Biology subjects

Zhu, B.

Publications and source records attributed to Zhu, B..

At least 163 records · Page 9Linked to original sources

bioRxiv2021

Altered effective connectivity from the posterior insula to the amygdala mediates the relationship between psychopathic traits and concern with the Harm foundation

Psychopathic traits have been demonstrated to be associated with different types of morality; however, the neuropsychological mechanism underlying the relationship between psychopathic traits and morality remains unclear. Our study examined the effective connectivity (EC) of psychopathic traits-related brain regions and its association to concern with different moral foundations by combining behavioral measures with resting-state fMRI. We administered the Levenson Self-Report Psychopathy Scale (LSRP) and Moral Foundation Questionnaire (MFQ) to 78 college students after resting-state fMRI scanning. Our results showed that total and primary psychopathic traits score predicted concern with the Harm foundation. The EC from the posterior insula to the amygdala was negatively correlated with psychopathic traits and positively with concern with the Harm foundation. Altered posterior insula-amygdala EC partially mediated the relationship between psychopathic traits and concern with the Harm foundation. Our findings indicated that individuals with elevated psychopathic traits may have atypical processes in recognizing and integrating bodily state information into emotional responses, leading to less concern for harm-related morality. The study deepened our understanding of the neuropsychological mechanism underlying the relationship between psychopathic traits and morality and may have implications for the prevention of higher psychopathic traits individuals from committing serious transgressions.

neuroscience↗

bioRxiv2021

Reporting amyloid beta levels via bioluminescence imaging with amyloid reservoirs inAlzheimer's disease models

Bioluminescence imaging has changed daily practice in preclinical research of cancers and other diseases in the last decades; however, it has been rarely applied in preclinical research of Alzheimers disease (AD). In this report, we demonstrated that bioluminescence imaging could be used to report the levels of amyloid beta (A{beta}) species in vivo. We hypothesized that AkaLumine, a newly discovered substrate for luciferase, could bind to A{beta} aggregates and plaques. We further speculated that the A{beta} species have the reservoir capacity to sequester and release AkaLumine to control the bioluminescence intensity, which could be used to report the levels of A{beta}s. Our hypotheses have been validated via in vitro solution tests, mimic studies with brain tissues and mice, two-photon imaging with AD mice, and in vivo bioluminescence imaging using transgenic AD mice that were virally transduced with aka Luciferase (AkaLuc), a new luciferase that generates bioluminescence in the near infrared window. As expected, compared to the control group, we observed that the A{beta} group showed lower bioluminescence intensity due to AkaLumine sequestering at early time points, while higher intensity due to AkaLumine releasing at later time points. Lastly, we demonstrated that this method could be used to monitor AD progression and therapeutic effectiveness of avagacestat, a well-studied gamma-secretase inhibitor. Importantly, a good correlation (R2 = 0.81) was established between in vivo bioluminescence signals and A{beta} burdens of the tested AD mice. We believe that our approach can be easily implemented into daily imaging experiments and has tremendous potential to change daily practice of preclinical AD research.

neuroscience↗

bioRxiv2021

Virus-Dependent Immune Conditioning of Tissue Microenvironments

A thorough understanding of complex spatial host-disease interactions in situ is necessary in order to develop effective preventative measures and therapeutic strategies. Here, we developed Protein And Nucleic acid IN situ Imaging (PANINI) and coupled it with Multiplexed Ion Beam Imaging (MIBI) to sensitively and simultaneously quantify DNA, RNA, and protein levels within the microenvironments of tissue compartments. The PANINI-MIBI approach was used to measure over 30 parameters simultaneously across large sections of archival lymphoid tissues from non-human primates that were healthy or infected with simian immunodeficiency virus (SIV), a model that accurately recapitulates human immunodeficiency virus infection (HIV). This enabled multiplexed dissection of cellular phenotypes, functional markers, viral DNA integration events, and viral RNA transcripts as resulting from viral infection. The results demonstrated immune coordination from an unexpected upregulation of IL10 in B cells in response to SIV infection that correlated with macrophage M2 polarization, thus conditioning a potential immunosuppressive environment that allows for viral production. This multiplexed imaging strategy also allowed characterization of the coordinated microenvironment around latently or actively infected cells to provide mechanistic insights into the process of viral latency. The spatial multi-modal framework presented here is applicable to deciphering tissue responses in other infectious diseases and tumor biology.

