Biology subjects

Chen, Q.

Publications and source records attributed to Chen, Q..

At least 451 records · Page 25Linked to original sources

bioRxiv2019

Universal and naked-eye gene detection platform based on CRISPR/Cas12a/13a system

Colorimetric gene detection based on gold nanoparticles (AuNPs) is an attractive detection format due to its simplicity. Here, we report a new design for a colorimetric gene-sensing platform based on the CRISPR/Cas system that has improved specificity, sensitivity, and universality. CRISPR/Cas12a and CRISPR/Cas13a have two distinct catalytic activities and are used for specific target gene recognition. Programmable recognition of DNA by Cas12a/crRNA and RNA by Cas13a/crRNA with a complementary sequence activates the nonspecific trans-ssDNA or -RNA cleavage, respectively, thus degrading the ssDNA or RNA linkers which are designed as a hybridization template for the AuNP-DNA probe pair. Target-induced trans -ssDNA or RNA cleavage leads to a distance-dependent color change for the AuNP-DNA probe pair. In this platform, naked eye detection of transgenic rice, African swine fever virus (ASFV), and a miRNA can be completed within 1 hour. Our colorimetric gene-sensing method shows superior characteristics, such as probe universality, isothermal reaction conditions, on-site detection capability, and sensitivity that is comparable to that of the fluorescent detection; thus, this method represents a robust next generation gene detection platform.

bioengineering↗

bioRxiv2019

Genomic islets in a sea of introgressions between sympatric species

In the conventional view, species are separate gene pools delineated by reproductive isolation (RI). However, species may also be delineated by merely a small set of "speciation genes" without full RI. It is thus important to know whether "good species" (defined by the "secondary sympatry" test) do continue to exchange genes. Here, we carry out sequencing and de novo high-quality assembly of the genomes of two closely related mangrove species (Rhizophora mucronata and R. stylosa). Whole-genome re-sequencing of individuals across their range on the tropical coasts shows their genomes to be well delineated in allopatry. They became sympatric in northeastern Australia but remain distinct species in contact. Nevertheless, their genomes harbor [~] 4,000 to 10,000 introgression blocks, each averaging only about 3-4 Kb. These fine-grained introgressions indicate that gene flow has continued long after speciation. Non-introgressable "genomic islets," averaging only 1.4 Kb, may contribute to speciation as they often harbor diverging genes underlying flower development and gamete production. In conclusion, RI needs not be the main criterion of species delineation even though all species would eventually be fully reproductively isolated.

evolutionary biology↗

bioRxiv2019

Accelerated trans-sulfuration metabolically defines a discrete subclass of ALS patients

Amyotrophic lateral sclerosis (ALS) is a disease characterized by progressive paralysis and death. Most ALS cases are sporadic (sALS) and patient heterogeneity poses a formidable challenge for the development of viable biomarkers and effective therapies. Applying untargeted metabolite profiling on 77 sALS patient-derived primary dermal fibroblast lines and 45 sex/age matched controls, we found that [~]25% of cell lines (termed sALS-1) are characterized by upregulated trans-sulfuration, where methionine-derived homocysteine is channeled into cysteine and glutathione synthesis. sALS-1 fibroblasts exhibit a growth defect when grown under oxidative conditions, that can be fully-rescued by N-acetylcysteine. [U-13C]-glucose tracing shows that activation of the trans-sulfuration pathway is associated with accelerated glucose flux into the TCA cycle. Based on four metabolites, we developed a support vector machine model capable of distinguishing sALS-1 with 97.5% accuracy. Importantly, plasma metabolite profiling identifies a systemic perturbation of cysteine metabolism as a hallmark of sALS-1. These results indicate that sALS patients can be stratified into distinct metabotypes, differently sensitive to metabolic stress, and provides new insights into metabolic biomarkers for personalized sALS therapy.

neuroscience↗

bioRxiv2019

Integrative analysis identifies key molecular signatures underlying neurodevelopmental deficits in Fragile X Syndrome

