Biology subjects

Wang, B.

Publications and source records attributed to Wang, B..

At least 685 records · Page 38Linked to original sources

bioRxiv2019

The identification of mecciRNAs and their roles in mitochondrial entry of proteins

Mammalian mitochondria have small genomes encoding very limited numbers of proteins. Over one thousand proteins and noncoding RNAs encoded by nuclear genome have to be imported from the cytosol into the mitochondria. Here we report the identification of hundreds of circular RNAs (mecciRNAs) encoded by mitochondrial genome. We provide both in vitro and in vivo evidence to show that mecciRNAs facilitate mitochondrial entry of nuclear-encoded proteins by serving as molecular chaperones in the folding of imported proteins. Known components of mitochondrial protein and RNA importation such as TOM40 and PNPASE interact with mecciRNAs and regulate protein entry. Expression of mecciRNAs is regulated, and these transcripts are critical for mitochondria in adapting to physiological conditions and diseases such as stresses and cancers by modulating mitochondrial protein importation. mecciRNAs and their associated physiological roles add categories and functions to eukaryotic circular RNAs, and shed novel lights on communication between mitochondria and nucleus.

molecular biology↗

bioRxiv2019

High-throughput proteomic analysis of FFPE tissue samples facilitates tumor stratification

Formalin-fixed, paraffin-embedded (FFPE), biobanked tissue samples offer an invaluable resource for clinical and biomarker research. Here we developed a pressure cycling technology (PCT)-SWATH mass spectrometry workflow to analyze FFPE tissue proteomes and applied it to the stratification of prostate cancer (PCa) and diffuse large B-cell lymphoma (DLBCL) samples. We show that the proteome patterns of FFPE PCa tissue samples and their analogous fresh frozen (FF) counterparts have a high degree of similarity and we confirmed multiple proteins consistently regulated in PCa tissues in an independent sample cohort. We further demonstrate temporal stability of proteome patterns from FFPE samples that were stored between one to 15 years in a biobank and show a high degree of the proteome pattern similarity between two types histological region of small FFPE samples, i.e. punched tissue biopsies and thin tissue sections of micrometer thickness, despite the existence of certain degree of biological variations. Applying the method to two independent DLBCL cohorts we identified myeloperoxidase (MPO), a peroxidase enzyme, as a novel prognostic marker. In summary, this study presents a robust proteomic method to analyze bulk and biopsy FFPE tissues and reports the first systematic comparison of proteome maps generated from FFPE and FF samples. Our data demonstrate the practicality and superiority of FFPE over FF samples for proteome in biomarker discovery. Promising biomarker candidates for PCa and DLBCL have been discovered.

biochemistry↗

bioRxiv2019

New mechanism-based inhibitors of aspartate transcarbamoylase for anticancer drug development

Aspartate transcarbamoylase (ATCase) is a key enzyme which regulates and catalyzes the second step of de novo pyrimidine synthesis in all organisms. E. coli ATCase is a prototypic enzyme regulated by both product feedback and substrate cooperativity, whereas human ATCase is a potential anticancer target. Through structural and biochemical analyses, we revealed that R167/130s loop region in ATCase serves as a gatekeeper for the active site, playing a new and unappreciated role in feedback regulation. Based on virtual compound screening simultaneously targeting the new regulatory region and active site of human ATCase, two compounds were identified to exhibit strong inhibition of ATCase activity, proliferation of multiple cancer cell lines, and growth of xenograft tumors. Our work has not only revealed a previously unknown regulatory region of ATCase that helps explain feedback regulation, but also successfully guided the identification of new ATCase inhibitors for anticancer drug development using a dual-targeting strategy.

cancer biology↗

bioRxiv2019

Variant Phasing and Haplotypic Expression from Single-molecule Long-read Sequencing in Maize