immunology↗

bioRxiv2021

Whole Genome and Exome Sequencing Reference Datasets from A Multi-center and Cross-platform Benchmark Study

With the rapid advancement of sequencing technologies in the past decade, next generation sequencing (NGS) analysis has been widely applied in cancer genomics research. More recently, NGS has been adopted in clinical oncology to advance personalized medicine. Clinical applications of precision oncology require accurate tests that can distinguish tumor-specific mutations from errors or artifacts introduced during NGS processes or data analysis. Therefore, there is an urgent need to develop best practices in cancer mutation detection using NGS and the need for standard reference data sets for systematically benchmarking sequencing platforms, library protocols, bioinformatics pipelines and for measuring accuracy and reproducibility across platforms and methods. Within the SEQC2 consortium context, we established paired tumor-normal reference samples, a human triple-negative breast cancer cell line and a matched normal cell line derived from B lymphocytes. We generated whole-genome (WGS) and whole-exome sequencing (WES) data using 16 NGS library preparation protocols, seven sequencing platforms at six different centers. We systematically interrogated somatic mutations in the paired reference samples to identify factors affecting detection reproducibility and accuracy in cancer genomes. These large cross-platform/site WGS and WES datasets using well-characterized reference samples will represent a powerful resource for benchmarking NGS technologies, bioinformatics pipelines, and for the cancer genomics studies.

bioinformatics↗

bioRxiv2021

Prolonged epigenetic and synaptic plasticity alterations following single exposure to a psychedelic in mice

Clinical evidence suggests a potential therapeutic effect of classic psychedelics for the treatment of depression. The most outstanding and distinct characteristic is the rapid and sustained antidepressant action with one single exposure to the drug. However, the biological substrates and key mediators of psychedelics enduring action remain unknown. Here, we show that a single administration of the psychedelic DOI produced fast-acting effects on frontal cortex dendritic spine structure and acceleration of fear extinction via the 5-HT2A receptor. Additionally, a single dose of DOI led to changes in chromatin organization particularly at enhancer regions of genes involved in synaptic assembly that stretched for days after the psychedelic exposure. DOI-induced alterations in neuronal epigenome overlapped with genetic loci associated with schizophrenia, depression and attention deficit hyperactivity disorder. Together, these data support the notion that epigenetic-driven changes in synaptic plasticity operate as the mechanistic substrate of psychedelics long-lasting antidepressant action but also warn on the limitations in individuals with underlying risk for psychosis.

neuroscience↗

bioRxiv2021

Production of SARS-CoV-2 virus-like particles in insect cells

Coronavirus disease (COVID-19) causes a serious threat to human health. To production of SARS-COV-2 virus-like particles (VLPs) in insect cells for vaccine development and scientific research. The E, M and S genes were cloned into multiple cloning sites of the new triple expression plasmid with one p10 promoter, two pPH promoters and three multiple cloning sites. The plasmid was transformed into DH10 BacTM Escherichia coli competent cells to obtain recombinant bacmid. Then the recombinant bacmid was transfected in ExpiSf9 insect cells to generate recombinant baculovirus. After ExpiSf9 infected with the recombinant baculovirus, the E, M, and S protein co-expressed in insect cells. Finally, SARS-CoV-2 VLPs were self-assembled in insect cells after infection. The morphology and the size of SARS-CoV-2 VLPs are similar to the native virions.

bioengineering↗

bioRxiv2021

A single mutation attenuates both the transcription termination and RNA-dependent RNA polymerase activity of T7 RNA polymerase

