Biology subjects

Ma, G.

Publications and source records attributed to Ma, G..

51 records · Page 3Linked to original sources

bioRxiv2021

Three bacterium-plasmid golden combinations facilitate the spread of ST11/CG258 carbapenemase-producing Klebsiella pneumoniae in China

Carbapenemase-producing Klebsiella pneumoniae (cpKP) poses serious threats to public health. Previous studies showed that only ST11/CG258-cpKP successfully disseminated in China, however, the underlying genetic bases are still unknown. We conducted a comprehensive genomic-epidemiology analysis on 420 cpKP isolates from 70 hospitals in 24 Chinese provinces during 2009-2017 based on short-/long-reads sequencing. Three golden combinations of host--blaKPC-carrying plasmids (Clade 3.1+3.2--IncFIIpHN7A8, Clade 3.1+3.2--IncFIIpHN7A8:IncR, Clade 3.3--IncFIIpHN7A8:IncpA1763-KPC) endowed cpKP with advantages both in genotypes (strong-correlation/co-evolution) and phenotypes (resistance/growth/competition), thereby facilitating nationwide spread of ST11/CG258-cpKP. Intriguingly, Bayesian skyline illustrated that the three golden combinations might directly lead to the strong population expansion during 2007-2008 and subsequent maintenance of the dissemination of ST11/CG258-cpKP after 2008. We tested drug-resistance profiles and proposed combination treatment regimens for CG258/non-CG258 cpKP. Our findings systematically revealed the molecular-epidemiology and genetic-basis for dissemination of Chinese ST11/CG258 cpKP and reminded us to monitor the golden combinations of cpKP- plasmid closely.

microbiology↗

bioRxiv2021

A programmable system to methylate and demethylate m6A on specific mRNAs

RNA N6-Methyladenosine (m6A) is the most abundant mRNA modification, and forms part of an epitranscriptomic system that modulates RNA function. RNA modifications can be reversibly catalyzed by several specific enzymes, and those modifications can be recognized by RNA binding proteins that in turn regulate biological processes. Although there are many reports demonstrating m6A participation in critical biological functions, this exploration has mainly been conducted through the global knockout or knockdown of the writers, erasers, or readers of m6A. Consequently, there is a lack of information about the role of m6A on single transcripts in biological processes, posing a challenge in understanding the biological functions of m6A. Here, we demonstrate a CRISPR/dCas13a-based RNA m6A-editor which can target mRNAs using single crRNA or multiple crRNAs array to methylate or demethylate m6A. We systematically assay its capabilities to enable the targeted rewriting of m6A dynamics, including modulation of circular RNA translation and transcript half-life. Finally, we demonstrate the utility of the system by specifically modulating XIST m6A levels, which can control X chromosome silencing and activation. Based on our editors, m6A on single and multiple transcripts can be modified to allow the exploration of the role of m6A on in biological processes.

cell biology↗

bioRxiv2020

Transposable Element-Gene Splicing Modulates the Transcriptional Landscape of Human Pluripotent Stem Cells

Transposable elements (TEs) occupy nearly 50% of mammalian genomes and are both potential dangers to genome stability and functional genetic elements. TEs can be expressed and exonised as part of a transcript, however, their full contribution to the transcript splicing remains unresolved. Here, guided by long and short read sequencing of RNAs, we show that 26% of coding and 65% of non-coding transcripts of human pluripotent stem cells (hPSCs) contain TEs. Different TE families have unique integration patterns with diverse consequences on RNA expression and function. We identify hPSC-specific splicing of endogenous retroviruses (ERVs) as well as LINE L1 elements into protein coding genes that generate TE-derived peptides. Finally, single cell RNA-seq reveals that proliferating hPSCs are dominated by ERV-containing transcripts, and subpopulations express SINE or LINE-containing transcripts. Overall, we demonstrate that TE splicing modulates the pluripotency transcriptome by enhancing and impairing transcript expression and generating novel transcripts and peptides.

genomics↗

bioRxiv2020

Unveiling transposable element expression heterogeneity in cell fate regulation at the single-cell level

