Biology subjects

Zhang, C.

Publications and source records attributed to Zhang, C..

At least 919 records · Page 51Linked to original sources

bioRxiv2020

WISP2/CCN5 gene knockdown in vitro and in vivo exhibits proliferation promotion of breast cancer through targeting Skp2 and p27Kip1

BackgroundEmerging evidence has demonstrated that WISP2/CCN5 is critically involved in tumorigenesis. However, the function of WISP2/CCN5 in breast cancer carcinogenesis is largely unclear. Methodswe aim to explore the effects and potential mechanisms of WISP2/CCN5 on proliferation of breast cancer cells and carcinogenesis of breast cancer xenograft. Lentivirus vector with WISP2/CCN5shRNA was transfected into MCF-7, and breast cancer cells and xenograft were conducted. Effect of WISP2/CCN5 on growth and carcinogenesis of breast cancer cells and xenografts was evaluated by MTT assay and tumor volume. The relationship between WISP2/CCN5, Skp2 and p27Kip1 was detected in vitro and in vivo by RT-PCR at mRNA level and Western blotting at protein level. ResultsThe result of MTT assay indicated that MCF-7 cell growth viability in WISP2/CCN5 gene knockdown group was significantly higher than negative vector group(P<0.05) or control group (P<0.05). It suggested that knockdown of WISP2/CCN5 gene by shRNA lentivirus plasmid promoted proliferation of MCF-7 cells. The growth curves of breast cancer xenograft showed that xenografts in WISP2/CCN5 knockdown group grew more quickly than negative vector group(P< 0.05) or control group (P< 0.05). Subsequently, the results of RT-PCR and Western blotting revealed that WISP2/CCN5 gene knockdown led to increased Skp2 and decreased p27Kip1 at mRNA and protein levels. WISP2/CCN5 exerts its inhibition on proliferation of MCF-7 cell line and suppressive functions on growth of breast carcinoma via regulation of Skp2 and p27Kip1at mRNA and protein levels. However, WISP2/CCN5 gene knockdown resulted in loss of inhibition effect on MCF-7 and breast cancer. ConclusionsOur findings suggest that WISP2/CCN5 could be a useful therapeutic strategy for the treatment of breast cancer through targeting Skp2 and p27Kip1.

molecular biology↗

bioRxiv2020

Origin time and epidemic dynamics of the 2019 novel coronavirus

The 2019 novel coronavirus (2019-nCoV) have emerged from Wuhan, China. Studying the epidemic dynamics is crucial for further surveillance and control of the outbreak. We employed a Bayesian framework to infer the time-calibrated phylogeny and the epidemic dynamics represented by the effective reproductive number (Re) changing over time from 33 genomic sequences available from GISAID. The time of the most recent common ancestor (MRCA) was December 17, 2019 (95% HPD: December 7, 2019 - December 23, 2019). The median estimate of Re shifted from 1.6 to 1.1 on around January 1, 2020. This study provides an early insight of the 2019-nCoV epidemic. However, due to limited amount of data, one should be cautious when interpreting the results at this stage.

microbiology↗

bioRxiv2020

Construction of Nanobody Library in Mammalian Cells by Linear-double-stranded DNA Based AND Gate Genetic Circuit

Nanobody is one special type of single-domain antibody fragment with multiple advantages over traditional antibody. Our previous work established linear-double-stranded DNA (ldsDNA, or PCR amplicon) as novel biological parts for building AND gate genetic circuits in mammalian cells. During this AND-gate circuit formation process, the co-transfected up- and down-stream ldsDNAs could be linked together to form intact gene expression cassette. Here, we employed this ldsDNA-based AND-gate (LBAG) strategy to construct nanobody library in mammalian cells. The sequence complexity of complementary determining regions (CDRs) was introduced into ldsDNA by PCR amplification. After being co-transfected into mammalian cells, the up- and down-stream ldsDNAs undergo AND gate linkage and form full nanobody coding regions, containing CDR1-3. High throughput sequencing identified 22,173 unique oligonucleotide sequences in total generated by this strategy. Thus, we developed a novel method to construct nanobody library, which is a start point for building high content nanobody library in mammalian cells.

immunology↗

bioRxiv2020

AmpliconReconstructor: Integrated analysis of NGS and optical mapping resolves the complex structures of focal amplifications in cancer