AbstractHaplotype phasing of genetic variants in maize is important for interpretation of the genome, population genetic analysis and functional genomic analysis of allelic activity. Accordingly, accurate methods for phasing the full-length isoforms are essential for functional genomics studies. We performed an isoform-level phasing study in maize, using two inbred lines and their reciprocal crosses, based on the single-molecule full-length cDNA sequencing. To phase and analyze the full-length transcripts between hybrids and parents, we developed a tool called IsoPhase. Using this tool, we validated the majority of SNPs called against matching short-read data and identified cases of allele-specific, gene-level and isoform-level expression. Our results revealed that maize parental lines and hybrid lines exhibit different splicing activities. After phasing 6,907 genes in two reciprocal hybrids using embryo, endosperm and root tissues, we annotated the SNPs and identified large-effect genes. In addition, based on single-molecule sequencing, we identified parent-of-origin isoforms in maize hybrids, distinct novel isoforms in maize parent and hybrid lines, and imprinted genes from different tissues. Finally, we characterized variation in cis- and trans-regulatory effects. Our study provides measures of haplotypic expression that could increase accuracy in studies of allelic expression.

bioinformatics↗

bioRxiv2019

Mechanically Resolved Imaging of Bacteria using Expansion Microscopy

Imaging dense and diverse microbial communities has broad applications in basic microbiology and medicine, but remains a grand challenge due to the fact that many species adopt similar morphologies. While prior studies have relied on techniques involving spectral labeling, we have developed an expansion microscopy method (ExM) in which cells are physically expanded prior imaging and their expansion patterns depend on the structural and mechanical properties of their cell walls, which vary across species and conditions. We use this phenomenon as a quantitative and sensitive phenotypic imaging contrast orthogonal to spectral separation in order to resolve bacterial cells of different species or in distinct physiological states. Focusing on host-microbe interactions that are difficult to quantify through fluorescence alone, we demonstrate the ability of ExM to distinguish species within a dense community through in vivo imaging of a model gut microbiota, and to sensitively detect cell-envelope damage caused by antibiotics or previously unrecognized cell-to-cell phenotypic heterogeneity among pathogenic bacteria as they infect macrophages.

biophysics↗

bioRxiv2019

Brown adipocyte NOSEMPE promotes nonmitochondrial thermogenesis and improves systemic metabolism through ATF4 activation

Mitochondrial transcription factor A (Tfam)-mediated mtDNA maintenance and transcription, as well as leucine-rich PPR motif-containing protein (Lrpprc)-mediated mtRNA maturation and translation are essential steps of mtDNA-encoded electron transport chain (ETC) protein expression. ETC is essential for mitochondrial thermogenesis, the process of oxygen-dependent heat production inside the mitochondria in brown adipocytes. Here we describe that Tfam or Lrpprc deficiency in brown adipocytes cause non-synchronized ETC mRNA and protein expression (NOSEMPE) and mitochondrial ETC imbalance, ultimately abolish mitochondrial thermogenesis. However, mice with NOSEMPE in brown adipocytes are cold resistant upon an acute 4{degrees}C cold challenge, because of augmented nonmitochondrial thermogenesis driven by the \"NOSEMPE[->]ATF4[->]proteome turnover\" pathway. Importantly, mice with either NOSEMPE or ATF4 overexpression in brown adipocytes are protected against high-fat-diet-induced metabolic abnormalities, indicating a positive association between nonmitochondrial thermogenesis in brown adipocytes and metabolic fitness. Thus, although brown adipocytes are defined by their unique ability to produce heat through mitochondrial respiration, our study demonstrates a novel cytosolic nonmitochondrial thermogenesis in brown adipocytes. Targeting this ATF4-dependent nonmitochondrial thermogenesis in brown adipocytes may represent a new therapeutic strategy for combating metabolic disorders.

physiology↗

bioRxiv2019

Anticipatory distractor suppression elicited by statistical regularities in visual search

Salient yet irrelevant objects often capture our attention and interfere with our daily tasks. Distraction by salient objects can be reduced by suppressing the location where they are likely to appear. The question we addressed here was whether suppression of frequent distractors is already implemented beforehand, in anticipation of the stimulus. Using electroencephalography (EEG), we recorded cortical activity of human participants searching for a target while ignoring a salient distractor. The distractor was presented more often at one location than at any other location. We found reduced capture for distractors at frequent locations, indicating that participants learned to avoid distraction. Critically, we found evidence for proactive suppression as already prior to display onset, there was enhanced power in parieto-occipital alpha oscillations contralateral to the frequent distractor location - a signal known to occur in anticipation of irrelevant information. Locked to display onset, event-related potentials analysis showed a distractor-suppression-related PD component for this location. Importantly, this PD was found regardless of whether distracting information was presented at the frequent location. In addition, there was an early PD component representing an early attentional index of the frequent distractor location. Our results are show anticipatory (proactive) suppression of frequent distractor locations in visual search already starting prior to display onset.