Transposable elements (TEs) make up a majority of a typical eukaryotes genome, and contribute to cell heterogeneity and fate in unclear ways. Single cell-sequencing technologies are powerful tools to explore cells, however analysis is typically gene-centric and TE activity has not been addressed. Here, we developed a single-cell TE processing pipeline, scTE, and report the activity of TEs in single cells in a range of biological contexts. Specific TE types were expressed in subpopulations of embryonic stem cells and were dynamically regulated during pluripotency reprogramming, differentiation, and embryogenesis. Unexpectedly, TEs were expressed in somatic cells, including human disease-specific TEs that are undetectable in bulk analyses. Finally, we applied scTE to single cell ATAC-seq data, and demonstrate that scTE can discriminate cell type using chromatin accessibly of TEs alone. Overall, our results reveal the dynamic patterns of TEs in single cells and their contributions to cell fate and heterogeneity.

bioinformatics↗

bioRxiv2020

Population Pharmacokinetics of an Anti-PD-1 Antibody Camrelizumab in Patients with Multiple tumor types and model informed dosing strategy

Objective Camrelizumab, a programmed cell death 1 (PD-1) inhibitor, has been approved for the treatment of relapsed or refractory classical Hodgkin lymphoma. The aim of this study was to perform a population pharmacokinetics (PK) analysis of camrelizumab to quantify the impact of patient characteristics on PK and to investigate the appropriateness of flat dose in the dosing regimen.Methods A total of 3298 camrelizumab concentrations from 133 patients from four studies were analyzed using nonlinear mixed effects modeling. Covariate model building was conducted using stepwise forward addition and backward elimination. Monte Carlo simulation was conducted to compare exposures of 200 mg and 3 mg/kg every 2-week regimens.Results The PK of camrelizumab were adequately described by a two-compartment model with parallel linear and nonlinear clearances. Baseline albumin had significant effects on linear clearance, and weight had effects on inter-compartmental clearance. Moreover, 200 mg and 3 mg/kg regimens provide similar exposure distributions with no advantage to either dosing approach.Conclusion Population PK analysis provided an integrated evaluation of the impact of albumin and weight on the PK of camrelizumab. It also provided evidence that neither the flat-dose nor the weight-based dose regimen was advantageous over the other for most patients with tumors.Competing Interest StatementThis study was sponsored by Jiangsu Hengrui Medicine Co. Ltd. Guang-li Ma, Da Xu and Yu-ya Wang are employees of Jiangsu Hengrui Medicine Co. Ltd.AbbreviationsAICAkaike’s information criterion;AUCarea under the concentration-time curve;AUCsssteady-state area under the concentration-time curve;BSVBetween-subject variability;Ccamrelizumab concentration;C1concentration of central compartment;Caverage,sssteady-state average concentration;CLclearance;CLlinearclearance of linear elimination;Cmax,sssteady-state peak concentration;CLnonlinearclearance of nonlinear elimination;Cmin,sssteady-state trough concentration;CWRESconditional weighted residuals;DVobserved concentration;IPREDindividual predicted concentrations;k0infusion rate;k23elimination rate from central compartment to peripheral compartment;k32elimination rate from peripheral compartment to central compartment;klinearlinear elimination rate;knonlinearnonlinear elimination rate;KmMichaelis–Menten constant;mAbmonoclonal antibody;NONMEMnonlinear mixed effect modeling;PD-1programmed cell death 1 receptor;PKpharmacokinetics;PREDpopulation predicted concentration;Qinter-compartmental clearance;Q2Wevery 2 weeks;Vmmaximum elimination rate;VPCvisual predictive check;V1distribution volume of central compartment;V2distribution volume of peripheral compartment;WBCwhite blood cell.View Full Text