Fragile X syndrome (FXS) is an incurable neurodevelopmental disorder with no effective treatment. FXS is caused by epigenetic silencing of FMR1 and loss of FMRP expression. To investigate the consequences of FMRP deficiency in the context of human physiology, we established isogenic FMR1 knockout (FMR1KO) human embryonic stem cells (hESCs). Integrative analysis of the transcriptomic and proteomic profiles of hESC-derived FMRP-deficient neurons revealed several dysregulated pathways important for brain development including processes related to axon development, neurotransmission, and the cell cycle. We functionally validated alterations in a number of these pathways, showing abnormal neural rosette formation and increased neural progenitor cell proliferation in FMR1KO cells. We further demonstrated neurite outgrowth and branching deficits along with impaired electrophysiological network activity in FMRP-deficient neurons. Using isogenic FMR1KO hESC-derived neurons, we reveal key molecular signatures and neurodevelopmental abnormalities arising from loss of FMRP. We anticipate that the FMR1KO hESCs and the neuronal transcriptome and proteome datasets will provide a platform to delineate the pathophysiology of FXS in human neural cells.

neuroscience↗

bioRxiv2019

Cox regression increases power to detect genotype-phenotype associations in genomic studies using the electronic health record

BackgroundThe growth of DNA biobanks linked to data from electronic health records (EHRs) has enabled the discovery of numerous associations between genomic variants and clinical phenotypes. Nonetheless, although clinical data are generally longitudinal, standard approaches for detecting genotype-phenotype associations in such linked data, notably logistic regression, do not naturally account for the times at which events occur. Here we explored the advantages of quantifying associations using Cox proportional hazards regression, which can account for the age at which a patient first visited the healthcare system (left truncation) and the age at which a patient either last visited the healthcare system or acquired a particular phenotype (right censoring).\n\nResultsUsing simulated data, we found that, compared to logistic regression, Cox regression had greater power at equivalent Type I error. We then scanned for genotype-phenotype associations using logistic regression and Cox regression on 50 phenotypes derived from the electronic health records of 49 792 genotyped individuals. In terms of effect sizes, the hazard ratios estimated by Cox regression were nearly identical to the odds ratios estimated by logistic regression. Consistent with the findings from our simulations, Cox regression had approximately 10% greater relative sensitivity for detecting known associations from the NHGRI-EBI GWAS Catalog.\n\nConclusionsAs longitudinal health-related data continue to grow, Cox regression may improve our ability to identify the genetic basis for a wide range of human phenotypes.

bioinformatics↗

bioRxiv2019

Laser Recording of Subcellular Neuron Activities

Advances in imaging and recording of neural activities with a single neuron resolution have played a significant role in understanding neurological diseases in the past decade. Conventional methods relying on patch-clamp and electrodes are regarded as invasive, whereas fluorescence-based imaging tools are useful but still suffer from a low signal-to-noise ratio and low sensitivity. Here we developed a novel optical imaging and recording system by employing laser emissions to record the action potentials in single neurons and neuronal networks caused by subtle transients (Ca2+ concentration) in primary neurons in vitro with a subcellular and single-spike resolution. By recording the laser emissions from neurons, we discovered that lasing emissions could be biologically modulated by intracellular activities and extracellular stimulation with >100-fold improvement in detection sensitivity over traditional fluorescence-based measurement. Finally, we showed that ultrasound can wirelessly activate neurons adsorbed with piezoelectric BaTiO3 nanoparticles, in which the neuron laser emissions were modulated by ultrasound. Our findings show that ultrasound stimulation can significantly increase the lasing intensity and neuron network response. This work not only opens the door to laser emission recording of intracellular dynamics in neuronal networks but may provide an ultra-sensitive detection method for brain-on-chip applications, optogenetics, and neuro-analysis.

