Biology subjects

Cheng, Z.

Publications and source records attributed to Cheng, Z..

At least 127 records · Page 7Linked to original sources

bioRxiv2020

Probing neuronal functions with precise and targeted laser ablation in the living cortex

Targeted cell ablation is an important strategy for dissecting the function of individual cells within biological tissues. Here we developed an amplified femtosecond laser-coupled two-photon microscopy (AFL-TPM) system that allows instantaneous and targeted ablation of individual cells and real-time monitoring of neuronal network changes in the living mouse cortex. Through precise and iterative control of the laser power and position, individual cells could be ablated by a single femtosecond light pulse with minimum collateral damage. We further show that ablation of individual somatostatin-expressing interneuron increases the activity of nearby neurons in the primary motor cortex during motor learning. Through precise dendrotomy, we reveal that different dendritic branches of layer 5 pyramidal neurons are structurally and functionally independent. By ablating individual cells and their processes in a spatiotemporally specific manner, the AFL-TPM system could serve as an important means for understanding the functions of cells within the complicated neuronal network.

neuroscience↗

bioRxiv2020

De Novo Mutation in an Enhancer of EBF3 in simplex autism

Previous research in autism and other neurodevelopmental disorders (NDDs) has indicated an important contribution of de novo protein-coding variants within specific genes. The role of de novo noncoding variation has been observable as a general increase in genetic burden but has yet to be resolved to individual functional elements. In this study, we assessed whole-genome sequencing data in 2,671 families with autism, with a specific focus on de novo variation in enhancers with previously characterized in vivo activity. We identified three independent de novo mutations limited to individuals with autism in the enhancer hs737. These mutations result in similar phenotypic characteristics, affect enhancer activity in vitro, and preferentially occur in AAT motifs in the enhancer with predicted disruptions of transcription factor binding. We also find that hs737 is enriched for copy number variation in individuals with NDDs, is dosage sensitive in the human population, is brain-specific, and targets the NDD gene EBF3 that is genome-wide significant for protein coding de novo variants, demonstrating the importance of understanding all forms of variation in the genome. One Sentence SummaryWhole-genome sequencing in thousands of families reveals variants relevant to simplex autism in a brain enhancer of the well-established neurodevelopmental disorder gene EBF3.

genomics↗

bioRxiv2020

A spatially-tracked single cell transcriptomics map of neuronal networks in the intrinsic cardiac nervous system

We developed a spatially-tracked single neuron transcriptomics map of an intrinsic cardiac ganglion - the right atrial ganglionic plexus (RAGP) that is a critical mediator of vagal control of the sinoatrial node (SAN) activity. We developed a 3D representation of RAGP with extensive mapping of neurons and used neuronal tracing to identify the spatial distribution of the subset of neurons that project to the SAN. RNAseq of laser capture microdissected neurons revealed a distinct composition of RAGP neurons compared to CNS neuronal subtypes. High-throughput qPCR of hundreds of laser capture microdissected single neurons led to a surprising finding that cholinergic and catecholaminergic neuronal markers Th and Chat were correlated, suggesting multipotential phenotypes that can drive neuroplasticity within RAGP. Interestingly, no single gene or module was an exclusive marker of RAGP neuronal connectivity to SAN. Neuropeptide-receptor coexpression analysis revealed a combinatorial paracrine neuromodulatory network within RAGP, informing follow-on studies on the vagal control of RAGP to regulate cardiac function in health and disease.

neuroscience↗

bioRxiv2020

3D single cell scale anatomical map of sex-dependent variability of the rat intrinsic cardiac nervous system