Oncogene amplification, a major driver of cancer pathogenicity, is often mediated through focal amplification of genomic segments. Recent results implicate extrachromosomal DNA (ecDNA) as the primary mechanism driving focal copy number amplification (fCNA) - enabling gene amplification, rapid tumor evolution, and the rewiring of regulatory circuitry. Resolving an fCNAs structure is a first step in deciphering the mechanisms of its genesis and the subsequent biological consequences. Here, we introduce a powerful new computational method, AmpliconReconstructor (AR), for integrating optical mapping (OM) of long DNA fragments (>150kb) with next-generation sequencing (NGS) to resolve fCNAs at single-nucleotide resolution. AR uses an NGS-derived breakpoint graph alongside OM scaffolds to produce high-fidelity reconstructions. After validating performance by extensive simulations, we used AR to reconstruct fCNAs in seven cancer cell lines to reveal the complex architecture of ecDNA, breakage-fusion-bridge cycles, and other complex rearrangements. By distinguishing between chromosomal and extrachromosomal origins, and by reconstructing the rearrangement signatures associated with a given fCNAs generative mechanism, AR enables a more thorough understanding of the origins of fCNAs, and their functional consequences.

bioinformatics↗

bioRxiv2020

Chinese Glioma Genome Atlas (CGGA): A Comprehensive Resource with Functional Genomic Data for Chinese Glioma Patients

Gliomas are the most common and malignant intracranial tumours in adults. Recent studies have shown that functional genomics greatly aids in the understanding of the pathophysiology and therapy of glioma. However, comprehensive genomic data and analysis platforms are relatively limited. In this study, we developed the Chinese Glioma Genome Atlas (CGGA, http://www.cgga.org.cn), a user-friendly data portal for storage and interactive exploration of multi-dimensional functional genomic data that includes nearly 2,000 primary and recurrent glioma samples from Chinese cohorts. CGGA currently provides access to whole-exome sequencing (286 samples), messenger RNA sequencing (1,018 samples) and microarray (301 samples), DNA methylation microarray (159 samples), and microRNA microarray (198 samples) data, as well as detailed clinical data (e.g., WHO grade, histological type, critical molecular genetic information, age, sex, chemoradiotherapy status and survival data). In addition, we developed an analysis tool to allow users to browse mutational, mRNA/microRNA expression, and DNA methylation profiles and perform survival and correlation analyses of specific glioma subtypes. CGGA greatly reduces the barriers between complex functional genomic data and glioma researchers who seek rapid, intuitive, and high-quality access to data resources and enables researchers to use these immeasurable data sources for biological research and clinical application. Importantly, the free provision of data will allow researchers to quickly generate and provide data to the research community.

genomics↗

bioRxiv2020

Ultra-deep sequencing differentiates patterns of skin clonogenic mutations associated with sun-exposure status