bioengineering↗

bioRxiv2019

Streamlined low-input transcriptomics through EASY-RNAseq

High-throughput sequencing for transcriptome profiling is an increasingly accessible and important tool for biological research. However, accurate profiling of small cell populations remains challenging due to issues with gene detection sensitivity and experimental complexity. Here we describe a streamlined RNAseq protocol (EASY RNAseq) for sensitive transcriptome assessment starting from low amount of input materials. EASY RNAseq is technically robust enough for sequencing small pools of homogenous and heterogeneous cells, recovering higher numbers of genes and with a more even expression distribution pattern than other commonly used methods. Application of EASY RNAseq to single human embryos at the 8-cell stage was able to achieve detection of 70% protein-coding genes. This workflow may thus serve as a useful tool for sensitive interrogation of rare cell populations.

genomics↗

bioRxiv2019

siRNA containing a unique 5-nucleotide motif acts as a quencher of IFI16-mediated innate immune response

We previously reported that small interfering RNA (siRNA) enhances DNA or DNA virus mediated-interferon (IFN)-{lambda}1 induction through retinoic acid-inducible gene I (RIG-I) and interferon gamma-inducible protein 16 (IFI16) crosstalk-signalling pathway. Here we provide further evidence of a new role for siRNA. siRNA containing a 5-nucleotide (nt) motif sequence suppresses DNA-mediated IFNs and inflammatory cytokines. We determined that motif siRNA inhibits the induction when the motif is located at the 3 or 5-terminus of siRNA. Using THP1-Lucia ISG cells with various DNA stimulants, it is demonstrated that motif siRNA inhibits DNA or DNA virus but not RNA virus-mediated signaling. Motif siRNA specifically interrupts IFI16 binding to DNA and has 2.5-fold higher affinity to IFI16 than that of siRNA without the motif. Collectively, these findings may shed lights on a novel function of siRNA with the 5-nt motif as a quencher of innate immunity and facilitate the development of potential therapeutics to treat diseases in which this pathway is dysregulated.

immunology↗

bioRxiv2019

Identification and characterization of a novel pic gene cluster responsible for picolinic acid degradation in Alcaligenes faecalis JQ135

Picolinic acid (PA) is a natural toxic pyridine derivative. Microorganisms can degrade and utilize PA for growth. However, the full metabolic pathway and its physiological and genetic foundation remain unknown. In this study, we identified the pic gene cluster responsible for the complete degradation of PA from Alcaligenes faecalis JQ135. PA was initially 6-hydroxylated into 6-hydroxypicolinic acid (6HPA) by PA dehydrogenase (PicA). 6HPA was then 3-hydroxylated by a four-component 6HPA monooxygenase (PicB) to form 3,6-dihydroxypicolinic acid (3,6DHPA), which was then converted into 2,5-dihydroxypyridine (2,5DHP) by a decarboxylase (PicC). The 2,5DHP was further degraded into fumaric acid, through PicD (2,5DHP dioxygenase), PicE (N-formylmaleamic acid deformylase), PicF (maleamic acid amidohydrolase), and PicG (maleic acid isomerase). Homologous pic gene clusters with diverse organizations were found to be widely distributed in -, {beta}-, and{gamma} -Proteobacteria. Our findings provide new insights into the microbial metabolism of environmental toxic pyridine derivatives. ImportancePicolinic acid is a common metabolite of L-tryptophan and some aromatic compounds and is an important intermediate of industrial concern. Although the microbial degradation/detoxification of picolinic acid has been studied for over 50 years, the underlying molecular mechanisms are still unknown. Here, we show the pic gene cluster responsible for the complete degradation of picolinic acid into the tricarboxylic acid cycle. This gene cluster was found to be widespread in other -, {beta}-, and{gamma} -Proteobacteria. These findings provide new perspective for understanding the mechanisms of picolinic acid biodegradation in bacteria.

microbiology↗

bioRxiv2019

Evolution in small steps - A nearly-universal rule of negative selection on codons