neuroscience↗

bioRxiv2019

PERK inhibition mitigates restenosis and thrombosis - a potential low-thrombogenic anti-restenotic paradigm

BackgroundDrug-eluting stents (DES) represent the main-stream management of restenosis following treatments of occlusive cardiovascular diseases. However, DES cannot eliminate instent restenosis yet exacerbate thrombogenic risks. To achieve dual inhibition of restenotic smooth muscle cell (SMC) de-differentiation/proliferation and thrombogenic endothelial cell (EC) dysfunction, a common target in both cell types, has been long-sought after. We evaluated the potential of protein kinase RNA-like endoplasmic reticulum kinase (PERK) as such a target for low-thrombogenic anti-restenotic intervention.\n\nMethods and ResultsWe used a rat angioplasty model of restenosis and a FeCl3-induced mouse model of thrombosis. Loss-or gain-of-function was achieved by PERK inhibition (GSK2606414, siRNA) or overexpression (adenovirus). Restenosis was robustly mitigated by GSK2606414 administered either via injected (i.v.) lesion-homing platelet membrane-coated nanoclusters or a perivascular hydrogel; it was enhanced by PERK transgene. Whereas PERK inhibition blocked, its overexpression exacerbated PDGF-induced human aortic SMC de-differentiation (reduced smooth muscle -actin or SMA) and proliferation (BrdU incorporation). Further, PERK activity promoted STAT3 activation but inhibited SRF transcriptional (luciferase) activity; its protein co-immunoprecipitated with STAT3 and also MRTF-A, the SRF activator for SMA transcription. Importantly, PERK inhibition also prevented TNF-induced impairment of human EC growth and upregulation of thrombogenic tissue factor, both in vitro and ex vivo. In vivo, oral gavage of GSK2606414 preserved ~50% of the normal blood flow 60 min after FeCl3-induced vascular injury.\n\nConclusionsPERK inhibition is dual beneficial in mitigating restenosis and thrombosis, thus implicating a potential design for anti-restenotic intervention to overcome the thrombogenicity of DES.

pathology↗

bioRxiv2019

Characterization and repurposing of the endogenous Type I-F CRISPR-Cas system of Zymomonas mobilis for genome engineering

Establishment of production platform organisms through prokaryotic engineering represents an efficient means to generate alternatives for yielding renewable biochemicals and biofuels from sustainable resources. Zymomonas mobilis, a natural facultative anaerobic ethanologen, possesses many attractive physiological attributes, making it an important industrial microorganism. To facilitate the broad applications of this strain for biorefinery, an efficient genome engineering toolkit for Z. mobilis was established in this study by repurposing the endogenous Type I-F CRISPR-Cas system upon its functional characterization, and further updated. This toolkit includes a series of genome engineering plasmids, each carrying an artificial self-targeting CRISPR and a donor DNA for the recovery of recombinants. Using the updated toolkit, genome engineering purposes were achieved with efficiencies of up to 100%, including knockout of cas3 gene, replacement of cas3 with the mCherry-encoding rfp gene, nucleotide substitutions in cas3, and deletion of two large genomic fragments up to 10 kb. This study established thus far the most efficient, straightforward and convenient genome engineering toolkit for Z. mobilis, and laid a foundation for further native CRISPRi studies in Z. mobilis, which extended the application scope of CRISPR-based technologies, and could also be applied to other industrial microorganisms with unexploited endogenous CRISPR-Cas systems.

bioengineering↗

bioRxiv2019

A non-canonical role of polo-like kinase-4 in adventitial fibroblast cell type transition

