub typhus. The results showed that scrub typhus had significant seasonality, with a q value of 0.52, and spatial heterogeneity, with a q-value of 0.64. Scrub typhus mainly occurred in summer and autumn, and high-risk areas were mainly distributed in southern China (Yunnan, Hainan and Guangdong). These heterogeneity were closely related to the vector and host. Whether in the South or the north, scrub typhus was closely related to risk factors such as temperature, per capita GDP, NDVI, altitude and the percentage of children aged 0-14. These results suggest that the relevant departments should strengthen the monitoring of the ecological environment, the host and vector of Orientia tsutsugamushi, and strengthen the risk awareness, so as to prevent and control the possible increased risk of scrub typhus under these meteorological, environmental and socio-economic conditions. Considering the differences in different regions, resources should be allocated reasonably.

ecology↗

bioRxiv2022

Color-biased regions in the ventral visual pathway are food-selective

The ventral visual pathway is well known to be involved in recognizing and categorizing objects (Kanwisher and Dilks, 2013). Three color-biased areas have also been found between face and place selective areas in the ventral visual pathway (Lafer-Sousa et al., 2016). To understand the function of these color-biased areas in a region known for object recognition, we analyzed the Natural Scenes Dataset (NSD; Allen et al., 2022), a large 7T fMRI dataset from 8 participants who viewed up to 30,000 trials of images of colored natural scenes. In a whole-brain analysis, we correlated the average color saturation of the images and the voxel responses, revealing color-biased areas that diverge into two streams in the ventral visual pathway, beginning in V4 and extending medially and laterally of the Fusiform Face Area in both hemispheres. We drew regions of interest (ROIs) for the two streams and found that the images for each ROI that evoked the largest responses had certain characteristics: They contained food, contained circular objects, had higher color saturation, contained warmer hues, and had more luminance entropy. A multiple linear regression showed that the presence of food in images was the strongest predictor of voxel responses in the medial and lateral color-biased regions for all eight participants, but that color saturation also contributed independently to voxel responses. Our results show that these areas are food-selective and color biased. We suggest that these streams might be involved in using color to recognize and judge the properties of food.

neuroscience↗

bioRxiv2022

Robust and Versatile Arrayed Libraries for Human Genome-Wide CRISPR Activation, Deletion and Silencing

Arrayed CRISPR libraries extend the scope of gene-perturbation screens but require large numbers of efficacious sgRNA-expressing vectors. Using a newly invented liquid-phase plasmid cloning methodology, we constructed genome-wide arrayed libraries for human gene ablation (19,936 plasmids), activation, and epigenetic silencing (22,442 plasmids). At least 76% of each plasmid preparation encoded an intact array of 4 non-overlapping sgRNAs designed to tolerate most human DNA polymorphisms. We achieved perturbation efficacies of 75-99%, 76-92% and up to 10,000x in deletion, silencing and activation experiments, respectively. Upon conversion into massively parallel lentiviral vectors, an arrayed activation screen of 1,634 human transcription factors yielded 11 novel regulators of the cellular prion protein PrPC. Furthermore, a screen using a pooled version of the ablation library identified 5 novel modifiers of autophagy that went undetected with either of two 1sgRNA libraries. The CRISPR libraries described here represent a powerful resource for the targeted perturbation of human protein-coding genes.

genomics↗

bioRxiv2022

Site-specific validation and quantification of RNA 2'-O-methylation by qPCR with RNase H

RNA 2'-O-methylation, one of the most abundant modifications on RNAs, is crucial for diverse intracellular biological processes. In the past several years, several high-throughput screening methods have been developed, resulting in the identification of thousands of new 2'-O-methylation (Nm) sites. However, due to the high variability in these high-throughput methods, accurate and rapid low-throughput validation assays are needed to confirm and quantify the 2'-O-methylation status of screened candidate sites. Although several low-throughput Nm site detection methods have been reported, precise location and quantitative assays are still challenging to achieve. Based on the characteristic that RNase H would be inhibited by Nm modification, we developed Nm-VAQ (site-specific 2'-O-methylation (Nm) Validation and Absolute Quantification resolution). In this study, with multiple tests of reagents and conditions, Nm-VAQ was established with a chimera probe of RNA/DNA, RNase H site-specific cleavage, and qRT-PCR, which demonstrated precise absolute quantification of modification ratios and methylation copy numbers. With the help of Nm-VAQ, the 2'-O-methylation status of 5 sites in rRNA was evaluated.

biochemistry↗

bioRxiv2022

Guided construction of single cell reference for human and mouse lung

Accurate cell type identification is a key and rate-limiting step in single cell data analysis. Single cell references with comprehensive cell types, reproducible and functional validated cell identities, and common nomenclatures are much needed by the research community to optimize automated cell type annotation and facilitate data integration, sharing, and collaboration. In the present study, we developed a novel computational pipeline to utilize the LungMAP CellCards as a dictionary to consolidate single-cell transcriptomic datasets of 104 human lungs and 17 mouse lung samples and constructed "LungMAP CellRef" and "LungMAP CellRef Seed" for both normal human and mouse lungs. "CellRef Seed" has an equivalent prediction power and produces consistent cell annotation as does "CellRef" but improves computational efficiency and simplifies its utilization for fast automated cell type annotation and online visualization. This atlas set incorporates 48 human and 40 mouse well-defined lung cell types catalogued from diverse anatomic locations and developmental time points. Using independent datasets, we demonstrated the utility of our CellRefs for automated cell type annotation analysis of both normal and disease lungs. User-friendly web interfaces were developed to support easy access and maximal utilization of the LungMAP CellRefs. LungMAP CellRefs are freely available to the pulmonary research community through fast interactive web interfaces to facilitate hypothesis generation, research discovery, and identification of cell type alterations in disease conditions.

bioinformatics↗

bioRxiv2022

Self-organized canals enable long range directed material transport in bacterial communities

Long-range material transport is essential to maintain the physiological functions of multicellular organisms such as animals and plants. By contrast, material transport in bacteria is often short-ranged and limited by diffusion. Here we report a unique form of actively regulated long-range directed material transport in structured bacterial communities. Using Pseudomonas aeruginosa colonies as a model system, we discover that a large-scale and temporally evolving open channel system spontaneously develops in the colony via shear-induced banding. Fluid flows in the open channels support high-speed (up to 450 {micro}m/s) transport of cells and outer membrane vesicles over centimeters, and help to eradicate colonies of a competing species Staphylococcus aureus. The open channels are reminiscent of human-made canals for cargo transport, and the channel flows are driven by interfacial tension mediated by cell-secreted biosurfactants. The spatial-temporal dynamics of fluid flows in the open channels are qualitatively described by flow profile measurement and mathematical modeling. Our findings demonstrate that mechanochemical coupling between interfacial force and biosurfactant kinetics can coordinate large-scale material transport in primitive life forms, suggesting a new principle to engineer self-organized microbial communities.

biophysics↗