The widely accepted view that evolution proceeds in small steps is based on two premises: i) negative selection acts strongly against large differences (Kimura 1983); and ii) positive selection favors small-step changes (Fisher 1930). The two premises are not biologically connected and should be evaluated separately. We now extend the approach of Tang et al. (2004) to codon evolution for the entire genome. Codon substitution rate is a function of the physico-chemical distance between amino acids (AAs), equated with the step size of evolution. This step size depends on a large number of physico-chemical properties as 46 of the 48 properties examined affect the rate. Between 9 pairs of closely-related species of plants, invertebrates and vertebrates, the evolutionary rate is indeed strongly and negatively correlated with the AA distance ({Delta}U, scaled to [0, 1]). While the analyses corroborate the published results that relied on partial genomes, there is an important difference: {Delta}U is strongly correlated with the evolutionary rate (R2 > 0.8) only when the genes are under predominant negative selection. Nevertheless, since most genes in most taxa are strongly constrained by negative selection, {Delta}U would appear to be a nearly-universal measure of codon evolution. In conclusion, the driving force of the small-step evolution at the codon level is negative selection. The unanswered question of whether positive selection may, or may not, follow the small-step rule will be addressed in a companion study (Chen, et al. 2019).

evolutionary biology↗

bioRxiv2019

Molecular evolution in large steps - Codon substitutions under positive selection

Molecular evolution is believed to proceed in small steps. The step size can be defined by a distance reflecting physico-chemical disparities between amino acid (AA) pairs that can be exchanged by single 1 bp mutations. We show that AA substitution rates are strongly and negatively correlated with this distance but only when positive selection is relatively weak. We use the McDonald and Kreitman (MK) test to separate the influences of positive and negative selection. While negative selection is indeed stronger on AA substitutions generating larger changes in chemical properties of amino acids, positive selection operates by different rules. For 65 of the 75 possible pairs, positive selection is comparable in strength regardless of AA distance. However, the 10 pairs under the strongest positive selection all exhibit large leaps in chemical properties. Five of the 10 pairs are shared between hominoids and Drosophila, thus hinting at a common but modest biochemical basis of adaptation across taxa. The hypothesis that adaptive changes often take large functional steps will need to be extensively tested. If validated, molecular models will need to better integrate positive and negative selection in the search for adaptive signal.

evolutionary biology↗

bioRxiv2018

Loss of muscle stem cells in aged mice is replenished by muscle-secreted niche factor G-CSF

Function and number of muscle stem cells (satellite cells, SCs) declines with muscle aging. Although SCs are heterogeneous and different subpopulations have been identified, it remains unknown if a specific subpopulation of muscle SCs selectively decreases during aging. Here, we find Pax7Hi cells are dramatically reduced in aged mice and this aged-dependent loss of Pax7Hi cells is metabolically mediated by myofiber-secreted granulocyte-colony stimulating factor G-CSF as the Pax7Hi SCs are replenished by exercise-induced G-CSF in aged mice. Mechanistically, we show that transcription of G-CSF (Csf3) gene in myofibers is regulated by MyoD in a metabolism-dependent manner and the myofibers-secreted G-CSF acts as a metabolic niche factor required for establishing and maintaining the Pax7Hi SC subpopulation in adult and physiological aged mice by promoting the asymmetric division of Pax7Hi and Pax7Mi SCs. Together, our findings uncover a metabolic niche role of muscle metabolism in regulating Pax7 SC heterogeneity in mice.\n\nHighlightsO_LISingle cell RNA-seq unveils Pax7Hi and Pax7Lo cells are two distinct subpopulations.\nC_LIO_LIPax7Hi SCs are enriched in glycolytic fibers and reduced in aging muscle.\nC_LIO_LIMetabolic niche factor G-CSF is required for regulating dynamic change of Pax7 SCs.\nC_LIO_LIG-CSF replenishes Pax7Hi cells by stimulating asymmetric division of Pax7Mi cells.\nC_LI

cell biology↗

bioRxiv2018

APOE4 is Associated with Differential Regional Vulnerability to Bioenergetic Deficits in Aged APOE Mice

