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Chen, N.

Publications and source records attributed to Chen, N..

At least 73 records · Page 4Linked to original sources

Genetically-encoded markers for confocal visualization of single dense core vesicles

Neuronal dense core vesicles (DCVs) store and release a diverse array of neuromodulators, trophic factors and bioamines. The analysis of single DCVs has largely been possible only using electron microscopy, which makes understanding cargo segregation and DCV heterogeneity difficult. To address these limitations, we developed genetically-encoded markers for DCVs that can be used in combination with standard immunohistochemistry and expansion microscopy, to enable single-vesicle resolution with confocal microscopy.

neuroscience↗

Lifetime fitness benefits of breeding site fidelity and low costs of inbreeding permit inbreeding tolerance in an avian cooperative breeder

Breeding with relatives often decreases individual fitness but can be mitigated via inbreeding avoidance mechanisms such as dispersal, mate-switching, and kin avoidance. Disentangling the effects of multiple mechanisms is challenging without detailed individual-level data and incomplete without considering all life stages. We leveraged 32 years of data from a population of Florida Scrub-Jays (Aphelocoma coerulescens) to investigate how dispersal and mate choice contribute to inbreeding risk across individuals lifetimes and quantify the associated fitness outcomes. We find that when mating is random, inbreeding is two-fold higher when dispersal is limited, but adding sex-biased dispersal significantly reduces inbreeding risk. Limited movement away from the breeding territory for later pairings (site fidelity) contributes more to higher-than-expected levels of inbreeding than limited natal dispersal. Within observed dispersal constraints, however, inbreeding is lower than expected under random mating, suggesting Florida Scrub-Jays actively avoid mating with relatives, especially first-order kin. Finally, males paired with close relatives and females who move farther for later pairings experience equivalently lower lifetime reproductive success. Breeders who move farther for first pairings have lower survival. Overall, our results show how variation in fitness tradeoffs and mate choice behavior across life stages affects inbreeding in a long-lived, social species.

animal behavior and cognition↗

A neural mechanism for compositional generalization of structure in humans

A human ability to adapt to the dynamics of novel environments relies on abstracting and generalizing from past experiences. Previous research has focused on how humans generalize from isolated sequential processes, yet we know little about mechanisms that enable adaptation to more complex dynamics, including those that govern much everyday experience. Here, using a novel sequence learning task based on graph factorization, coupled with simultaneous magnetoencephalography (MEG) recordings, we ask whether reuse of experiential "building blocks" enable inference and generalization. Behavioral results were consistent with participants decomposing task experience into subprocesses, abstracting their dynamical structure away from their sensory specifics and transferring these to a new task environment. Neurally, this transfer was associated with a learning-induced, condition-specific, increase in neural similarity among stimuli conditional on their structural role in abstract subprocesses that were shared across task phases. Consistent with this, a complementary dynamical role decoding analysis revealed enhanced neural decodability for stimuli sharing identical structural transition types across experiential contexts, specifically when prior experience included the relevant graph structure. Decoding strength for these role representations was positively related to behavioral success in subprocess knowledge transfer, a relationship that will require future independent replication. These findings provide neural evidence consistent with the idea that a structural scaffolding mechanism supports generalization of experience to new contexts.

neuroscience↗

Unveiling multi-scale architectural features in single-cell Hi-C data using scCAFE

Single-cell Hi-C (scHi-C) has provided unprecedented insights into the heterogeneity of 3D genome organization. However, its sparse and noisy nature poses challenges for computational analyses, such as chromatin architectural feature identification. Here, we introduce scCAFE, a deep learning model for the multi-scale detection of architectural features at the single-cell level. scCAFE provides a unified framework for annotating chromatin loops, TAD-like domains (TLDs), and compartments across individual cells. Our model outperforms previous scHi-C loop calling methods and delivers accurate predictions of TLDs and compartments that are biologically consistent with previous studies. The resulting single-cell annotations also offer a measure to characterize the heterogeneity of different levels of architectural features across cell types. We leverage this heterogeneity and identify a series of marker loop anchors, which demonstrate the potential of the 3D genome data to annotate cell identities without the aid of simultaneously sequenced omics data. Overall, scCAFE not only serves as a useful tool for analyzing single-cell genomic architecture, but also paves the way for precise cell-type annotations solely based on 3D genome features.