Transcription termination is one of the least understood processes of gene expression. As the prototype model for transcription studies, the single-subunit T7 RNA polymerase (RNAP) was known to response to two types of termination signals, while the mechanism underlying such termination especially the specific elements of the polymerase involved in is still unclear, due to the lack of a termination complex structure. Here we applied phage-assisted continuous evolution to obtain variants of T7 RNAP that can bypass the typical class I T7 terminator with stem-loop structure. Through in vivo selection and in vitro characterization, we discovered a single mutation S43Y that significantly decreased the termination efficiency of T7 RNAP at all transcription terminators tested. Coincidently, the S43Y mutation almost eliminates the RNA-dependent RNA polymerase (RdRp) of T7 RNAP without affecting the major DNA-dependent RNA polymerase (DdRp) activity of the enzyme, indicating the relationship between transcription termination and RdRp activity, and suggesting a model in which the stem-loop terminator induces the RdRp activity which competes with the ongoing DdRp activity to cause transcription termination. The T7 RNAP S43Y mutant as an enzymatic reagent for in vitro transcription reduces the undesired termination in run-off RNA synthesis and produces RNA with higher terminal homogeneity.

molecular biology↗

bioRxiv2020

A nucleotide-sensing endonuclease from the Gabija bacterial defense system

The arms race between bacteria and phages has led to the development of exquisite bacterial defense systems including a number of uncharacterized systems distinct from the well-known Restriction-Modification and CRISPR/Cas systems. Here, we report functional analyses of the GajA protein from the newly predicted Gabija system. The GajA protein is revealed as an endonuclease unique in that: 1. It may function as a restriction enzyme or a site-specific nicking enzyme, depending on the arrangement of the recognition sequences; 2. Its activity is strictly regulated by nucleotides concentration. NTP and dNTP at physiological concentrations can fully inhibited the robust DNA cleavage activity of GajA. Interestingly, the nucleotide inhibition is mediated by an ATPase-like domain, which usually hydrolyzes ATP to stimulate the DNA cleavage when associated with other nucleases. These features suggested the mechanism of the Gabija defense in which an endonuclease activity was suppressed at normal condition, while activated by the depletion of NTP and dNTP upon the replication and transcription by invaded phages. This work highlights a concise strategy to utilize a single protein for phage resistance via nucleotide regulatory.

biochemistry↗

bioRxiv2020

Body site-specific and disease-specific virulome in the human microbiome

Human body habitats are home to a diverse array of microbes, and within these microbial ecosystems, there are exchanges of genetic material, including virulence factors (VFs). Little is known about the diversity and abundance of VFs in different body sites and different types of diseases. We developed a virulome analysis pipeline using the species-specific sequence identity inferred from intraspecies ANI values to precisely assign reads to virulence factors. We characterized the human virulome from four body habitats, including the gut, oral cavity, skin, and vagina. Specifically, the diversity and abundance of VFs in the oral cavity were significantly higher than those in other body sites, including stool. We highlight the importance of sex-specific analysis when studying the human virulome. We analyzed data from more than 4,000 samples across healthy and diseased subjects and 13 types of diseases from different metagenomic sequencing studies to characterize the disease-specific virulome. Atherosclerotic cardiovascular disease (ACVD) has a more diverse virulome than other diseases tested. Notably, many VFs, including genes for secretion systems and toxins, are more abundant in diseased subjects than in healthy controls. We present, to our knowledge, the most comprehensive healthy and diseased virulome dataset yet created.

genomics↗

bioRxiv2020

New insight into bacterial iron competition: A-to-I editing in ferric enterobactin uptake

