Biology subjects

Guo, W.

Publications and source records attributed to Guo, W..

At least 199 records · Page 11Linked to original sources

bioRxiv2020

The RNA-binding protein HuR promotes nonalcoholic steatohepatitis (NASH) progression by enhancing death signaling pathway

Hepatocyte death triggers liver inflammation, liver injury, and fibrosis, which contributes to non-alcoholic steatohepatitis (NASH) pathogenesis. However, whether RNA processing regulates death signaling pathway during NASH progression is not investigated. In this study, we show that HuR, a widely expressed RNA-binding protein, promotes NASH progression by increasing DR5/caspase8/caspase3-mediated hepatocyte death. Cytosolic HuR levels are abnormally elevated in human patients with NASH. Hepatocyte-specific deletion of HuR protects against MCD-induced NASH by decreasing liver steatosis, inflammation and cell death, whereas hepatic overexpression of HuR induces liver injury by increasing DR5-induced hepatocyte death. Furthermore, in primary hepatocytes, HuR deficiency ameliorates PA&TNF-induced hepatocyte death due to decreased DR5/caspase8/caspase 3 signaling pathway while overexpression of HuR induces hepatocyte death by increasing DR5/caspase8/caspase 3 signaling pathway. Mechanistically, HuR directly binds to 3'-UTR of DR5 transcript and promotes its mRNA stability, contributing to the hepatocyte death during NASH progression. Our data reveal a novel mechanism by which HuR promotes mRNA stability of DR5, which contributes to NASH progression.

physiology↗

bioRxiv2020

Live imaging of breast tumors shows macrophage-dependent induction and TMEM-mediated enrichment of cancer stem cells during metastatic dissemination

Cancer stem cells (CSCs) play an important role during metastasis, but the dynamic behavior and induction mechanisms of CSCs are not well understood. We employed high-resolution intravital microscopy using a CSC biosensor to directly observe CSCs in live mice with mammary tumors. CSCs display the slow-migratory, invadopod-rich phenotype that is the hallmark of disseminating tumor cells. CSCs are enriched near macrophages, particularly near macrophage-containing intravasation sites called Tumor Microenvironment of Metastasis (TMEM) doorways. A dramatic enrichment of CSCs occurs on association with TMEM doorways, contributing to the finding that CSCs represent [~]>60% of circulating tumor cells. Mechanistically, stemness is induced in non-stem cancer cells upon their direct contact with macrophages via Notch signaling. In breast cancers from patients, the density of TMEM doorways correlates strongly with the proportion of cancer cells expressing stem cell markers, indicating that in human breast cancer TMEM doorways are not only cancer cell intravasation portals but also CSC programming sites. One Sentence SummaryIntravital imaging reveals macrophage-mediated induction of cancer stem cells in vivo and their dramatic enrichment on dissemination through TMEM doorways.

cancer biology↗

bioRxiv2020

RBM20 phosphorylation on serine/arginine domain is crucial to regulate pre-mRNA splicing and protein shuttling in the heart

Molecular and cellular mechanisms of mutations of splicing factors in heart function are not well understood. The splicing of precursor mRNA is dependent on an essential group of splicing factors containing serine-arginine (SR) domain(s) that are critical for protein-RNA and protein-protein interaction in the spliceosome assembly. Phosphorylation of SR domains plays a key role in splicing control and the distribution of splicing factors in the cell. RNA binding motif 20 (RBM20) is a splicing factor predominantly expressed in muscle tissues with the highest expression level in the heart. However, its phosphorylation status is completely unknown up-to-date. In this study, we identified sixteen amino acid residues that are phosphorylated by middle-down mass spectrometry. Four of them are located in the SR domain, and two out of the four residues, S638 and S640, play an essential role in splicing control and facilitate RBM20 shuttling from the nucleus to the cytoplasm. Re-localization of RBM20 promotes protein aggregation in the cytoplasm. We have also verified that SR-protein kinases (SRPKs), cdc2-like kinases (CLKs) and protein kinase B (PKB or AKT) phosphorylate S638 and S640. Mutations of S638A and S640G reduce RBM20 phosphorylation and disrupt the splicing. Taken together, we determine the phosphorylation status of RBM20 and provide the first evidence that phosphorylation on SR domain is crucial for pre-mRNA splicing and protein trafficking. Our findings reveal a new role of RBM20 via protein shuttling in cardiac function.

cell biology↗

bioRxiv2020

Distinct mechanisms for TMPRSS2 expression explain organ-specific inhibition of SARS-CoV-2 infection by enzalutamide

The coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has rapidly become a global public health threat due to the lack of effective drugs or vaccines against SARS-CoV-2. The efficacy of several repurposed drugs has been evaluated in clinical trials. Among these drugs, a relatively new antiandrogen agent, enzalutamide, was proposed because it reduces the expression of transmembrane serine protease 2 (TMPRSS2), a key component mediating SARS-CoV-2-driven entry into host cells, in prostate cancer cells. However, definitive evidence for the therapeutic efficacy of enzalutamide in COVID-19 is lacking. Here, we evaluated the antiviral efficacy of enzalutamide in prostate cancer cells, lung cancer cells, human lung organoids and SARS-CoV-2-infected Ad-ACE2-transduced Tmprss2 knockout (Tmprss2-KO) and wild-type (WT) mice. TMPRSS2 knockout significantly inhibited SARS-CoV-2 infection in vivo. Enzalutamide effectively inhibited SARS-CoV-2 infection in human prostate cancer cells (LNCaP) but not in human lung cancer cells or patient-derived lung organoids. Although Tmprss2 knockout effectively blocked SARS-CoV-2 infection in ACE2-transduced mice, enzalutamide showed no antiviral activity due to the AR independence of TMPRSS2 expression in mouse and human lung epithelial cells. Moreover, we observed distinct AR binding patterns between prostate cells and lung cells and a lack of direct binding of AR to TMPRSS2 in human lung cells. Thus, our findings do not support the postulated protective role of enzalutamide in treating COVID-19.

microbiology↗

bioRxiv2020

A Targeted Vaccine against COVID-19: S1-Fc Vaccine Targeting the Antigen-Presenting Cell Compartment Elicits Protection against SARS-CoV-2 Infection

Vaccination efficacy is enhanced by targeting the antigen-presenting cell compartment. Here, we show that S1-Fc antigen delivery targeting the Fc{gamma}R+ antigen-presenting cell compartment elicits anti-SARS-CoV-2 S1-antigen specific IgG production in vivo exerting biologically functional and protective activity against live virus infection, assessed in a stringent experimental virus challenge assay in vitro. The S1-domain of the SARS-CoV-2 spike protein was genetically fused to a human immunoglobulin Fc moiety, which contributes to mediate S1-Fc cellular internalization by Fc{gamma}R+ antigen-presenting cells. Immediately upon administration intramuscularly, our novel vaccine candidate recombinant rS1-Fc homes to lymph nodes in vivo where Fc{gamma}R+ antigen-presenting cells reside. Seroconversion is achieved as early as day 7, mounting considerably increased levels of anti-S1 IgGs in vivo. Interestingly, immunization at elevated doses with non-expiring S1-Fc encoding dsDNA favors the education of a desired antigen-specific adaptive T cell response. However, low-dose immunization, safeguarding patient safety, using recombinant rS1-Fc, elicits a considerably elevated protection amplitude against live SARS-CoV-2 infection. Our promising findings on rS1-Fc protein immunization prompted us to further develop an affordable and safe product for delivery to our communities in need for COVID-19 vaccinations.

microbiology↗

bioRxiv2020

Chromatin accessibility landscapes activated by cell surface and intracellular immune receptors

Activation of cell Surface and Intracellular Receptor-Mediated Immunity (SRMI and IRMI) results in rapid transcriptional reprogramming that underpins disease resistance. However, the mechanisms by which SRMI and IRMI lead to transcriptional changes are not clear. Here, we combine RNA-seq and ATAC-seq to define changes in gene expression and chromatin accessibility; both SRMI and IRMI increase chromatin accessibility at induced defense genes. Analysis of ATAC-seq and RNA-seq data combined with publicly available information on transcription factor DNA-binding motifs enabled comparison of individual gene regulatory networks activated by SRMI and IRMI, and by both. These results and analyses reveal overlapping and conserved transcriptional regulatory mechanism between the two immune systems.

plant biology↗

bioRxiv2020

Examining sex-differentiated genetic effects across neuropsychiatric and behavioral traits