bioinformatics↗

Systematic Targeting of Protein Complexes with Molecular COUPLrs

Molecular glues that engage protein complexes have transformed the study of cell biology and have had a direct impact on clinical oncology. However, the identification of new glue classes and their corresponding protein complexes has remained largely serendipitous. To overcome this challenge, we report the development of molecular COUPLrs, elaborated small molecules flanked by two cysteine-reactive warheads, as well as CONNECT, an integrated chemical proteomic platform for target deconvolution. By profiling a library of molecular COUPLrs across 13 cancer cell lines, we uncovered hundreds of proteins that can be coupled together, including in some cases in mutant selective fashions. We develop an advanced COUPLr for the oncogene EML4-ALK, which engages the fusion outside of its kinase domain, restricts protein dynamics, and disrupts EML4-ALK signaling. Collectively, molecular COUPLrs substantially expand the scope of proteins that can be chemically connected, providing an unbiased approach to identify small molecules that target protein complexes.

systems biology↗

Temperature-Dependent Fold-Switching Mechanism of the Circadian Clock Protein KaiB

The oscillator of the cyanobacterial circadian clock relies on the ability of the KaiB protein to switch reversibly between a stable ground-state fold (gsKaiB) and an unstable fold-switched fold (fsKaiB). Rare fold-switching events by KaiB provide a critical delay in the negative feedback loop of this post-translational oscillator. In this study, we experimentally and computationally investigate the temperature dependence of fold switching and its mechanism. We demonstrate that the stability of gsKaiB increases with temperature compared to fsKaiB and that the Q10 value for the gsKaiB [->] fsKaiB transition is nearly three times smaller than that for the reverse transition. Simulations and native-state hydrogen-deuterium exchange NMR experiments suggest that fold switching can involve both subglobally and near-globally unfolded intermediates. The simulations predict that the transition state for fold switching coincides with isomerization of conserved prolines in the most rapidly exchanging region, and we confirm experimentally that proline isomerization is a rate-limiting step for fold switching. We explore the implications of our results for temperature compensation, a hallmark of circadian clocks, through a kinetic model.

biochemistry↗

Supervised and unsupervised deep learning-based approaches for studying DNA replication spatiotemporal dynamics

In eukaryotic cells, DNA replication is organised both spatially and temporally, as evidenced by the stage-specific spatial distribution of replication foci in the nucleus. Despite the genetic association of aberrant DNA replication with numerous human diseases, the labour-intensive methods employed to study DNA replication have hindered large-scale analyses of its roles in pathological processes. In this study, we first demonstrate that a convolutional neural network trained to classify S-phase stages based on DAPI and EdU patterns could identify altered replication dynamics in Rif1-deficient mouse embryonic stem cells (mESCs), revealing a skewed distribution across the various S-phase stages. Given the possible practical limitations associated with a supervised framework, we proceed to show that the abnormal replication profile of Rif1-deficient mESCs could further be detected by an unsupervised approach (based on self-supervised representation learning), which could additionally reconstruct progression through S-phase. Finally, we extend our approach to a well-characterised cellular model of inducible deregulated origin firing, involving cyclin E overexpression. Through parallel EdU- and PCNA-based analyses, we demonstrate the potential applicability of our method to patient samples, offering a means to identify the contribution of deregulated DNA replication to a plethora of pathogenic processes.

cell biology↗

A new high-quality genome assembly and annotation for the threatened Florida Scrub-Jay (Aphelocoma coerulescens)