Adenosine-to-inosine (A-to-I) RNA editing, which is catalyzed by the adenosine deaminase RNA-specific family of enzymes, is a frequent post-transcriptional modification in metazoans. Research on A-to-I editing in bacteria is limited, and the importance is underestimated. In this study, we show that bacteria may use A-to-I editing as an alternative strategy to promote uptake of metabolic iron. The T408A editing event of xfeA in Xanthomonas oryzae pv. oryzicola (Xoc) senses extracytoplasmic iron and changes the hydrogen bonding network of ligand channel domains. The frequency of A-to-I RNA editing during iron-deficient conditions increased by 76.87%, which facilitated the passage of iron through the XfeA outer membrane channel. When bacteria were subjected to high iron concentrations, the percentage of A-to-I editing in xfeA decreased, which reduced iron transport via XfeA. Furthermore, A-to-I RNA editing increased expression of multiple genes in the chemotaxis pathway, including methyl-accepting chemotaxis proteins (MCPs) that sense concentrations of exogenous ferric enterobactin (Fe-Ent) at the cytoplasmic membrane. A-to-I RNA editing helps Xoc move towards an iron-rich environment and supports our contention that editing in xfeA facilitates entry of a ferric siderophore. Overall, our results reveal a new signaling mechanism that bacteria use to facilitate iron uptake and improve their competitiveness.

microbiology↗

bioRxiv2020

β-Amyloid peptides tailor switching behaviors of Donor-Acceptor Stenhouse Adducts

Molecular switching plays a critical role in biological and displaying systems. Here we demonstrate the first use of peptides to operate molecular switches of donor-acceptor Stenhouse adducts (DASAs), a series of negative photochromes that are highly promising for applications ranging from smart material to biological systems. Fluorescence imaging proved A{beta}40 species could make SHA-2 more stable in the linear configuration than without peptide and decrease the rate of molecular switching. According to molecular dynamics simulation, SHA-2 bound to protein resulted in substantial changes in the tertiary structure of A{beta}40 monomer with the region of Glu22-Ala30 partially unfolded and being more exposed to water. This structural change is likely to impede the aggregation of A{beta}40, as evidenced by fluorescence and ProteoStat(R) aggresome detection experiments. SHA-2 is able to inhibit the aggregation of A{beta}40 by producing the off-pathway structures. These results open ample opportunities for optically addressable potential widely apply DASAs in the biological system based on this peptides-tailor process.

biochemistry↗

bioRxiv2020

In vitro transcription using psychrophilic phage VSW-3 RNA polymerase

RNA research and applications were underpinned by in vitro transcription (IVT), while the RNA impurity resulted from the enzymatic reagents severely impede downstream applications. To improve the stability and purity of synthesized RNA we had characterized a novel single-subunit RNA polymerase (RNAP) encoded by a psychrophilic phage VSW-3 from plateau lake to produce RNA at low temperature. The VSW-3 RNAP is capable of carrying out in vitro RNA synthesis at low temperature (4-25{degrees}C) to reduce RNA degradation and alleviate the need of costly RNase inhibitor. Compared to routinely used T7 RNAP, VSW-3 RNAP provides comparable yield of transcripts, but is insensitive to class II transcription terminators and synthesizes RNA without redundant 3 -cis extension. More importantly, through dot-blot detection with the J2 monoclonal antibody, we found that the RNA products synthesized by VSW-3 RNAP contain much lower amount of or virtually no double-stranded RNA (dsRNA) by-products, which are significant in most T7 RNAP products and may cause severe cellular immune response. Combining these advantages, the VSW-3 RNAP is an advantageous enzyme for IVT, especially to produce RNA for in vivo use.

molecular biology↗

bioRxiv2020

Manganese depletion leads to multisystem changes in the transcriptome of the opportunistic pathogen Streptococcus sanguinis