We developed and analyzed a single cell scale anatomical map of the rat intrinsic cardiac nervous system (ICNS) across 4 male and 3 female hearts. We find the ICNS has a reliable structural organizational plan across individuals that may provide the foundation for further analyses of the ICNS in cardiac function and disease. The distribution of the ICNS was evaluated by 3D visualization and using data-driven clustering. The pattern, distribution and clustering of ICNS neurons across all male and female rat hearts is highly conserved, demonstrating a coherent organizational plan in which distinct clusters of neurons are consistently localized. Female hearts had fewer neurons, lower packing density and slightly reduced distribution, but with the same localization. We registered the anatomical data from each heart to a geometric scaffold, normalizing their 3D coordinates for the standardization of common anatomical planes and providing the path where multiple experimental results and data types can be integrated and compared.

neuroscience↗

bioRxiv2020

AD-linked R47H-TREM2 mutation induces disease-enhancing proinflammatory microglial states in mice and humans

The hemizygous R47H variant of TREM2, a microglia-specific gene in the brain, increases risk for late-onset Alzheimers disease (AD). In this study, we identified a subpopulation of microglia with disease-enhancing proinflammatory signatures associated with the R47H mutation in human AD brains and tauopathy mouse brains. Using transcriptomic analysis at the single-nuclei level from AD patients with the R47H or the common variant (CV)-TREM2 with matched sex, pathology and APOE status, we found that the R47H mutation was associated with cell type- and sex-specific transcriptional changes in human AD brains, with microglia exhibiting the most robust alterations. Further characterization revealed that R47H-associated microglial subpopulations had enhanced inflammatory signatures including hyperactivation of Akt, one of the signaling pathways downstream of TREM2. In a newly-generated tauopathy knock-in mouse model expressing one allele of human TREM2 (hTREM2) with either the R47H mutation or CV, R47H induced and exacerbated tau-mediated spatial memory deficits in female mice. Single-cell transcriptomic analysis of microglia from these mice also revealed transcriptomic changes induced by R47H that had significant overlaps with R47H microglia in human AD brains, including robust increases in proinflammatory cytokines, activation of Syk-Akt-signaling, and elevation of a subset of disease-associated microglial signatures. Strikingly, pharmacological Akt inhibition largely reversed the enhanced inflammatory signatures induced by R47H in primary microglia treated with tau fibrils. By unraveling the disease-enhancing properties of the R47H mutation in mouse and human, our findings shed light on an immune-linked AD subtype and provide new directions for modulating microglial immune responses to treat AD.

neuroscience↗

bioRxiv2020

Listeria monocytogenes personalized cancer vaccines drive therapeutic immune responses to cancer derived neoantigens

BackgroundRecent advances in the field of cancer immunotherapy have identified CD8+ T cell responses against tumor-specific mutations as a key driver of tumor regression and overall survival. ADXS-NEO is a personalized Listeria monocytogenes (Lm)-based immunotherapy designed to target a patients mutation-derived tumor-specific neoantigens. The objective of this study is to demonstrate the feasibility of using the ADXS-NEO platform to target tumor-specific point mutations and control tumor growth by generating neoantigen-specific T cell responses using a pre-clinical mouse tumor model. MethodsWhole-exome sequencing of the MC38 mouse tumor cell line identified 2870 unique non-synonymous mutations. The netMHCcons algorithm was used to predict 137 potential neoantigens. We validated 20 immunogenic neoantigens either by peptide immunization followed by ELISPOT or by the presence of CD8+ T cells recognizing the neoantigen peptide following checkpoint inhibitor treatment. Two ADXS-NEO vectors were constructed; Lm20, targeting 20 validated immunogenic neoantigens, and Lm19, targeting most of the non-validated NSMs. ResultsBoth Lm19 & Lm20 significantly slowed tumor growth in C57BL/6 mice compared to control. An accumulation of ADXS-NEO-specific TILs was observed in tumor bearing mice treated with either Lm19 or Lm20. Examination of the tumor microenvironment in Lm19 or Lm20 treated mice revealed a decrease in the frequency and absolute number of Tregs, TAMs, MDSCs, and PD1high exhausted CD8+ T cells as well as an increase in the frequency and absolute number of effector CD8+ T cells, relative to control. ConclusionADXS-NEO is a potent immunotherapy capable of driving immune responses against tumor-specific mutations and leading to tumor control in mice.