Non-melanoma skin cancer is the most common human malignancy and is primarily caused by exposure to ultraviolet (UV) radiation. The earliest detectable precursor of UV-mediated skin cancer is the growth of cell groups harboring clonal mutation (CM) in clinically normal appearing skin. Systematic evaluation of CMs is crucial to understand early photo-carcinogenesis. Previous studies confirmed the presence of CMs in sun-exposed skin. However, the relationship between UV-exposure and the accumulation of CMs, and the correlation of CMs with skin cancer risk remain poorly understood. To elucidate the exact molecular and clinical effects of long-term UV-exposure on skin, we performed targeted ultra-deep sequencing in 450 individual-matched sun-exposed (SE) and non-sun-exposed (NE) epidermal punch biopsies obtained from clinically normal skin from 13 donors. A total of 638 CMs were identified, including 298 UV-signature mutations (USMs). The numbers of USMs per sample were three times higher in the SE samples and were associated with significantly higher variant allele frequencies (VAFs), compared with the NE samples. We identified genomic regions in TP53, NOTCH1 and GRM3 where mutation burden was significantly associated with UV-exposure. Six mutations were almost exclusively present in SE epidermis and accounted for 42% of the overall difference between SE and NE mutation burden. We defined Cumulative Relative Clonal Area (CRCA), a single metric of UV-damage calculated by the overall relative percentage of the sampled skin area affected by CMs. The CRCA was dramatically elevated by a median of 11.2 fold in SE compared to NE samples. In an extended cohort of SE normal skin samples from patients with a high- or low-burden of cutaneous squamous cell carcinoma (cSCC), the SE samples in high-cSCC patients contained significantly more USMs than SE samples in low-cSCC patients, with the difference mostly conferred by mutations from low-frequency clones (defined by VAF≤1%) but not expanded clones (VAF>1%). Our studies of differential mutational features in normal skin between paired SE/NE body sites and high/low-cSCC patients provide novel insights into the carcinogenic effect of UV exposure, and suggest CMs might be used to develop novel biomarkers for predicting cancer risk.Significance statement In UV radiation exposed skin, mutations fuel clonal cell growth. We established a sequencing-based method to objectively assess the mutational differences between sun-exposed (SE) and non-sun-exposed (NE) areas of normal human skin. Striking differences, in both the numbers of mutations and variant allele frequencies, were found between SE and NE areas. Furthermore, we identified specific genomic regions where mutation burden is significantly associated with UV-exposure status. These findings revealed previously unknown mutational patterns associated with UV-exposure, providing important insights into UV radiation’s early carcinogenic effects. Additionally, in an extended cohort, we identified preliminary association between normal skin mutation burden and cancer risk. These findings pave the road for future development of quantitative measurement of subclinical UV damage and skin cancer risk.Competing Interest StatementThe authors have declared no competing interest.Common AbbreviationsUVUltravioletCMClonogenic mutationNMSCNonmelanoma skin cancerSESun-exposedNENon-sun-exposedUSMUV-signature mutationNUSMNon-UV-signature mutationCRCACumulative Relative Clonal AreacSCCCutaneous squamous cell carcinomaAKActinic keratosisSNVSingle nucleotide variant Indels – Insertions/deletionsDNVDinucleotide variantCSNVCluster of single nucleotide variantMACMulti-Nucleotide Variant Annotation CorrectorVAFVariant allele frequencyView Full Text

cancer biology↗

bioRxiv2020

MSIsensor-pro: fast, accurate and matched-normal-sample-free detection of microsatellite instability

We developed MSIsensor-pro (https://github.com/xjtu-omics/msisensor-pro), an open-source single sample microsatellite instability (MSI) scoring method for research and clinical applications. MSIsensor-pro introduces a multinomial distribution model to quantify polymerase slippages for each tumor sample and a discriminative sites selection method to enable MSI detection without matched normal samples. For samples of various sequencing depths and tumor purities, MSIsensor-pro significantly outperformed the current leading methods in terms of both accuracy and computational cost.

bioinformatics↗

bioRxiv2020

Single cell atlas of trisomy 21 cerebral cortex

Down syndrome (DS) is one of the most common human birth defects caused by trisomy 21 (T21), leading to a variety of cognitive impairments. The cellular composition of human brain has been explored using single cell RNA sequencing in both physiological and pathological conditions. However, the cellular heterogeneity of human brain with chromosome aneuploidy is largely unknown. Here, we profiled the transcriptome of 36046 cells in cerebral cortex of T21 human fetus, covering frontal lobe, parietal lobe, occipital lobe and temporal lobe. Intriguingly, we detected several genes positively associated with neurons maturation was dysregulated in T21 frontal cortex (HIC2, POU2F2, ZGLP1 and FOXK1). To share, explore and utilized the data resources of T21 cerebral cortex, we developed a comprehensive platform named T21atlas, composing of two functional modules (T21cluster and T21talk). Overall, our study provides, as far as we know, the first single cell atlas for T21 cerebral cortex, which could promote our understanding of the molecular mechanism of DS at an unprecedented resolution and could potentially facilitate the development of novel clinical therapeutics against T21.

neuroscience↗

bioRxiv2019

Microbial microdroplet culture system (MMC): an integrated platform for automated, high-throughput microbial cultivation and adaptive evolution