The Florida Scrub-Jay (Aphelocoma coerulescens), a Federally Threatened, cooperatively-breeding bird, is an emerging model system in evolutionary biology and ecology. Extensive individual-based monitoring and genetic sampling for decades has yielded a wealth of data, allowing for the detailed study of social behavior, demography, and population genetics of this natural population. Here, we report a linkage map and a chromosome-level genome assembly and annotation for a female Florida Scrub-Jay made with long-read sequencing technology, chromatin conformation data, and the linkage map. We constructed a linkage map comprising 4,468 SNPs that had 34 linkage groups and a total sex-averaged autosomal genetic map length of 2446.78 cM. The new genome assembly is 1.32 Gb in length, consisting of 33 complete or near-complete autosomes and the sex chromosomes (ZW). This highly contiguous assembly has an N50 of 68 Mb and a Benchmarking Universal Single-Copy Orthologs (BUSCO) completeness score of 97.1% with respect to the Aves database. The annotated gene set has a BUSCO transcriptome completeness score of 95.5% and 18,051 identified protein-coding genes, 92.2% of which have associated functional annotations. This new, high-quality genome assembly and linkage map of the Florida Scrub-Jay provides valuable tools for future research into the evolutionary dynamics of small, natural populations of conservation concern. ARTICLE SUMMARYWe present a new high-quality genome assembly and annotation for the Florida Scrub-Jay (Aphelocoma coerulescens), a Federally Threatened bird species. In comparison to other genome assemblies of this species, our assembly is the first to be made using long-read sequencing technology and is the first generated from a female individual. We also constructed the first linkage map for this species using a population pedigree. Our genome assembly is highly contiguous and is of similar quality to other bird genome assemblies.

genomics↗

Pseudocobalamin production and use in Synechococcus cultures and communities

Cobalamin influences marine microbial communities because an exogenous source is required by most eukaryotic phytoplankton, and demand can exceed supply. Pseudocobalamin is a cobalamin analog that is produced and used by most cyanobacteria but is not directly available to eukaryotic phytoplankton. Some microbes can remodel pseudocobalamin into cobalamin, but a scarcity of pseudocobalamin measurements impedes our ability to evaluate its importance for marine cobalamin production. Here, we perform simultaneous measurements of pseudocobalamin and methionine synthase (MetH), the key protein that uses it as a co-factor, in Synechococcus cultures and communities. In Synechococcus sp. WH8102, pseudocobalamin quota decreases in low temperature (17 {degrees}C) and low N:P, while MetH did not. Pseudocobalamin and MetH quotas were influenced by culture methods and growth phase. Despite the variability present in cultures, we found a comparably consistent quota of 300 {+/-} 100 pseudocobalamin molecules per cyanobacterial cell in the Northwest Atlantic Ocean, suggesting that cyanobacterial cell counts may be sufficient to estimate pseudocobalamin inventories in this region. This work offers insights into cellular pseudocobalamin metabolism, and the environmental and physiological conditions that may influence it, and provides environmental measurements to further our understanding of when and how pseudocobalamin can influence marine microbial communities.

microbiology↗

Granzyme B-based CAR T cells block metastasis by eliminating circulating tumor cells

Chimeric antigen receptor (CAR) T cells have limited efficacy against solid tumors due to the hostile microenvironment. Circulating tumor cells (CTCs) are essential to metastasis, which is the cause of most of cancer-related death. Here, we generated GrB-CAR T cells targeting membrane-bound HSP70 (mHSP70), a highly tumor-specific antigen detected in numerous cancers. GrB-CAR T cells exhibited potent cytotoxicity against a broad spectrum of cancer cell lines and stem-like cancer cells in vitro and effectively inhibited xenograft tumor growth in vivo. Importantly, GrB-CAR T cells markedly decreased the number of CTCs and, therefore, hindered cancer metastasis in spontaneous metastasis models with uncontrollable primary tumor growth, a scenario commonly encountered in clinical trials of CAR T therapies for solid tumors. Furthermore, despite the 100% homology between human and macaque HSP70 protein, the autotransplantation of macaque T cells expressing human GrB-CAR did not cause any obvious toxic effects. These results not only demonstrate GrB-CAR T cells as a safe and effective tactic with broad-spectrum anticancer activity, but also offer strong experimental evidence and proof-of-concept validation for CAR T cell-mediated metastasis inhibition by targeting CTCs.

immunology↗

Glioma-associated fibroblasts promote glioblastoma resistance to temozolomide through CCL2-CCR2 paracrine signaling