cancer biology↗

bioRxiv2020

Efficient detection of pre-proinsulin by double antibody sandwich ELISA

Abstract ObjectiveTo generate monoclonal antibodies against pre-proinsulin (PPI), and establish sandwich ELISA method to provide a basis for PPI detection in recombinant human insulin production. MethodsThe Balb /c mice were immunized with PPI, and the hybridomas secreting anti-PPI monoclonal antibodies were obtained by conventional cell fusion technique and ELISA screening.The antibody was purified using a Protein G gel column and identified for purity by SDS-PAGE. Pairing effect was found by the sandwich ELISA, and the specificity of the paired antibody was determined. A paired antibody with better specificity was selected to establish sandwich ELISA, and construct a quantitative curve, the accuracy and sensitivity of the method were evaluated. ResultsSix anti-PPI monoclonal antibodies were obtained, named P1, P2, P3, P4, P5 and P6, of which P5 had the highest titer. The sandwich ELISA method was established with P5 for plating and P2 for detection antibodie. The linear range of the quantitative curve of PPI by sandwich ELISA was 0. 645-82.5 pg/mL, the recovery was 89%-95%, and the limit of detection was 3.06 pg/mL. ConclusionSix monoclonal antibodies against PPI were generated and the sandwich ELISA method was established to detect PPI in process control and product release control for recombinant human insulin production.

bioengineering↗

bioRxiv2019

An upstream open reading frame represses translation of chicken PPARγ transcript variant 1

Peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}) is a master regulator of adipogenesis. The PPAR{gamma} gene produces various transcripts with different 5'-untranslated regions (5' UTRs) because of alternative promoter usage and splicing. The 5' UTR plays important roles in posttranscriptional gene regulation. However, to date, the regulatory role and underlying mechanism of 5' UTRs in the posttranscriptional regulation of PPAR{gamma} expression remain largely unclear. In this study, we investigated the effects of 5' UTRs on posttranscriptional regulation using reporter assays. Our results showed that the five PPAR{gamma} 5' UTRs exerted different effects on reporter gene activity. Bioinformatics analysis showed that chicken PPAR{gamma} transcript 1 (PPAR{gamma}1) possessed an upstream open reading frame (uORF) in its 5' UTR. Mutation analysis showed that a mutation in the uORF led to increased Renilla luciferase activity and PPAR{gamma} protein expression, but decreased Renilla luciferase and PPAR{gamma}1 mRNA expression. mRNA stability analysis using real-time RT-PCR showed that the uORF mutation did not interfere with mRNA stability, but promoter activity analysis of the cloned 5' UTR showed that the uORF mutation reduced promoter activity. Furthermore, in vitro transcription/translation assays demonstrated that the uORF mutation markedly increased the translation of PPAR{gamma}1 mRNA. Collectively, our results indicate that the uORF represses the translation of chicken PPAR{gamma}1 mRNA.

molecular biology↗

bioRxiv2019

A cortico-collicular pathway for motor planning in a memory-dependent perceptual decision task

Survival in a dynamic environment requires animals to plan future actions based on past sensory evidence. However, the neural circuit mechanism underlying this crucial brain function, referred to as motor planning, remains unclear. Here, we employ projection-specific imaging and perturbation methods to investigate the direct pathway linking two key nodes in the motor planning network, the secondary motor cortex (M2) and the midbrain superior colliculus (SC), in mice performing a memory-dependent perceptual decision task. We find dynamic coding of choice information in SC-projecting M2 neurons during motor planning and execution, and disruption of this information by inhibiting M2 terminals in SC selectively impaired decision maintenance. Furthermore, cell-type-specific optogenetic circuit mapping shows that M2 terminals modulate both excitatory and inhibitory SC neurons with balanced synaptic strength. Together, our results reveal the dynamic recruitment of the premotor-collicular pathway as a circuit mechanism for motor planning.

neuroscience↗

bioRxiv2019

Parasites Acquired Beta Satellite DNAs from Hominid Hosts via Horizontal Gene Transfer