immunology↗

bioRxiv2020

Crystal Structure of INTS3/INTS6 complex reveals Functional Importance of INTS3 dimerization in DSB Repair

SOSS1 is a single-stranded DNA (ssDNA)-binding protein complex that plays a critical role in double-strand DNA break (DSB) repair. SOSS1 consists of three subunits: INTS3, SOSSC, and hSSB1 with INTS3 serving as a scaffold to stabilize this complex. Moreover, the integrator complex subunit 6 (INTS6) participates in the DNA damage response through direct binding to INTS3 but how INTS3 interacts with INTS6 thereby, impacting DBS repair is not clear. Here, we determined the crystal structure of the C-terminus of INTS3 (INTS3c) in complex with the C-terminus of INTS6 (INTS6c) at a resolution of 2.4 [A]. Structure analysis revealed that two INTS3c subunits dimerize and interact with INTS6c via conserved residues. Subsequent biochemical analyses confirmed that INTS3c forms a stable dimer and INTS3 dimerization is important for recognizing the longer ssDNA. Perturbation of INTS3c dimerization and disruption of the INTS3c/INTS6c interaction, impair the DSB repair process. Altogether, these results unravel the underappreciated role of INTS3 dimerization and the molecular basis of INTS3/INTS6 interaction in DSB repair.

biochemistry↗

bioRxiv2020

Facing small and biased data dilemma in drug discovery with federated learning

Artificial intelligence (AI) models usually require large amounts of high-quality training data, which is in striking contrast to the situation of small and biased data faced by current drug discovery pipelines. The concept of federated learning has been proposed to utilize distributed data from different sources without leaking sensitive information of these data. This emerging decentralized machine learning paradigm is expected to dramatically improve the success of AI-powered drug discovery. We here simulate the federated learning process with 7 aqueous solubility datasets from different sources, among which there are overlapping molecules with high or low biases in the recorded values. Beyond the benefit of gaining more data, we also demonstrate federated training has a regularization effect making it superior than centralized training on the pooled datasets with high biases. Further, two more cases are studied to test the usability of federated learning in drug discovery. Our work demonstrates the application of federated learning in predicting drug related properties, but also highlights its promising role in addressing the small data and biased data dilemma in drug discovery.

pharmacology and toxicology↗

bioRxiv2020

Differential responses of transplanted stem cells to the diseased environment unveiled by a single molecular NIR II cell tracker

Stem cell therapy holds high promises in regenerative medicine. The major challenge of clinical translation is to precisely and quantitatively evaluate the in vivo cell distribution, migration, and engraftment, which cannot be easily achieved by current techniques. To address this issue, for the first time, we have developed a single molecular cell tracker with a strong fluorescence signal in the second near-infrared (NIR-II) window (1000-1700 nm) for real-time monitoring of in vivo cell behaviors in both healthy and diseased animal models. The NIR-II tracker (CelTrac1000) has shown complete cell labeling with low cytotoxicity and profound long-term tracking ability for 30 days in high temporospatial resolution for semi-quantification of the biodistribution of primary mesenchymal stem cell and induced pluripotent stem cell-derived endothelial cells. Taking advantage of the unique merits of CelTrac1000, the responses of transplanted stem cells to different diseased environments have been discriminated and unveiled. Furthermore, we also demonstrate CelTrac1000 as a universal and effective technique for ultrafast real-time tracking of the cellular migration and distribution in a single cell cluster resolution, along with the lung contraction and heart beating. As such, this single molecular NIR-II tracker will shift the optical cell tracking into a single cell cluster and millisecond temporospatial resolution for better evaluating and understanding stem cell therapy, affording optimal doses and efficacy. Significance StatementFor the first time, we synthesized a NIR-II tracker (CelTrac1000) for ultrafast real-time tracking of the migration trajectory of transplanted mesenchymal stem cells in the circulatory system with a single cell cluster resolution. Taking advantage of the merits of CelTrac1000, the responses of transplanted stem cells to different diseased environments, including acute lung injury, myocardial infarction, and middle cerebral artery occlusion, have been discriminated and unveiled in mice models. As such, our approach can help correlate critical biomedical information in stem cell therapies, such as stem cell dosing and engraftment and their relationships with efficacy, providing more accurate therapeutic treatment and outcomes in certain diseases during a long evaluation period (>30 days) in comparison with the commercial Qtracker (7-10 days).