Complicated tumor microenvironment contributes mostly to chemoresistance in glioblastoma. Glioma-associated fibroblasts (GAFs) were recently identified as non-tumor stromal cells in the glioblastoma microenvironment, whereas their function in glioblastoma chemoresistance is unclear. Herein, we interrogated the correlation between GAFs and chemoresistance of glioblastoma by examining a series of patient-derived GAFs and glioblastoma organoids (GBOs), revealing that GAFs could promote temozolomide resistance in glioblastoma. Mechanistically, C-C motif chemokine ligand 2 (CCL2) secreted by GAFs selectively activated the ERK1/2 signaling in glioblastoma cells to potentiate temozolomide resistance. Pharmacologically disrupting the CCL2-CCR2 axis or MEK1/2-ERK1/2 pathway effectively improved the therapeutic efficacy of temozolomide in GBM cells and patient-derived GBOs, and both decreased phospho-ERK1/2 expression. Collectively, our results identified that targeting the GAF-dominated CCL2-ERK1/2 pathway may be an alternative strategy to alleviate the GAF-mediated chemoresistance of glioblastoma. SignificanceComprehensive interpretation of the mutual support between tumor microenvironment and cancer cells is demanded for glioma with poor response rates to chemotherapy. This study demonstrates that GAFs promote the temozolomide resistance of glioblastoma by secreting cytokine CCL2 to activate ERK1/2 pathway, which may serve as a potential druggable candidate. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/581575v1_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@167b955org.highwire.dtl.DTLVardef@b892bcorg.highwire.dtl.DTLVardef@1a9b082org.highwire.dtl.DTLVardef@18327a8_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphic abstract.C_FLOATNO Schematic illustration for GAFs mediated chemoresistance of GBMs and underlying mechanisms. We demonstrate that Glioma-associated Fibroblasts (GAFs) grow in gliomas by isolating and identifying a panel of patient-derived GAFs. CCL2 secreted by GAFs stimulates CCR2 in GBM cells, which promotes activation of the ERK1/2 expression to potentiate GBM chemoresistance. C_FIG

cancer biology↗

Immunomodulatory Porous Regenerative Scaffolds for in situ Vascular Engineering

The 70-year quest for synthetic vascular graft (sVG) endothelialization has not led to completely healed endothelium in clinically used sVGs. In humans, healing is limited to the vicinity of anastomotic regions (pannus ingrowth) and does not reach the middle regions of sVGs. Here, we conducted proof-of-concept implantation of immunomodulatory porous regenerative scaffolds for in situ vascular engineering (IMPRESSIVE) as interposition grafts in sheep carotid arteries. These scaffolds are based on a new polyurethane (PU) material featuring a 40 {micro}m precision porous structure optimized for angiogenesis. The modulus of the PU was adjusted to match that of natural arteries. The implantation study revealed rapid healing in IMPRESSIVE sVGs. In side-by-side comparison with standard polytetrafluoroethylene (PTFE) grafts, the luminal surfaces of PU grafts were almost completely covered with nucleated cells, while healing in PTFE grafts was limited to several millimeters within anastomotic regions. Endothelialization was observed in the middle regions of PU grafts and overall endothelialization increased significantly compared to PTFE grafts. Densities of mononuclear cells, foreign body giant cells (FBGCs), and endothelial cells within graft walls of PU grafts were also significantly higher than those in PTFE grafts, suggesting transmural cellular infiltration may play a key role in overall improved healing. High percentages of macrophages in pores of PU grafts show Type 1 (CCR7+) and Type 2 (mannose receptor, MR+) characteristics. We also discovered that FBGCs exist in a diverse spectrum of phenotypes. Dually activated FBGCs (CCR7+MR+, G1/2) dominate the population of FBGCs associated with pro-healing PU grafts. These observations suggest a complex, balanced pro-healing response from macrophages and FBGCs. The IMPRESSIVE approach may enable complete endothelialization in pro-healing sVGs and have wide applications in implantable devices and tissue engineering.

bioengineering↗

A high coverage Mesolithic aurochs genome and effective leveraging of ancient cattle genomes using whole genome imputation.