BackgroundThe origin of sex differences in prevalence and presentation of neuropsychiatric and behavioral traits is largely unknown. Given established genetic contributions and correlations across these traits, we tested for a sex-differentiated genetic architecture within and between traits. MethodsUsing genome-wide association study (GWAS) summary statistics for 20 neuropsychiatric and behavioral traits, we tested for differences in SNP-based heritability (h2) and genetic correlation (rg<1) between sexes. For each trait, we computed z-scores from sex-stratified GWAS regression coefficients and identified genes with sex-differentiated effects. We calculated Pearson correlation coefficients between z-scores for each trait pair, to assess whether specific pairs share variants with sex-differentiated effects. Finally, we tested for sex differences in between-trait genetic correlations. ResultsWith current sample sizes (and power), we found no significant, consistent sex differences in SNP-based h2. Between-sex, within-trait genetic correlations were consistently high, although significantly less than 1 for educational attainment and risk-taking behavior. We identified genome-wide significant genes with sex-differentiated effects for eight traits. Several trait pairs shared sex-differentiated effects. The top 0.1% of genes with sex-differentiated effects across traits overlapped with neuron- and synapse-related gene sets. Most between-trait genetic correlation estimates were similar across sex, with several exceptions (e.g. educational attainment & risk-taking behavior). ConclusionsSex differences in the common autosomal genetic architecture of neuropsychiatric and behavioral phenotypes are small and polygenic, requiring large sample sizes. Genes with sex-differentiated effects are enriched for neuron-related gene sets. This work motivates further investigation of genetic, as well as environmental, influences on sex differences.

genetics↗

bioRxiv2020

Downy Mildew effector HaRxL21 interacts with the transcriptional repressor TOPLESS to promote pathogen susceptibility

Hyaloperonospora arabidopsidis (Hpa) is an oomycete pathogen causing Arabidopsis downy mildew. Effector proteins secreted from the pathogen into the plant play key roles in promoting infection by suppressing plant immunity and manipulating the host to the pathogens advantage. One class of oomycete effectors share a conserved RxLR motif critical for their translocation into the host cell. Here we characterize the interaction between an RxLR effector, HaRxL21 (RxL21), and the Arabidopsis transcriptional co-repressor Topless (TPL). We establish that RxL21 and TPL interact via an EAR motif at the C-terminus of the effector, mimicking the host plant mechanism for recruiting TPL to sites of transcriptional repression. We show that this motif, and hence interaction with TPL, is necessary for the virulence function of the effector. Furthermore, we provide evidence that RxL21 uses the interaction with TPL, and its close relative TPL-related 1, to repress plant immunity and enhance host susceptibility to both biotrophic and necrotrophic pathogens.

plant biology↗

bioRxiv2020

Hypoxia Depletes Contaminating CD45+ Hematopoietic Cells from Bone Marrow Stromal Cell (BMSC) Cultures: Methods for BMSC Culture Purification

Culture expanded bone marrow stromal cells (BMSCs) are easily isolated, can be grown rapidly en masse, and contain both skeletal stem cells (SSCs) and multipotent mesenchymal progenitors (MMPs). Despite this functional heterogeneity, BMSC cultures continue to be utilized for many applications due to the lack of definitive and universally accepted markers to prospectively identify and purify SSCs. Isolation of BMSCs is widely based on their adherence to tissue culture plastic; however, this method poses unique challenges when working with murine models. In particular, hematopoietic cell contamination is a significant impediment during the isolation or murine BMSCs. Several strategies to reduce hematopoietic contamination have been developed with varying levels of success. In this study, we found that up to 60% of cells isolated in standard BMSC cultures expressed the hematopoietic marker CD45. Remarkably, when cultured at a physiological oxygen tension of 1% O2, there was a 10-fold reduction in hematopoietic cells associated with a concomitant increase in PDGFR+ expressing stromal cells. This finding was due, in part, to a differential response of the two populations to low oxygen tension, or hypoxia. Specifically, in standard tissue culture conditions of 21% O2, CD45+ hematopoietic cells showed increased proliferation coupled with no observable changes in cell death when compared to their counterparts grown at 1% O2. In contrast, PDGFR+ stromal cells responded to hypoxia by increasing proliferation and exhibiting a 10-fold decrease in cell death. In summary, we describe a simple and reliable method exploiting the divergent biological response of hematopoietic and stromal cells to hypoxia to significantly increase the PDGFR+ stromal cell population in murine BMSC cultures.

cell biology↗

bioRxiv2020

The flagellar motor of Vibrio alginolyticus undergoes major structural remodeling during rotational switching