bioengineering↗

bioRxiv2019

Genomic Characterization of Posttraumatic Stress Disorder in a Large US Military Veteran Sample

Individuals vary in their liability to develop Posttraumatic Stress Disorder (PTSD), the symptoms of which are highly heterogeneous, following exposure to life-threatening trauma. Understanding genetic factors that contribute to the biology of PTSD is critical for refining diagnosis and developing new treatments. Using genetic data from more than 250,000 participants in the Million Veteran Program, genomewide association analyses were conducted using a validated electronic health record-based algorithmically-defined PTSD diagnosis phenotype (48,221 cases and 217,223 controls), and PTSD quantitative symptom phenotypes (212,007 individuals). We identified several genome-wide significant loci in the case-control analyses, and numerous such loci in the quantitative trait analyses, including some (e.g., MAD1L1; TCF4; CRHR1) that were associated with multiple symptom sub-domains and total symptom score, and others that were more specific to certain symptom sub-domains (e.g., CAMKV to re-experiencing; SOX6 to hyperarousal). Genetic correlations between all pairs of symptom sub-domains and their total were very high (rg 0.93 - 0.98) supporting validity of the PTSD diagnostic construct. We also demonstrate strong shared heritability with a range of traits, show that heritability persists when conditioned on other major psychiatric disorders, present independent replication results, provide support for one of the implicated genes in postmortem brain of individuals with PTSD, and use this information to identify potential drug repositioning candidates. These results point to the utility of genetics to inform and validate the biological coherence of the PTSD syndrome despite considerable heterogeneity at the symptom level, and to provide new directions for treatment development.

genetics↗

bioRxiv2019

Meta-analysis of problematic alcohol use in 435,563 individuals identifies 29 risk variants and yields insights into biology, pleiotropy and causality

Problematic alcohol use (PAU) is a leading cause of death and disability worldwide. Although genome-wide association studies (GWASs) have identified PAU risk genes, the genetic architecture of this trait is not fully understood. We conducted a proxy-phenotype meta-analysis of PAU combining alcohol use disorder and problematic drinking in 435,563 European-ancestry individuals. We identified 29 independent risk variants, 19 of them novel. PAU was genetically correlated with 138 phenotypes, including substance use and psychiatric traits. Phenome-wide polygenic risk score analysis in an independent biobank sample (BioVU, n=67,589) confirmed the genetic correlations between PAU and substance use and psychiatric disorders. Genetic heritability of PAU was enriched in brain and in genomic conserved and regulatory regions. Mendelian randomization suggested causal effects on liability to PAU of substance use, psychiatric status, risk-taking behavior, and cognitive performance. In summary, this large PAU meta-analysis identified novel risk loci and revealed genetic relationships with numerous other outcomes.

genetics↗

bioRxiv2019

Enhance genome editing efficiency and specificity by a quick CRISPR/Cas9 system

CRISPR/Cas9 is a powerful genome editing tool that has been successfully applied to a variety of species, including zebrafish. However, targeting efficiencies vary greatly at different genomic loci, the underlying causes of which were still elusive. Here we report a quick CRISPR/Cas9 system, designated as qCas9, which exhibits accelerated turnover of Cas9 protein in zebrafish. Our data showed that qCas9 significantly improved targeting efficiency, including both knock-out and knock-in in F0 embryos, and yielded higher germline transmission rate in founder screen. Importantly, qCas9 showed little to no off-target editing in zebrafish and profoundly reduced off-target effect in HEK293T cell line. In summary, our findings demonstrate that qCas9 is a simple, economic and highly effective method to improve genome editing efficiency in zebrafish embryos and also holds great potential in reducing off-target effect in mammalian cell lines.