Ancient genomic analyses are often restricted to utilising pseudo-haploid data due to low genome coverage. Leveraging low coverage data by imputation to calculate phased diploid genotypes that enable haplotype-based interrogation and SNP calling at unsequenced positions is highly desirable. This has not been investigated for ancient cattle genomes despite these being compelling subjects for archaeological, evolutionary and economic reasons. Here we test this approach by sequencing a Mesolithic European aurochs (18.49x; 9852-9376 calBC), an Early Medieval European cow (18.69x; 427-580 calAD), and combine these with published individuals; two ancient and three modern. We downsample these genomes (0.25x, 0.5x, 1.0x, 2.0x) and impute diploid genotypes, utilising a reference panel of 171 published modern cattle genomes that we curated for 21.7 million (Mn) phased single-nucleotide polymorphisms (SNPs). We recover high densities of correct calls with an accuracy of >99.1% at variant sites for the lowest downsample depth of 0.25x, increasing to >99.5% for 2.0x (transversions only, minor allele frequency (MAF) [≥]2.5%). The recovery of SNPs correlates with coverage, on average 58% of sites are recovered for 0.25x increasing to 87% for 2.0x, utilising an average of 3.5 million (Mn) transversions (MAF [≥]2.5%), even in the aurochs which is temporally and morphologically distinct from the reference panel. Our imputed genomes behave similarly to directly called data in allele-frequency-based analyses; for example consistently identifying runs of homozygosity >2mb, including a long homozygous region in the Mesolithic European aurochs.

genomics↗

Impacts of increasing isolation and environmental variation on Florida Scrub-Jay demography

Isolation caused by anthropogenic habitat fragmentation can destabilize populations. Populations relying on the inflow of immigrants can face reduced fitness due to inbreeding depression as fewer new individuals arrive. Empirical studies of the demographic consequences of isolation are critical to understand how populations persist through changing conditions. We used a 34-year demographic and environmental dataset from a population of cooperatively-breeding Florida Scrub-Jays (Aphelocoma coerulescens) to create mechanistic models linking environmental and demographic factors to population growth rates. We found that the population has not declined despite both declining immigration and increasing inbreeding, owing to a coinciding response in breeder survival. We find evidence of density-dependent immigration, breeder survival, and fecundity, indicating that interactions between vital rates and local density play a role in buffering the population against change. Our study elucidates the impacts of isolation on demography and how long-term stability is maintained via demographic responses.

ecology↗

Ecological niche contributes to the persistence of the Western x Glaucous-winged Gull hybrid zone

Hybrid zones occur in nature when populations with limited reproductive barriers overlap in space. Many hybrid zones persist over time, and different models have been proposed to explain how selection can maintain hybrid zone stability. More empirical studies are needed to elucidate the role of ecological adaptation in maintaining stable hybrid zones. Here, we investigated the role of exogenous factors in maintaining a hybrid zone between western gulls (Larus occidentalis) and glaucous-winged gulls (L. glaucescens). We used ecological niche models (ENMs) and niche similarity tests to quantify and examine the ecological niches of western gulls, glaucous-winged gulls, and their hybrids. We found evidence of niche divergence between all three groups. Our results best support the bounded superiority model, providing further evidence that exogenous selection favoring hybrids may be an important factor in maintaining this stable hybrid zone.

ecology↗

A novel "ceasefire" model to explain efficient seed transmission of Xanthomonas citri pv. fuscans to common bean.

O_LIAlthough seed represents an important means of plant pathogen dispersion, the seed-pathogen dialogue remains largely unexplored. C_LIO_LIA multi-omic approach (i.e. dual RNAseq, plant small RNAs and methylome) was performed at different seed developmental stages of common bean (Phaseolus vulgaris L.) during asymptomatic colonization by Xanthomonas citri pv. fuscans (Xcf). C_LIO_LIIn this condition, Xcf did not produce disease symptoms, neither affect seed development. Although, an intense molecular dialogue, via important transcriptional changes, was observed at the early seed developmental stages with down-regulation of plant defense signal transduction, via action of plant miR, and upregulation of the bacterial Type 3 Secretion System. At later seed maturation stages, molecular dialogue between host and pathogen was reduced to few transcriptome changes, but marked by changes in DNA methylation of plant defense and germination genes, in response to Xcf colonization, potentially acting as defense priming to prepare the host for the post-germination battle. This distinct response of infected seeds during maturation, with a more active role at early stages refutes the widely diffused assumption considering seeds as passive carriers of microbes. C_LIO_LIFinally, our data support a novel plant-pathogen interaction model, specific to the seed tissues, which differs from others by the existence of distinct phases during seed-pathogen interaction with seeds first actively interacting with colonizing pathogens, then both belligerents switch to more passive mode at later stages. We contextualized this observed scenario in a novel hypothetical model that we called "ceasefire", where both the pathogen and the host benefit from temporarily laying down their weapons until the moment of germination. C_LI