Conventional microbial cell cultivation techniques are typically labor intensive, low throughput, and poor parallelization, rendering them inefficient. The development of automated, modular microbial cell micro-cultivation systems, particularly those employing droplet microfluidics, has gained attention for their high-throughput, parallel and highly efficient cultivation capabilities. Here, we report the development of a microbial microdroplet culture system (MMC), which is an integrated platform for automated, high-throughput cultivation and adaptive evolution of microorganisms. We demonstrated that the MMC yielded both accurate and reproducible results for the manipulation and detection of droplets. The superior performance of MMC for microbial cell cultivation was validated by comparing the growth curves of six microbial strains grown in MMC, conventional shake flasks or well plates. The highest incipient growth rate for all six microbial cell lines was achieved using MMC. We also conducted an 18-day process of adaptive evolution of a methanol-essential Escherichia coli strain in MMC and obtained two strains exhibiting higher growth rates compared with the parent strain. Our study demonstrates the power of MMC to provide an efficient and reliable approach for automated, high-throughput microbial cultivation and adaptive evolution.

bioengineering↗

bioRxiv2019

Boron deficiency-induced root growth inhibition is mediated by BR signal regulation in Arabidopsis

Brassinosteroid (BR) is a pivotal phytohormone involved in regulating root development. Boron (B) is an essential micronutrient for plant growth and development, and root growth of plants is rapidly inhibited under B deficiency condition, but the mechanisms are still elusive. Here, we demonstrate that BR plays crucial roles in these processes. We identify BR-related processes underlying B deficiency at the physiological, genetic, molecular/cell biological and transcriptome levels, and provide strong evidences that B deficiency can affect BR signalling, thereby altering root growth. RNA-sequencing analysis reveals a high co-regulation between BR-regulated genes and B deficiency-responsive genes. We found that low B negatively regulates BR signalling to control BR signalling-dependent root elongation, bes1-D exhibits insensitivity to low B stress, and bri1-301 mutants fails to respond to B depletion. Exogenous eBL application can rescue the inhibition of root growth under B deficiency condition, and application of BR biosynthesis inhibitor BRZ aggravates root growth inhibition of wild-type under B deficiency condition. B deficiency reduces the nuclear signal of BES1. We further found that B deficiency reduces the accumulation of brassinolide (BL) by reducing BR6ox1 and BR6ox2 mRNA level to down-regulate BR signalling and modulate root elongation. Altogether, our results uncover a role of BR signalling in root elongation under B deficiency. One-sentence summaryB deficiency reduces the accumulation of brassinolide by reducing BR6ox1 and BR6ox2 mRNA level to down-regulate BR signalling and modulate root elongation.

physiology↗

bioRxiv2019

ASTRAL-Pro: quartet-based species tree inference despite paralogy

Species tree inference via summary methods that combine gene trees has become an increasingly common analysis in recent phylogenomic studies. This broad adoption has been partly due to the greater availability of genome-wide data and ample recognition that gene trees and species trees can differ due to biological processes such as gene duplication and gene loss. This increase has also been encouraged by the recent development of accurate and scalable summary methods, such as ASTRAL. However, most of these methods, including ASTRAL, can only handle single-copy gene trees and do not attempt to model gene duplication and gene loss. In this paper, we introduce a measure of quartet similarity between single-copy and multi-copy trees (accounting for orthology and paralogy relationships) that can be optimized via a scalable dynamic programming similar to the one used by ASTRAL. We then present a new quartet-based species tree inference method: ASTRAL-Pro (ASTRAL for PaRalogs and Orthologs). By studying its performance on an extensive collection of simulated datasets and on a real plant dataset, we show that ASTRAL-Pro is more accurate than alternative methods when gene trees differ from the species tree due to the simultaneous presence of gene duplication, gene loss, incomplete lineage sorting, and estimation errors.

bioinformatics↗

bioRxiv2019

In vitro induced floral reversion in switchgrass (Panicum Virgatum L.)

Switchgrass (Panicum Virgatum L.) is a warm-season perennial grass native to North America, it was used as forage and vegetative filter strips in early days, and have developed into a bioenergy crop in recent years. In this study, we found that the switchgrass cultivar Alamo at elongation stage 4 have developed inflorescences about 1 cm in length, and in vitro incubation of the shoot apexes harboring inflorescences on Murashige and Skoogs basal medium supplemented with 3 mg/L 6-benzylaminopurine generated multiple shoot clumps. Anatomical study showed that some of the regenerated shoots originated from axillary buds on the explants, some of them originated from adventurous buds and some of them originated from young florets. Further study of shoots originated from young florets found that the floral organs degenerated or developed into leaf-like organs, and the flower terminal transformed into a vegetative shoot apical meristem, thats to say these shoots arise from flower reversion. In vitro induction of floral reversion provided a novel protocol to manipulate flower development in switchgrass, which might contribute a fundamental for flower development study in switchgrass and other plants.