Beta satellite DNA (satDNA) sequences are repeated DNA elements located in primate centromeres and telomeres, and might play roles in genome stability and chromosome segregation. Beta satDNAs mainly exist in great apes. Previous studies suggested that beta satDNAs may originate in old world monkeys. In this study, we searched both GenBank and SRA database, and identified beta satDNA sequences from the genomic sequences of 22 species. The beta satDNA sequences found in Prosimian, Dermoptera and Scandentia indicated that the origin of beta satDNAs might be as early as 80 MYA. Strikingly, beta satDNA sequences were also found in a number of some species evolutionarily far from primates, including several endoparasites of human and other great apes, which could be the results of multiple horizontal gene transfer (HGT) events. The similar phylogenic profiles between beta satDNAs in the parasite genomes and the human genome indicates that the parasite beta satDNAs have undergone similar concerted evolution and play similar roles as the beta satDNAs in primates.\n\nHighlightsO_LIThe ever largest scale analysis on beta satDNAs.\nC_LIO_LIThe origin of beta satDNAs was traced back to [~]80 MYA.\nC_LIO_LIMass existence of beta satDNAs in non-primate species was contributed by multiple HGT events.\nC_LI

evolutionary biology↗

bioRxiv2018

Optical imaging of single protein size, charge, mobility, binding and conformational change

Protein analysis has relied on electrophoresis, mass spectroscopy and immunoassay, which separate, detect and identify proteins based on the size, charge, mobility and binding to antibodies. However, measuring these quantities at the single molecule level has not been possible. We tether a protein to a surface with a flexible polymer, drive the protein into mechanical oscillation with an alternating electric field, and image the protein oscillation with a near field imaging method, from which we determine the size, charge, mobility of the protein. We also measure binding of antibodies to single proteins and ligand binding-induced conformational changes in single proteins. This work provides new capabilities for protein analysis and disease biomarker detection at the single molecule level.

biophysics↗

bioRxiv2018

Time-resolved digital immunoassay for rapid and sensitive quantitation of procalcitonin with plasmonic imaging

Timely diagnosis of acute diseases improves treatment outcomes and saves lives, but it requires fast and precision quantification of biomarkers. Here we report a time-resolved digital immunoassay based on plasmonic imaging of binding of single nanoparticles to biomarkers captured on a sensor surface. The real-time and high contrast of plasmonic imaging lead to fast and precise counting of the individual biomarkers over a wide dynamic range. We demonstrated the detection principle, evaluated the performance of the method using procalcitonin (PCT) as an example, and achieved a limit of detection of ~ 3 pg/mL, dynamic range of 4-12500 pg/mL, for a total detection time of ~ 25 mins.

bioengineering↗

bioRxiv2018

Development and Application of a High-Content Virion Display Human GPCR Array

G protein-coupled receptors (GPCRs) comprise the largest membrane protein family in humans and can respond to a wide variety of ligands and stimuli. Like other multi-pass membrane proteins, the biochemical properties of GPCRs are notoriously difficult to study because they must be embedded in lipid bilayers to maintain their native conformation and function. To enable an unbiased, high-throughput platform to profile biochemical activities of GPCRs in native conformation, we individually displayed 315 human non-odorant GPCRs (>85% coverage) in the envelope of human herpes simplex virus-1 and immobilized on glass to form a high-content Virion Display (VirD) array. Using this array, we found that 50% of the tested commercial anti-GPCR antibodies (mAbs) is ultra-specific, and that the vast majority of those VirD-GPCRs, which failed to be recognized by the commercial mAbs, could bind to their canonical ligands, indicating that they were folded correctly. Next, we used the VirD-GPCR arrays to examine binding specificity of two known peptide ligands and recovered expected interactions, as well as new off-target interactions, three of which were confirmed with real-time kinetics measurements. Finally, we explored the possibility of discovering novel pathogen targets by probing VirD-GPCR arrays with live group B Streptococcus (GBS), a common Gram-positive bacterium causing neonatal meningitis. Using cell invasion assays and a mouse model of hematogenous meningitis, we showed that inhibition of one of the five newly identified GPCRs, CysLTR1, greatly reduced GBS penetration in brain-derived endothelial cells and in mouse brains. Therefore, our work demonstrated that the VirD-GPCR array holds great potential for high-throughput, unbiased screening for small molecule drugs, affinity reagents, and deorphanization.