genetics↗

bioRxiv2019

A catalog of cis-regulatory mutations in 12 major cancer types

Despite the recent availability of complete genome sequences of tumors from thousands of patients, isolating disease-causing (driver) non-coding mutations from the plethora of somatic variants is notoriously challenging, and only a handful of validated examples exist. By integrating whole-genome sequencing, gene expression, chromatin accessibility, and genetic data from TCGA, we identified 301 non-coding somatic mutations that affect gene expression in cis. These mutations cluster into 36 hotspot regions with diverse molecular mechanisms of gene expression regulation. We further show that these mutations have hallmark features of noncoding drivers; namely, that they confer a positive selection on growth, functionally disrupt transcription factor binding sites, and contribute to disease progression reflected in decreased overall patient survival.

genomics↗

bioRxiv2019

Precise annotation of human, chimpanzee, rhesus macaque and mouse mitochondrial genomes using 5’ and 3’ end small RNAs

Using 5 and 3 end small RNAs, we annotated human, chimpanzee, rhesus macaque and mouse mitochondrial genomes at 1 base-pair (bp) resolution to cover both strands of the mammalian mitochondrial genome entirely without leaving any gaps or overlaps. The precise annotation of all coding and non-coding genes (e.g. ncMT1, MDL2 and MDL1AS) led to the discovery of novel functions and mechanisms of mitochondrion. In this study, we defined the conserved sequence block (CSB) region to span five CSBs (CSB1, CSB2, CSB3, LSP and HSP) and identified the motifs of five CSBs in the mitochondrial displacement loop (D-loop) regions of 52 mammals. The conserved arrangement of these five CSBs in 17 primates inspired us to investigate the function of the mtDNA D-loop, which has been puzzling scientists for more than 50 years. We found that 5 sRNAs of MDL1AS control the expression levels of mitochondrial genes as a whole by a negative feedback mechanism. Thus, the precise annotations of three CSBs (CSB2, LSP and HSP) in more species will help to understand the function of the mtDNA D-loop. The precision annotation of animal mitochondrial genomes also provides abundant information for studying the molecular phylogenetics and evolution of animals.

bioinformatics↗

bioRxiv2019

Single cell chromatin accessibility reveals pancreatic islet cell type- and state-specific regulatory programs of diabetes risk

Genetic risk variants for complex, multifactorial diseases are enriched in cis-regulatory elements. Single cell epigenomic technologies create new opportunities to dissect cell type-specific mechanisms of risk variants, yet this approach has not been widely applied to disease-relevant tissues. Given the central role of pancreatic islets in type 2 diabetes (T2D) pathophysiology, we generated accessible chromatin profiles from 14.2k islet cells and identified 13 cell clusters including multiple alpha, beta and delta cell clusters which represented hormone-producing and signal-responsive cell states. We cataloged 244,236 islet cell type accessible chromatin sites and identified transcription factors (TFs) underlying both lineage- and state-specific regulation. We measured the enrichment of T2D and glycemic trait GWAS for the accessible chromatin profiles of single cells, which revealed heterogeneity in the effects of beta cell states and TFs on fasting glucose and T2D risk. We further used machine learning to predict the cell type-specific regulatory function of genetic variants, and single cell co-accessibility to link distal sites to putative cell type-specific target genes. We localized 239 fine-mapped T2D risk signals to islet accessible chromatin, and further prioritized variants at these signals with predicted regulatory function and co-accessibility with target genes. At the KCNQ1 locus, the causal T2D variant rs231361 had predicted effects on an enhancer with beta cell-specific, long-range co-accessibility to the insulin promoter, and deletion of this enhancer reduced insulin gene and protein expression in human embryonic stem cell-derived beta cells. Our findings provide a cell type- and state-resolved map of gene regulation in human islets, illuminate likely mechanisms of T2D risk at hundreds of loci, and demonstrate the power of single cell epigenomics for interpreting complex disease genetics.