A divergent member of the polo-like kinase family, PLK4 is known for its canonical role in centriole duplication. Its non-canonical function and regulators are poorly defined. Here we investigated PLK4s activation and expression and regulations thereof in rat adventitial fibroblast cell-type transition induced by platelet-derived growth factor (PDGF-AA).\n\nExperiments using siRNA and selective inhibitor (centrinone-B) revealed a role for PLK4 not only in AA-induced proliferation/migration, but also in serum response factor (SRF) activation and smooth muscle -actin expression. PDGFR (receptor) inhibition abrogated AA-stimulated PLK4 activation (phosphorylation) and expression; P38 inhibition (siRNA, inhibitor) downstream of PDGFR also mitigated PLK4 activation. Furthermore, transcription of PLK4 (and PDGFR) was repressed by pan-inhibition of the bromodomain/extraterminal family of chromatin-bookmark readers (BRD2, BRD3, BRD4), an effect determined herein as mainly mediated by BRD4. In vivo, periadventitial administration of centrinone-B reduced collagen content and thickness of the adventitia in a rat model of carotid artery injury.\n\nIn summary, we have identified a non-canonical role for PLK4 in SRF activation and its regulations by BRD4/PDGFR-dominated pathways. Results in this study suggest PLK4 inhibition as a potential anti-fibrotic intervention.

cell biology↗

bioRxiv2019

Demographic histories and genome-wide patterns of divergence in incipient species of shorebirds

Understanding how incipient species are maintained with gene flow is a fundamental question in evolutionary biology. Whole genome sequencing of multiple individuals holds great potential to illustrate patterns of genomic differentiation as well as the associated evolutionary histories. Kentish (Charadrius alexandrinus) and the white-faced (C. dealbatus) plovers, which differ in their phenotype, ecology and behaviour, are two incipient species and parapatrically distributed in East Asia. Previous studies show evidence of genetic diversification with gene flow between the two plovers. Under this scenario, it is of great importance to explore the patterns of divergence at the genomic level and to determine whether specific regions are involved in reproductive isolation and local adaptation. Here we present the first population genomic analysis of the two incipient species based on the de novo Kentish plover reference genome and resequenced populations. We show that the two plover lineages are distinct in both nuclear and mitochondrial genomes. Using model-based coalescence analysis, we found that population sizes of Kentish plover increased whereas white-faced plovers declined during the Last Glaciation Period. Moreover, the two plovers diverged allopatrically, with gene flow occurring after secondary contact. This has resulted in low levels of genome-wide differentiation, although we found evidence of a few highly differentiated genomic regions in both the autosomes and the Z-chromosome. This study illustrates that incipient shorebird species with gene flow after secondary contact can exhibit discrete divergence at specific genomic regions and provides basis to further exploration on the genetic basis of relevant phenotypic traits.

evolutionary biology↗

bioRxiv2019

Exploring the influence of surface proteins on probiotic activity of Lactobacillus pentosus HC-2 in Litopenaeus vannamei midgut via Label-free quantitative proteomic analysis

Our previously work showed that Lactobacillus pentosus HC-2 as probiotic could improve the growth performance, immune response, gut bacteria diversity and disease resistance of Litopenaeus vannamei. However, the probiotic mechanism of was not fully characterized. In the present study, histology and proteomic analysis was performed to explore the influence of HC-2 surface protein on its probiotic effects to L. vannamei after feeding the intact surface proteins or the probiotic treated with lithium chloride (LiCl) to remove non-covalently bound surface proteins or no probiotic for four weeks. Histological observation found that feeding with normal HC-2 obviously improved the intestinal histology and enhanced the protective effect against pathogens damages, but fed with LiCl-treated HC-2 didnt improve the intestinal environment. A total over 2,764 Peptides and 1,118 uniproteins were identified from L. vannamei midgut, 211 proteins were significant differentially expressed normal HC-2 group compared with control, 510 proteins were significant differentially expressed in LiCl-tread HC-2 group compared with control, and 458 proteins were significant differentially expressed in LiCl-tread HC-2 group compared with normal HC-2 group. GO/KEGG enrichment analysis of the significantly different proteins demonstrated that fed with normal HC-2 mainly induced immune response, metabolic, cell adhesion and cell-cell signaling related proteins up-regulation, which were contributed to bacteria adhesion and colonization in midgut to improve shrimp immune system and growth, but these proteins were suppressed after feeding with deprived surface proteins bacteria. Taken together, these results indicating that the surface proteins were indispensable for HC-2 to execute probiotic effects in midgut of shrimp.