1Streptococcus sanguinis is a primary tooth colonizer and is typically considered beneficial due to its antagonistic relationship with the cariogenic pathogen Streptococcus mutans. However, S. sanguinis can also act as an opportunistic pathogen should it enter the bloodstream and colonize a damaged heart valve, leading to infective endocarditis. Studies have implicated manganese acquisition as an important virulence determinant in streptococcal endocarditis. A knockout mutant lacking the primary manganese import system in S. sanguinis, SsaACB, is severely attenuated for virulence in an in vivo rabbit model. Manganese is a known cofactor for several important enzymes in S. sanguinis, including superoxide dismutase, SodA, and the aerobic ribonucleotide reductase, NrdEF. To determine the effect of manganese depletion on S. sanguinis, we performed transcriptomic analysis on a {Delta}ssaACB mutant grown in aerobic fermentor conditions after the addition of the metal chelator EDTA. Despite the broad specificity of EDTA, analysis of cellular metal content revealed a decrease in manganese, but not in other metals, that coincided with a drop in growth rate. Subsequent supplementation with manganese, but not iron, zinc, or magnesium, restored growth in the fermentor post-EDTA. Reduced activity of Mn-dependent SodA and NrdEF likely contributed to the decreased growth rate post-EDTA, but did not appear entirely responsible. With the exception of the Dps-like peroxide resistance gene, dpr, manganese depletion did not induce stress response systems. By comparing the transcriptome of {Delta}ssaACB cells pre- and post-EDTA, we determined that manganese deprivation led to altered expression of diverse systems, including ethanolamine utilization, CRISPR/Cas, and a type IV pilus. Manganese depletion also led to an apparent induction of carbon catabolite repression in a glucose-independent manner. The combined results suggest that manganese limitation produces effects in S. sanguinis that are diverse and complex, with no single protein or system appearing entirely responsible for the observed growth rate decrease. This study provides further evidence for the importance of this trace element in streptococcal biology. Future studies will focus on determining mechanisms for regulation, as the multitude of changes observed in this study indicate that multiple regulators may respond to manganese levels.

microbiology↗

bioRxiv2020

Increasing cell-free gene expression yields from linear templates in Escherichia coli and Vibrio natriegens extracts by using DNA-binding proteins

In crude extract-based cell-free protein synthesis (CFPS), DNA templates are transcribed and translated into functional proteins. Although linear expression templates (LETs) are less laborious and expensive to generate, plasmid templates are often desired over PCR-generated LETs due to increased stability and protection against exonucleases present in the extract of the reaction. Here we demonstrate that addition of a dsDNA-binding protein to the CFPS reaction, termed single-chain Cro protein (scCro), achieves terminal protection of LETs. This CroP-LET (scCro-based Protection of LET) method effectively increases sfGFP expression levels from LETs in Escherichia coli CFPS reactions by 6-fold. Our yields are comparable to other strategies that provide chemical and enzymatic DNA stabilization in E. coli CFPS. Notably, we also report that the CroP-LET method successfully enhanced yields in CFPS platforms derived from non-model organisms. Our results show that CroP-LET increased sfGFP yields by 18-fold in the Vibrio natriegens CFPS platform. With the fast-expanding applications of CFPS platforms, this method provides a practical and generalizable solution to protect linear expression DNA templates.

synthetic biology↗

bioRxiv2020

The Forensic Landscape and the Population Genetic Analyses of Hainan Li Based on Massively Parallel Sequencing DNA Profiling

Due to the formation of the Qiongzhou Strait by climate change and marine transition, Hainan island isolated from the mainland southern China during the Last Glacial Maximum. Hainan island, located at the southernmost part of China and separated from the Leizhou Peninsula by the Qiongzhou Strait, laid on one of the modern human northward migration routes from Southeast Asia to East Asia. The Hlai-language speaking Li minority, the second largest population after Han Chinese in Hainan island, is the direct descendants of the initial migrants in Hainan island and has unique ethnic properties and derived characteristics, however, the forensic associated studies on Hainan Li population are still insufficient. Hence, 136 Hainan Li individuals were genotyped in this study using the MPS-based ForenSeq DNA Signature Prep Kit (DNA Primer Set A) to characterize the forensic genetic polymorphism landscape, and DNA profiles were obtained from 152 different molecular genetic markers (27 autosomal STRs, 24 Y-STRs, 7 X-STRs, and 94 iiSNPs). A total of 419 distinct length variants and 586 repeat sequence sub-variants, with 31 novel alleles (at 17 loci), were identified across the 58 STR loci from the DNA profiles of Hainan Li population. We evaluated the forensic characteristics and efficiencies of DAPA, it demonstrated that the STRs and iiSNPs in DAPA were highly polymorphic in Hainan Li population and could be employed in forensic applications. In addition, we set up three Datasets, which included the genetic data of (I). iiSNPs (27 populations, 2640 individuals), (II). Y-STRs (42 populations, 8281 individuals), and (III). Y-haplogroups (123 populations, 4837 individuals) along with the population ancestries and language families, to perform population genetic analyses separately from different perspectives. In conclusion, the phylogenetic analyses indicated that Hainan Li, with a southern East Asia origin and Tai-Kadai language-speaking language, is an isolated population relatively. But the genetic pool of Hainan Li influenced by the limited gene flows from other Tai-Kadai populations and Hainan populations. Furthermore, the establishment of isolated population models will be beneficial to clarify the exquisite population structures and develop specific genetic markers for subpopulations in forensic genetic fields.