genomics↗

bioRxiv2019

Comprehensive Integrated Anatomical And Molecular Atlas Of Rodent Intrinsic Cardiac Nervous System

In this study, we developed, coordinated, and integrated several technologies including novel whole organ imaging, software development to support the very first precise 3D neuroanatomical mapping and molecular phenotyping of the intracardiac nervous system (ICN). While qualitative and gross anatomical descriptions of the anatomy of the ICN have been presented, we here bring forth the first comprehensive atlas at large scale of the entire ICN in rat at a single cell resolution. Our work for the first time provides a novel 3D model to precisely integrate anatomical, functional and molecular data in the 3D digitally reconstructed whole heart with high resolution at the micron scale. This work represents the cutting edge in a long history of attempts to understand the anatomical substrate upon which the neuronal control of cardiac function is built. To our knowledge, there has not yet been a comprehensive histological mapping to generate a neurocardiac atlas at cellular and molecular level for the whole heart of any species. We now display the full extent and the position of neuronal clusters on the base and posterior left atrium, and the distribution of molecular phenotypes in that context. In addition we display in this context distinct molecular phenotypes that are defined along the base-to-apex axis, and the present novel discovery of their phenotypical spatial gradients, have not been previously described. The development of these approaches needed to acquire these data has produced method pipelines which can not only achieve the goals of anatomical and molecular mapping of the heart, but also provide the method pipelines for mapping other organs (e.g., stomach, lung, kidney, and liver).

neuroscience↗

bioRxiv2019

The Pseudomonas aeruginosa accessory genome elements, including bacterial immune systems effect virulence towards C. elegans

Multicellular animals and bacteria frequently engage in predator-prey and host-pathogen interactions, such as the well-studied relationship between Pseudomonas aeruginosa and the nematode Caenorhabditis elegans. This study investigates the genomic and genetic basis of bacterial-driven variability in P. aeruginosa virulence towards C. elegans. Natural isolates of P. aeruginosa that exhibit diverse genomes display a broad range of virulence towards C. elegans. Using gene association and genetic analysis, we identified accessory genome elements that correlate with virulence, including both known and novel virulence determinants. Among the novel genes, we found a viral-like mobile element, the teg block, that impairs virulence and whose acquisition is restricted by CRISPR-Cas systems. Further genetic and genomic evidence suggests that spacer-targeted elements preferentially associate with lower virulence and suggest a positive, albeit indirect, role for host CRISPR-Cas systems in the restriction of accessory genome elements that may be detrimental to virulence.

evolutionary biology↗

bioRxiv2019

Leaf shape is a predictor of fruit quality and cultivar performance in tomato

O_LICommercial tomato (Solanum lycopersicum) is one of the most widely grown vegetable crops worldwide. Heirloom tomatoes retain extensive genetic diversity and a considerable range of fruit quality and leaf morphological traits.\nC_LIO_LIHere the role of leaf morphology was investigated for its impact on fruit quality. Heirloom cultivars were grown in field conditions and BRIX by Yield (BY) and other traits measured over a fourteen-week period. The complex relationships among these morphological and physiological traits were evaluated using PLS-Path Modeling, and a consensus model developed.\nC_LIO_LIPhotosynthesis contributed strongly to vegetative biomass and sugar content of fruits but had a negative impact on yield. Conversely leaf shape, specifically rounder leaves, had a strong positive impact on both fruit sugar content and yield. Cultivars such as Stupice and Glacier, with very round leaves, had the highest performance in both fruit sugar and yield. Our model accurately predicted BY for two commercial cultivars using leaf shape data as input.\nC_LIO_LIThis study revealed the importance of leaf shape to fruit quality in tomato, with rounder leaves having significantly improved fruit quality. This correlation was maintained across a range of diverse genetic backgrounds and shows the importance of leaf morphology in tomato crop improvement.\nC_LI

plant biology↗