The bacterial flagellar motor is an intricate nanomachine that switches rotational directions between counterclockwise (CCW) and clockwise (CW) to direct the migration of the cell. The cytoplasmic ring (C-ring) of the motor, which is composed of FliG, FliM, and FliN, is known for controlling the rotational sense of the flagellum. However, the mechanism underlying rotational switching remains elusive. Here, we deployed cryo-electron tomography to visualize the C-ring in two rotational biased mutants (CCW-biased fliG-G214S and CW-locked fliG-G215A) in Vibrio alginolyticus. Sub-tomogram averaging was utilized to resolve two distinct conformations of the C-ring. Comparison of the C-ring structures in two rotational senses provide direct evidence that the C-ring undergoes major structural remodeling during rotational switch. Specifically, FliG conformational changes elicit a large rearrangement of the C-ring that coincides with rotational switching, whereas FliM and FliN form a spiral-shaped base of the C-ring, likely stabilizing the C-ring during the conformational remodeling.

microbiology↗

bioRxiv2020

Zika virus-based immunotherapy enhances long-term survival of rodents with brain tumors through upregulation of memory T-cells

Zika virus (ZIKV) exhibits a tropism for brain tumor cells and has been used as an oncolytic virus to target brain tumors in mice with modest effects on extending median survival. Recent studies have highlighted the potential for combining virotherapy and immunotherapy to target cancer. We postulated that ZIKV could be used as an adjuvant to enhance the long-term survival of mice with malignant glioblastoma and generate memory T-cells capable of providing long-term immunity against cancer remission. To test this hypothesis mice bearing malignant intracranial GL261 tumors were subcutaneously vaccinated with irradiated GL261 cells previously infected with the ZIKV. Mice also received intracranial injections of live ZIKV, irradiated ZIKV, or irradiated GL261 cells previously infected with ZIKV. Long-term survivors were rechallenged with a second intracranial tumor to examine their immune response and look for the establishment of protective memory T-cells. Mice with subcutaneous vaccination plus intracranial irradiated ZIKV or intracranial irradiated GL261 cells previously infected with ZIKV exhibited the greatest extensions to overall survival. Flow cytometry analysis of immune cells within the brains of long-term surviving mice after tumor rechallenge revealed an upregulation in the levels of T-cells, including CD4+ and tissue-resident memory CD4+ T-cells, in comparison to long-term survivors that were mock-rechallenged, and in comparison to naive untreated mice challenged with intracranial gliomas. These results suggest that ZIKV can serve as an adjuvant to subcutaneous tumor vaccines that enhance long-term survival and generate protective tissue-resident memory CD4+ T-cells.

cancer biology↗

bioRxiv2020

Structural basis for human TRPC5 channel inhibition by two distinct inhibitors

TRPC5 channel is a non-selective cation channel that participates diverse physiological processes. Human TRPC5 inhibitors show promise in the treatment of anxiety disorder, depression and kidney disease. Despite the high relevance of TRPC5 to human health, its inhibitor binding pockets have not been fully characterized due to the lack of structural information, which greatly hinders structure-based drug discovery. Here we show cryo-EM structures of human TRPC5 in complex with two distinct inhibitors, namely clemizole and HC-070, to the resolution of 2.7 [A]. Based on the high-quality cryo-EM maps, we uncover the different binding pockets and detailed binding modes for these two inhibitors. Clemizole binds inside the voltage sensor-like domain of each subunit, while HC-070 binds close to the ion channel pore and is wedged between adjacent subunits. Both of them exert the inhibitory function by stabilizing the ion channel in a closed state. These structures provide templates for further design and optimization of inhibitors targeting human TRPC5.

biophysics↗

bioRxiv2020

ERG orchestrates chromatin interactions to drive prostate cell fate reprogramming

While cancer is commonly perceived as a disease of dedifferentiation, the hallmark of early stage prostate cancer is paradoxically the loss of more plastic basal cells and the abnormal proliferation of more differentiated secretory luminal cells. However, the mechanism of prostate cancer pro-luminal differentiation is largely unknown. Through integrating analysis of the transcription factors (TFs) from 806 human prostate cancers, we have identified that ERG highly correlated with prostate cancer luminal subtyping. ERG overexpression in luminal epithelial cells inhibits its normal plasticity to transdifferentiate into basal lineage and ERG supersedes PTEN-loss which favors basal differentiation. ERG knock-out disrupted prostate cell luminal differentiation, whereas AR knock-out had no such effects. Trp63 is a known master regulator of prostate basal lineage. Through analysis of 3D chromatin architecture, we found that ERG binds and inhibits the enhancer activity and chromatin looping of a Trp63 distal enhancer, thereby silencing its gene expression. Specific deletion of the distal ERG binding site resulted in the loss of ERG-mediated inhibition of basal differentiation. Thus, ERG orchestrates chromatin interactions and regulates prostate cell lineage toward pro-luminal program, as its fundamental role on lineage differentiation in prostate cancer initiation.