The {varepsilon}4 allele of apolipoprotein E (APOE) is the dominant genetic risk factor for late-onset Alzheimers disease (AD). However, the reason for the association between APOE4 and AD remains unclear. While much of the research has focused on the ability of the apoE4 protein to increase the aggregation and decrease the clearance of A{beta}, there is also an abundance of data showing that APOE4 negatively impacts many additional processes in the brain, including bioenergetics. In order to gain a more comprehensive understanding of the APOE4s role in AD pathogenesis, we performed a multi-omic analysis of APOE4 vs. APOE3 expression in the entorhinal cortex (EC) and primary visual cortex (PVC) of aged APOE mice. These studies revealed region-specific alterations in several bioenergetic pathways, including oxidative phosphorylation (OxPhos), the TCA-cycle and fatty acid metabolism. Follow-up analysis utilizing the Seahorse platform revealed decreased mitochondrial respiration in the hippocampus and cortex of aged APOE4 vs. APOE3 mice, but not in the EC of these mice. Additional studies, as well as the original multi-omic data suggest that bioernergetic pathways in the EC of aged APOE mice may be differentially regulated by APOE4 expression. Given the importance of the EC as one of the first regions to be affected by AD pathology in humans, this differential bionenergetic regulation observed in the EC vs. other brain regions of aged APOE4 mice may play an important role in the pathogenesis of AD, particularly among APOE4 carriers.

neuroscience↗

bioRxiv2018

Spatiotemporal expression of FRIGIDA modulate flowering time in Arabidopsis thaliana

FRIGIDA (FRI) as the major regulator of flowering time in Arabidopsis accessions can activate its target FLOWERING LOCUS C (FLC) to delay flowering before vernalization. Besides FLC, other FRI targets also exist in Arabidopsis. Although leaves sense environmental cues to modulate flowering time, it is not known if roots also regulate the floral transition. In this study, we investigated the spatiotemporal effect of FRI on flowering time. Local expression of FRI in the phloem and leaves activated FLC to delay flowering. Furthermore, we found that local expression of FRI in the roots also delayed flowering by activating other targets MADS AFFECTING FLOWERING4 (MAF4) and MAF5 in the roots. Graft and genetic experiments revealed that the spatial expression of FRI in the root might generate a mobile signal, which is transmitted from roots to shoot and antagonizes the FT signal to delay flowering. Specifically expressing FRI in the embryo efficiently delayed flowering, even expressing FRI as early as pro-embryo stage is enough to upregulate FLC expression to delay flowering. Together, our findings confirm the spatiotemporal effect of FRI on delaying flowering, and propose that root tissue also perceives the flowering signal to fine-tune the flowering time through MAF4/5 as novel targets of FRI.\n\nHighlightRoot FRIDIGA activated the novel targets MAF4/5 to delay flowering; Temporal expressing FRIGIDA at as early as pro-embryo stage is efficient to delay flowering.

plant biology↗

bioRxiv2018

Molecular profiles and mutation burden analysis in Chinese patients with gastric carcinoma

The goal of this work was to investigate the molecular profiles and mutation burden in Chinese patients with gastric carcinoma (GC). In total, we performed whole exome sequencing (WES) on 74 GC patients with tumor and adjacent normal formalin-fixed, paraffin-embedded (FFPE) tissue samples. The mutation spectrum of these samples showed a high concordance with TCGA and other studies on GC. We found the alterations of 17 DNA repair genes (including BRCA2, POLE and MSH3, etc.) were strongly correlated with the tumor mutation burden (TMB) and tumor neoantigen burden (TNB) of GC patients. Patients with mutations of these genes tend to have high TMB (median of TMB = 12.77, p=2.3e-6) and TNB (median of TNB = 5.97, p= 2.8e-3). In addition, younger GC patients (age < 60) have lower TMB (p = 0.0021) and TNB (p = 0.034) than older patients (age >= 60). Furthermore, we found a list of 18 genes and two genomic regions (1p36.21 and Xq26.3) were associated with peritoneal metastasis (PM) of GC, and patients with amplification of 1p36.21 and Xq26.3 have a worse prognosis (p=0.002, 0.01, respectively). Our analysis provides GC patients with potential markers for single and combination therapies.