genetics↗

bioRxiv2020

Establishing Prokaryotic Expression System of Angiotensin- Converting Enzyme 2 (ACE2) gene in pigs

In this paper, ACE2 gene of pigs was cloned and the purified protein was obtained via the prokaryotic expression system. Polyclonal antibody of high titer and sensitivity was obtained using Wastar rats immunization method and is then used to determine of the expression of ACE2 using immunohistochemistry. The sequence of ACE2 in pigs covered 2418 nucleotides and coded 805 amino acid (aa) residues. Sequence homology analysis showed that the ACE2 sequence in pigs is highly conserved among species at the nucleotide and amino acid levels. Genetic evolution analysis revealed that ACE2 gene in pigs has the shortest genetic distance with that in goats while residing in a totally different branch from that in zebra fishes. Analysis of protein structure predicted that ACE2 protein is a transmembrane secreted protein with high hydrophilicity, containing a signal peptide sequences locating between 1aa to 17aa. The ACE2 fusion protein expressed (under the induction with 1.0 mmol/L IPTG for 10 h) was of approximately 100 kDa and mainly existed in inclusion body. Wastar rats immunization showed that the titer of the anti-ACE2 antiserum in rats was 1: 3200. Western blot showed that the antibody binds specifically. Immunohistochemistry showed that the ACE2 protein was expressed in all major tissues of pigs. It is the first time that polyclonal antibody of ACE2 in pigs was obtained and the expression of ACE2 was confirmed. These results will provide a basis for investigating on ACE2s biological activity in pigs.

biochemistry↗

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

Interneuron dysfunction in a new knock-in mouse model of SCN1A GEFS+

Advances in genome sequencing have identified over 1300 mutations in the SCN1A sodium channel gene that result in genetic epilepsies. However, how individual mutations within SCN1A produce seizures remains elusive for most mutations. Previous work from our lab has shown that the K1270T (KT) mutation, which is linked to GEFS+ (Genetic Epilepsy with Febrile Seizure plus) in humans, causes reduced firing of GABAergic neurons in a Drosophila knock-in model. To examine the effect of this mutation in mammals, we introduced the equivalent KT mutation into the mouse Scn1a (Scn1aKT) gene using CRISPR/Cas9. Mouse lines carrying this mutation were examined in two widely used genetic backgrounds, C57BL/6NJ and 129x1/SvJ. In both backgrounds, homozygous mutants had spontaneous seizures and died by postnatal day 23. There was no difference in the lifespan of mice heterozygous for the mutation in either background when compared to wild-type littermates up to 6 months. Heterozygous mutants had heat-induced seizures at ~42 deg. Celsius, a temperature that did not induce seizures in wild-type littermates. In acute hippocampal slices, current-clamp recordings revealed a significant depolarized shift in action potential threshold and reduced action potential amplitude in parvalbumin-expressing inhibitory interneurons in Scn1aKT/+ mice. There was no change in the firing properties of excitatory CA1 pyramidal neurons. Our results indicate that Scn1aKT/+ mice develop seizures, and impaired action potential firing of inhibitory interneurons in Scn1aKT/+ mice may produce hyperexcitability in the hippocampus.

neuroscience↗