pharmacology and toxicology↗

bioRxiv2018

The homoeologous genes for the Rec8-like meiotic cohesin in wheat: structure, function, and evolutionary implication

The Rec8-like cohesin is a cohesion protein essential for orderly chromosome segregation in meiosis. Here, we cloned two Rec8-like homoeologous genes (homoeoalleles) from tetraploid wheat (TtRec8-A1 and TtRec8-B1) and one from hexaploid wheat (TaRec8-D1), and performed expression and functional analyses of the homoeoalleles. Also, we identified other two Rec8 homoeoalleles in hexaploid wheat (TaRec8-A1 and TaRec8-B1) and the one in Aegilops tauschii (AetRec8-D1) by comparative analysis. The coding DNA sequences (CDS) of these six Rec8 homoeoalleles are all 1,827 bp in length, encoding 608 amino acids. They differed from each other primarily in introns although single nucleotide polymorphisms were detected in CDS. Substantial difference was observed between the homoeoalleles from the subgenome B (TtRec8-B1 and TaRec8-B1) and those from the subgenomes A and D (TtRec8-A1, TaRec8-A1, and TaRec8-D1). TtRec8-A1 expressed dominantly over TtRec8-B1, but comparably to TaRec8-D1. Therefore, the Rec8 homoeoalleles from the subgenomes A and D may be functionally more active than the one from the subgenome B in wheat. The structural variation and differential expression of the Rec8 homoeoalleles indicate a unique cross-genome coordination of the homoeologous genes in the polyploid, and imply the distinction of the wheat subgenome B from other subgenomes in the origin and evolution.\n\nHIGHLIGHTThis work revealed the structural and expression patterns of the Rec8-like homoeologous genes in polyploid wheat, implying a unique origin and evolutionary route of the wheat B subgenome.

genetics↗

bioRxiv2017

Clustering enzymes using E.coli inner cell membrane as scaffold in metabolic pathway

Clustering enzymes in the same metabolism pathway is a natural strategy to enhance the productivity. Several systems have been designed to artificially cluster desired enzymes in the cell, such as synthetic protein scaffold and nucleic acid scaffold. However, these scaffolds require complicated construction process and have limited slots for target enzymes. Following this direction, we designed a scaffold system based on natural cell membrane. Target enzymes (FabZ, FabG, FabI and TesA in fatty acid synthesis II pathway) are anchored on the E.coli inner membrane, showing the enhanced metabolism flux without the requirement of the further artificial interactions to force the clustering. Furthermore, anchoring the enzymes on the membrane enhances the products exportation, which further increases the productivity. Together, the proposed system has potential applications in producing valuable biomaterials.

synthetic biology↗

bioRxiv2017

Genome-wide homology analysis reveals new insights into the origin of the wheat B genome

Wheat is a typical allopolyploid with three homoeologous subgenomes (A, B, and D). The ancestors of the subgenomes A and D had been identified, but not for the subgenome B. The goatgrass Aegilops speltoides (genome SS) has been controversially considered a candidate for the ancestor of the wheat B genome. However, the relationship of the Ae. speltoides S genome with the wheat B genome remains largely obscure, which has puzzled the wheat research community for nearly a century. In the present study, the genome-wide homology analysis identified perceptible homology between wheat chromosome 1B and Ae. speltoides chromosome 1S, but not between other chromosomes in the B and S genomes. An Ae. speltoides-originated segment spanning a genomic region of approximately 10.46 Mb was identified on the long arm of wheat chromosome 1B (1BL). The Ae. speltoides-originated segment on 1BL was found to co-evolve with the rest of the B genome in wheat species. Thereby, we conclude that Ae. speltoides had been involved in the origin of the wheat B genome, but should not be considered an exclusive ancestor of this genome. The wheat B genome might have a polyphyletic origin with multiple ancestors involved, including Ae. speltoides. These novel findings provide significant insights into the origin and evolution of the wheat B genome, and will facilitate polyploid genome studies in wheat and other plants as well.

genetics↗