cancer biology↗

bioRxiv2018

CRISPR/Cas9 screens Reveal Dasatinib Targets of Inhibiting T cell Activation and Proliferation

Immune response by T cells is essential for a healthy body against cancer, infection, and pathophysiological alteration. The activation and expansion of T cells can be inhibited by dasatinib, a tyrosine inhibitor, thus improving the outcome of diseases, such as autoimmune disease, graft-versus-host disease, and transplant rejection. The underlying mechanism of inhibition by dasatinib is elusive. Here, we designed and synthesized a CRISPR/Cas9 screening library that includes 6,149 genes. Using the library, we performed dasatinib CRISPR/cas9 screening in Jurkat cell, a T lymphocyte cell. We firstly identified survival essential genes for Jurkat cells. Comparing with other CRISPR/Cas9 screenings, we obtained Jurkat cell specific essential genes. By comparing dasatinib treatment to control, we identified a set of dasatinib targets, which includes known targets: CSK, LCK, ZAP70, and previously unknown targets: ZFP36L2, LRPPRC, CFLAR, PD-1, CD45 et al. Visualizing these target genes on T cell receptor signaling pathway, we found several genes could be inhibited by dasatinib. Furthermore, we introduced a framework, 9-square, to classify genes and found a group of genes that are associated with dasatinib resistance, possibly linking the side effects of dasatinib. These data reveal a set of dasatinib targets and demonstrate the molecular potential functions of dasatinib. Identification of dasatinib targets will broaden our understanding to its molecular mechanism, and thus benefits to clinical outcome.

cancer biology↗

bioRxiv2018

Quantitative Analysis of Interactive Behavior of Mitochondria and Lysosomes Using Structured Illumination Microscopy

Super-resolution optical microscopy has extended the spatial resolution of cell biology from the cellular level to the nanoscale, enabling the observation of the interactive behavior of single mitochondria and lysosomes. Quantitative parametrization of interaction between mitochondria and lysosomes under super-resolution optical microscopy, however, is currently unavailable, which has severely limited our understanding of the molecular machinery underlying mitochondrial functionality. Here, we introduce an M-value to quantitatively investigate mitochondria and lysosome contact (MLC) and mitophagy under structured illumination microscopy. We found that the M-value for an MLC is typically less than 0.4, whereas in mitophagy it ranges from 0.5 to 1.0. This system permits further investigation of the detailed molecular mechanism governing the interactive behavior of mitochondria and lysosomes.

cell biology↗

bioRxiv2018

Bromodomain Protein, BRD4, Contributes to the Regulation of Alternative Splicing

Bromodomain protein 4 (BRD4) is an atypical kinase and a histone acetyl transferase (HAT) which plays an important role in chromatin remodeling and early transcriptional elongation. During transcription elongation, BRD4 travels with the elongation complex. Since most of the alternative splicing events take place co-transcriptionally, we asked if BRD4 plays a role in regulation of alternative splicing. We find that distinct patterns of alternative splicing are associated with conditional deletion of BRD4 during thymocyte differentiation in vivo. Similarly, depletion of BRD4 in T-ALL cells alters patterns of splicing. Most of the alternatively spliced events affected by BRD4 are usage of exon skipping. In an established insulin receptor minigene model of splicing, BRD4 over expression modulates alternative splicing. Importantly, as assessed by both immunoprecipitation (IP) and proximity ligation (PLA) assays, BRD4 interacts with components of the splicing machinery. BRD4 also co-localizes on chromatin with one of the splicing regulators. We propose that BRD4 contributes to patterns of alternative splicing through its interaction with the splicing machinery during transcription elongation.\n\nSignificance StatementThe bromodomain protein, BRD4, is a transcriptional and epigenetic regulator that plays a critical role in both cancer and inflammation. It has pleiotropic activities, including chromatin organization, transcriptional pause release and initiation. We now report that it also contributes to the regulation of alternative splicing. Taken together, these findings indicate that BRD4 functions to coordinate the various steps in gene expression.

molecular biology↗