developmental biology↗

bioRxiv2019

A screening platform based on epitope editing for drug discovery

The interaction between an antibody and its epitope has been daily utilized in various biological studies; however it has been rarely explored whether small molecules can alter the interaction. We discovered that small molecules could alter/edit surface properties of amyloid beta (A{beta}) epitopes, and consequently inhibit or enhance corresponding antibody recognition. Remarkably, this editing effect could generate functional changes including protein aggregation behaviors, cell cytokine secreting and in vivo microglia activation. According to this discovery, we proposed a screen platform based on epitope editing for drug discovery (SPEED). With a small library of compounds, we validated that SPEED could be used to seek new leads for A{beta} species. We also demonstrated that this platform could potentially be extended to other targets including tau protein and PD-L1 protein. The SPEED is a simple, fast and label-free screening method. We believe that the SPEED strategy could be universally applicable for seeking and validating drug candidates and imaging ligands.

biochemistry↗

bioRxiv2019

Genomic Characterization of SiGRFs in Foxtail Millet and SiGRF1-overexpression in Arabidopsis thaliana Promotes Plant Avoidance of Salt Stress

In plants, 14-3-3 proteins are recognized as mediators of signal transduction and function in both development and stress response. However, their functions have not been reported in the C4 crop foxtail millet. Here, phylogenetic analysis categorized foxtail millet 14-3-3s (SiGRFs) into ten discrete groups (Clusters I to {square}). Transcriptome and qPCR analyses showed that all the SiGRFs responded to at least one abiotic stress. All but one SiGRF-overexpressing (OE) Arabidopsis thaliana line (SiGRF1) exhibited insensitivity to abiotic stresses during seed germination and seedling growth. Compared with the Col-0 wild-type, SiGRF1-OEs had slightly lower germination rates and smaller leaves. However, flowering time of SiGRF1-OEs occurred earlier than that of Col-0 under high-salt stress. Interaction of SiGRF1 with a foxtail millet E3 ubiquitin-protein ligase (SiRNF1/2) indicates that the proteinase system might hydrolyse SiGRF1. Further investigation showed that SiGRF1 localized in the cytoplasm, and its gene was ubiquitously expressed in various tissues throughout various developmental stages. Additionally, flowering-related genes, WRKY71, FLOWERING LOCUS T, LEAFY and FRUITFULL, in SiGRF1-OEs exhibited considerably higher expression levels than those in Col-0 under salinity-stressed conditions. Results suggest that SiGRF1 hastens flowering, thereby providing a means for foxtail millet to complete its life cycle and avoid further salt stress. HighlightSiGRFs in foxtail millet: SiGRF1 hastens flowering in transgenic Arabidopsis thaliana exposed to salt stress

genomics↗

bioRxiv2019

Classification of clear cell renal cell carcinoma based on PKM alternative splicing

Clear cell renal cell carcinoma (ccRCC) accounts for 70-80% of kidney cancer diagnoses and displays high molecular and histologic heterogeneity. Hence, it is necessary to reveal the underlying molecular mechanisms involved in progression of ccRCC to better stratify the patients and design effective treatment strategies. Here, we analyzed the survival outcome of ccRCC patients as a consequence of the differential expression of four transcript isoforms of the pyruvate kinase muscle type (PKM). We first extracted a classification biomarker consisting of eight gene pairs whose within-sample relative expression orderings (REOs) could be used to robustly classify the patients into two groups with distinct molecular characteristics and survival outcomes. Next, we validated our findings in a validation cohort and an independent Japanese ccRCC cohort. We finally performed drug repositioning analysis based on transcriptomic expression profiles of drug-perturbed cancer cell lines and proposed that paracetamol, nizatidine, dimethadione and conessine can be repurposed to treat the patients in one of the subtype of ccRCC whereas chenodeoxycholic acid, fenoterol and hexylcaine can be repurposed to treat the patients in the other subtype.

systems biology↗

bioRxiv2019

Integrated Sequencing & Array Comparative Genomic Hybridization in Familial Parkinson’s Disease