microbiology↗

bioRxiv2019

The BD2 domain of BRD4 is a determinant in EndoMT and vein graft neointima formation

BackgroundVein-graft bypass is commonly performed to overcome atherosclerosis but is limited by high failure rates, principally due to neointimal wall thickening. Recent studies reveal that endothelial-mesenchymal transition (EndoMT) is critical for vein-graft neointima formation. BETs are a family of Bromo/ExtraTerminal domains-containing epigenetic reader proteins (BRD2, BRD3, BRD4). They bind acetylated histones through their unique tandem bromodomains (BD1, BD2), facilitating transcriptional complex formation and cell-state transitions. The role for BETs, including individual BRDs and their unique BDs, is not well understood in EndoMT and neointimal formation.\n\nMethods and ResultsRepression of BRD4 expression abrogated TGF{beta}1-induced EndoMT, with greater effects than BRD2 or BRD3 knockdown. An inhibitor selective for BD2 in all BETs, but not that for BD1, blocked EndoMT. Moreover, expression of a dominant-negative BRD4-specific BD2 fully abolished EndoMT. Concordantly, BRD4 knockdown repressed TGF{beta}1-stimulated increase of ZEB1 protein - a transcription factor integral in EndoMT. In vivo, lentiviral gene transfer of either BRD4 shRNA or dominant negative BRD4-specific BD2 mitigated neointimal development in rat jugular veins grafted to carotid arteries.\n\nConclusionsOur data reveal the BD2 domain of BRD4 as a determinant driving EndoMT in vitro and neointimal formation in vivo. These findings provide new insight into BET biology, while offering prospects of specific BET domain targeting as an approach to limiting neointima and extending vein graft patency.

pathology↗

bioRxiv2018

Tissue repair in the mouse liver following acute carbon tetrachloride depends on injury-induced Wnt/β-catenin signaling

In the liver, Wnt/{beta}-catenin signaling is involved in regulating zonation and hepatocyte proliferation during homeostasis. We have examined Wnt gene expression and signaling after injury and we show by in situ hybridization that Wnts are activated by acute carbon tetrachloride (CCl4) toxicity. Following injury, peri-injury hepatocytes become Wnt-responsive, expressing the Wnt target gene Axin2. Lineage tracing of peri-injury Axin2+ hepatocytes shows that during recovery, the injured parenchyma becomes repopulated and repaired by Axin2+ descendants. Using single cell RNA sequencing (scRNA-seq), we show that endothelial cells are the major source of Wnts following acute CCl4 toxicity. Induced loss of {beta}-catenin in peri-injury hepatocytes results in delayed repair and ultimately to injury-induced lethality, while loss of Wnt production from endothelial cells leads to a delay in the proliferative response after injury.\n\nConclusionOur Pindings highlight the importance of the Wnt/{beta}-catenin signaling pathway in restoring tissue integrity following acute liver toxicity and establishes a role of endothelial cells as an important Wnt-producing regulator of liver tissue repair following localized liver injury.

developmental biology↗

bioRxiv2018

Simulation of transcranial magnetic stimulation in head model with morphologically-realistic cortical neurons

Transcranial magnetic stimulation (TMS) enables non-invasive modulation of brain activity with both clinical and research applications, but fundamental questions remain about the neural types and elements it activates and how stimulation parameters affect the neural response. We integrated detailed neuronal models with TMS-induced electric fields in the human head to quantify the effects of TMS on cortical neurons. TMS activated with lowest intensity layer 5 pyramidal cells at their intracortical axonal terminations in the superficial gyral crown and lip regions. Layer 2/3 pyramidal cells and inhibitory basket cells may be activated too, whereas direct activation of layers 1 and 6 was unlikely. Neural activation was largely driven by the field magnitude, contrary to theories implicating the field component normal to the cortical surface. Varying the induced currents direction caused a waveform-dependent shift in the activation site and provided a mechanistic explanation for experimentally observed differences in thresholds and latencies of muscle responses. This biophysically-based simulation provides a novel method to elucidate mechanisms and inform parameter selection of TMS and other forms of cortical stimulation.