plant biology↗

Connecting single neuron transcriptomes to the projectome in mouse visual cortex

The mammalian brain is composed of diverse neuron types that play different functional roles. Recent single-cell RNA sequencing approaches have led to a whole brain taxonomy of transcriptomically-defined cell types, yet cell type definitions that include multiple cellular properties can offer additional insights into a neurons role in brain circuits. While the Patch-seq method can investigate how transcriptomic properties relate to the local morphological and electrophysiological properties of cell types, linking transcriptomic identities to long-range projections is a major unresolved challenge. To address this, we collected coordinated Patch-seq and whole brain morphology data sets of excitatory neurons in mouse visual cortex. From the Patch-seq data, we defined 16 integrated morphoelectric-transcriptomic (MET)-types; in parallel, we reconstructed the complete morphologies of 300 neurons. We unified the two data sets with a multi-step classifier, to integrate cell type assignments and interrogate cross-modality relationships. We find that transcriptomic variations within and across MET-types correspond with morphological and electrophysiological phenotypes. In addition, this variation, along with the anatomical location of the cell, can be used to predict the projection targets of individual neurons. We also shed new light on infragranular cell types and circuits, including cell-type-specific, interhemispheric projections. With this approach, we establish a comprehensive, integrated taxonomy of excitatory neuron types in mouse visual cortex and create a system for integrated, high-dimensional cell type classification that can be extended to the whole brain and potentially across species.

neuroscience↗

Translocations spur population growth but exacerbate inbreeding in an imperiled species

Land and natural resource usage that supports human society can pose a risk to the survival of other species, spurring biodiversity loss. In extreme cases, when development threatens the existence of individuals, wildlife managers may perform mitigation translocations, relocating individuals out of harms way. We investigated the efficacy of mitigation translocations as a conservation strategy in Federally Threatened Florida Scrub-Jays using a dataset that provided unprecedented resolution into both the demographic and genomic outcomes of translocations. Over the course of seven years, a total of fourteen groups (51 jays) from five subpopulations that had been declining from agriculture and lack-of-fire driven habitat degradation were translocated to a larger site of more contiguous restored habitat with only four family groups, to mitigate for loss of these subpopulations from mining activity. Habitat restoration and translocations established a core population that increased 10-fold in size after only 17 years from the first translocations. Pedigree analyses of this population revealed that a small subset of mostly translocated individuals fueled the demographic expansion, with a single breeding pair responsible for [~]24% of the ancestral genetic contributions since 2008. Genomic comparisons between translocated individuals and individuals from the core population before and after translocations revealed that the high reproductive skew led to increased inbreeding and loss of genetic diversity. This study stresses the importance of demographic and genetic monitoring following translocations, and that subsequent, genetic-rescue-oriented translocations may be necessary in mitigation scenarios to counter the genetic consequences of reproductive skew in fragmented populations. Significance StatementThere is ongoing debate surrounding the effectiveness of mitigation-driven translocations for conservation, however we show that translocations to mitigate the effects of mining on Federally Threatened Florida Scrub-Jays spurred population growth; a major boon to the viability of this species. We translocated individuals from at-risk subpopulations that were demographic sinks into recently restored habitat, which quickly established a rapidly growing core population. We demonstrate that demographic and genetic recovery do not necessarily go hand-in-hand, as reproduction was highly skewed towards a small subset of mostly translocated individuals, which increased inbreeding and eroded genetic diversity. This stresses the importance of demographic and genetic monitoring for identifying reproductive skew, allowing for adaptive management that addresses inbreeding and achieves broader conservation goals.

evolutionary biology↗