BackgroundParkinsons disease (PD) is a genetically heterogeneous condition; both single nucleotide variants (SNVs) and copy number variants (CNVs) are important genetic risk factors. We examined the utility of combining exome sequencing and genome-wide array-based comparative genomic hybridization (aCGH) for identification of PD genetic risk factors. MethodsWe performed exome sequencing on 110 subjects with PD and a positive family history; 99 subjects were also evaluated using genome-wide aCGH. We interrogated exome sequencing and array comparative genomic hybridization data for pathogenic SNVs and CNVs at Mendelian PD gene loci. SNVs were confirmed via Sanger sequencing. CNVs were confirmed with custom-designed high-density aCGH, droplet digital PCR, and breakpoint sequencing. ResultsUsing exome sequencing, we discovered individuals with known pathogenic single nucleotide variants in GBA (p.E365K, p.T408M, p.N409S, p.L483P) and LRRK2 (p.R1441G and p.G2019S). Two subjects were each double heterozygotes for variants in GBA and LRRK2. Based on aCGH, we additionally discovered cases with an SNCA duplication and heterozygous intragenic GBA deletion. Five additional subjects harbored both SNVs (p.N52fs, p.T240M, p.P437L, p.W453*) and likely disrupting CNVs at the PARK2 locus, consistent with compound heterozygosity. In nearly all cases, breakpoint sequencing revealed microhomology, a mutational signature consistent with CNV formation due to DNA replication errors. ConclusionsIntegrated exome sequencing and aCGH yielded a genetic diagnosis in 19.3% of our familial PD cohort. Our analyses highlight potential mechanisms for SNCA and PARK2 CNV formation, uncover multilocus pathogenic variation, and identify novel SNVs and CNVs for further investigation as potential PD risk alleles.

genetics↗

bioRxiv2019

α-ketoglutaric acid stimulates muscle hypertrophy and fat loss through OXGR1-dependent adrenal activation

Beneficial effects of resistance exercise on metabolic health and particularly muscle hypertrophy and fat loss are well established, but the underlying chemical and physiological mechanisms are not fully understood. Here we identified a myometabolite-mediated metabolic pathway that is essential for the beneficial metabolic effects of resistance exercise in vivo. We showed that substantial accumulation of the tricarboxylic acid cycle intermediate -ketoglutaric acid (AKG) is a metabolic signature of resistance exercise performance. Interestingly, human plasma AKG level is also negatively correlated with BMI. Pharmacological elevation of circulating AKG induces muscle hypertrophy, brown adipose tissue (BAT) thermogenesis, and white adipose tissue (WAT) lipolysis in vivo. We further found that AKG stimulates the adrenal release of adrenaline through 2-oxoglutarate receptor 1 (OXGR1) expressed in adrenal glands. Finally, by using both loss-of-function and gain-of-function mouse models, we showed that OXGR1 is essential for AKG-mediated exercise-induced beneficial metabolic effects. These findings reveal an unappreciated mechanism for the salutary effects of resistance exercise, using AKG as a systemically-derived molecule for adrenal stimulation of muscle hypertrophy and fat loss.

molecular biology↗

bioRxiv2019

LIMD1 phase separation contributes to cellular mechanics and durotaxis by regulating focal adhesion dynamics in response to force

The mechanical environement affects cell morphology, differentiation and motility. The ability of cells to follow gradients of extracellular matrix stiffness-durotaxis has been implicated in development, fibrosis, and cancer. Cells sense and respond to extra-cellular mechanical cues through cell-matrix adhesions. Interestingly, the maturation of focal adhesions (FAs) is reciprocally force-dependent. How biomechanical cues dictate the status of cell motility and how FAs coordinate force sensing and self-organization remain enigmatic. LIMD1, a member of the LIM domain proteins, localizes to the FAs and has been reported to negatively regulate the Hippo-YAP pathway in response to tension. Here we identify the force sensitive recruitment of LIMD1 to the FAs. We discover that LIMD1 regulates cell spreading, maintains FA dynamics and cellular force, and is critical for durotaxis. Intriguingly, LIMD1 selectively recruits late but not early FA proteins through phase separation at the FAs under force. We suggest a model in which localization of LIMD1 to the FAs, triggered by mechanical force, serves as a phase separation hub for assembling and organizing late FA proteins, allowing for effective FA maturation and efficient cellular mechano-transduction.

cell biology↗