bioengineering↗

bioRxiv2018

βVPE is involved in tapetal degradation and pollen development by activating proprotease maturation in Arabidopsis thaliana

Vacuolar processing enzyme (VPE) is responsible for the maturation and activation of vacuolar proteins in plants. We found that {beta}VPE was involved in tapetal degradation and pollen development by transforming proproteases into mature protease in Arabidopsis thaliana. {beta}VPE was expressed specifically in the tapetum from stages 5-8 of anther development. The {beta}VPE protein first appeared as a proenzyme and transformed into the mature enzyme before stages 7-8. The recombinant {beta}VPE protein self-cleaved and transformed to a 27-kD mature protein at pH 5.2. The mature {beta}VPE protein could induce the maturation of CEP1 in vitro. {beta}vpe mutants exhibited delayed vacuolar degradation and decreased pollen fertility. The maturation of CEP1, RD19A, and RD19C were seriously inhibited in {beta}vpe mutants. Our results indicate that {beta}VPE is a crucial processing enzyme that directly participates in the maturation of cysteine proteases before vacuolar degradation, and is indirectly involved in pollen development and tapetal cell degradation.\n\nHighlight{beta}VPE is a crucial processing enzyme that directly participates in the maturation of cysteine proteases before vacuolar degradation, and is indirectly involved in pollen development and tapetal cell degradation.

plant biology↗

bioRxiv2018

Phase Transitions in Mutualistic Communities under Invasion

Predicting the outcome of species invasion in ecosystems is a challenge of both theoretical and practical significance. Progress so far has been limited by the intractability of the far-from-equilibrium nature of the transitions that occur during invasion events. Here, we address this limitation by solving for the transition dynamics of a cross-feeding community along an analytically tractable manifold defined by the system carrying capacity. We find that species invasion induces discontinuous transitions in the community composition, resembling phase transitions in physical systems. These sharp transitions are emergent properties of species-resource interactions and relate directly to the extent of niche overlap between invasive and native species. The high susceptibility of community structure to small variations in species phenotype and resource conditions near the phase boundaries can explain empirically observed stochasticity and the emergence of tipping points in ecosystems. Moreover, we demonstrate that these phase transitions can be modulated by environmental variations to construct nonlinear organization of species both in space and time.

ecology↗

bioRxiv2018

Reference genes for RT-qPCR normalisation in different tissues, developmental stages and stress conditions of Hypericum perforatum

AbstactHypericum perforatum is a widely known medicinal herb used mostly as a remedy for depression because of its abundant secondary metabolites. Quantitative real-time PCR (qRT-PCR) is an optimized method for the efficient and reliable quantification of gene expression studies. In general, reference genes are used in qRT-PCR analysis because of their known or suspected housekeeping roles. However, their expression level cannot be assumed to remain stable under all possible experimental conditions. Thus, the identification of high quality reference genes is very necessary for the interpretation of qRT-PCR data. In this study, we investigated the expression of fourteen candidate genes, including nine housekeeping genes and five potential candidate genes. Additionally, the HpHYP1 gene, belonging to the PR-10 family associated with stress control, was used for validation of the candidate reference genes. Three programs were applied to evaluate the gene expression stability across four different plant tissues, three developmental stages and a set of abiotic stress and hormonal treatments. The candidate genes showed a wide range of Ct values in all samples, indicating that they are differentially expressed. Integrating all of the algorithms and evaluations, ACT2 and TUB-{beta} were the most stable combination overall and for different developmental stages samples. Moreover, ACT2 and EF1- were considered to be the two most applicable reference genes for different tissues and for stress samples. Majority of the conventional housekeeping genes exhibited better than the potential reference genes. The obtained results will contribute to improving credibility of standardization and quantification of transcription levels in future expression research of H. perforatum.

plant biology↗