cancer biology↗

bioRxiv2020

New gene association measures by joint network embedding of multiple gene expression datasets

Large number of samples are required to construct a reliable gene co-expression network, the samples from a single gene expression dataset are obviously not enough. However, batch effect may widely exist among datasets due to different experimental conditions. We proposed JEBIN (Joint Embedding of multiple BIpartite Networks) algorithm, it can learn a low-dimensional representation vector for each gene by integrating multiple bipartite networks, and each network corresponds to one dataset. JEBIN owns many inherent advantages, such as it is a nonlinear, global model, has linear time complexity with the number of genes, dataset or samples, and can integrate datasets with different distribution. We verified the effectiveness and scalability of JEBIN through a series of simulation experiments, and proved better performance on real biological data than commonly used integration algorithms. In addition, we conducted a differential co-expression analysis of hepatocellular carcinoma between the single-cell and bulk RNA-seq data, and also a contrast between the hepatocellular carcinoma and its adjacency samples using the bulk RNA-seq data. Analysis results prove that JEBIN can obtain comprehensive and stable gene co-expression networks through integrating multiple datasets and has wide prospect in the functional annotation of unknown genes and the regulatory mechanism inference of target genes.

bioinformatics↗

bioRxiv2020

Accurate Diagnosis of Colorectal Cancer Based on Histopathology Images Using Artificial Intelligence

BackgroundAccurate and robust pathological image analysis for colorectal cancer (CRC) diagnosis is time-consuming and knowledge-intensive, but is essential for CRC patients treatment. The current heavy workload of pathologists in clinics/hospitals may easily lead to unconscious misdiagnosis of CRC based on their daily image analyses. MethodsBased on a state-of-the-art transfer-learned deep convolutional neural network in artificial intelligence (AI), we proposed a novel patch aggregation strategy for clinic CRC prediction/diagnosis using weakly labeled pathological whole slide image (WSI) patches. This approach was trained and validated using an unprecedented and enormously large number of 170,099 patches, >14,680 WSIs, from >9,631 subjects that covered diverse and representative clinical cases from multi-independent-sources across China, U.S., and Germany. ResultsOur innovative AI tool was consistently nearly perfectly agreed with (average Kappa-statistic 0.896) and even often better than most of the experienced expert pathologists when tested in diagnosing CRC WSIs from multi-centers. The average area under the receiver operating characteristics curve (AUC) of AI was greater than that of the pathologists (0.981 vs 0.970) and achieved the best performance among the application of other AI methods to CRC diagnosis. Our AI-generated heatmap highlights the image regions of cancer tissue/cells. ConclusionsThis first-ever generalizable AI system can handle large amounts of WSIs consistently and robustly without potential bias due to fatigue commonly experienced by clinical pathologists. Hence, it will drastically alleviate the heavy clinical burden of daily pathology diagnosis, and improve the treatment for CRC patients. This tool is generalizable to other cancer diagnosis based on image recognition.

pathology↗

bioRxiv2020

Precision engineering of the transcription factor Cre1 in Hypocrea jecorina (Trichoderma reesei) for efficient cellulase production in the presence of glucose

In Trichoderma reesei, carbon catabolite repression (CCR) significantly downregulates the transcription of cellulolytic enzymes, which is usually mediated by the zinc finger protein Cre1. It was found that there is a conserved region at the C-terminus of Cre1/CreA in several cellulase-producing fungi that contains up to three continuous S/T phosphorylation sites. Here, S387, S388, T389, and T390 at the C-terminus of Cre1 in T. reesei were mutated to valine for mimicking an unphosphorylated state, thereby generating the transformants Tr_Cre1S387V, Tr_Cre1S388V, Tr_Cre1T389V, and Tr_Cre1T390V, respectively. Transcription of cel7a in Tr_ Cre1S388V was markedly higher than that of the parent strain when grown in glucose-containing media. Under these conditions, both filter paperase (FPase) and p-nitrophenyl-{beta}-D-cellobioside (pNPCase) activities, as well as soluble proteins from Tr_Cre1S388V were significantly increased by up to 2- to 3-fold compared with that of other transformants and the parent strain. To our knowledge, this is the first report demonstrating an improvement of cellulase production in fungal species under CCR by mimicking dephosphorylation at the C-terminus of Cre1. Taken together, we developed a precision engineering strategy based on the modification of phosphorylation sites of Cre1 transcription factor to enhance the production of cellulase in fungal species under CCR.

bioengineering↗

bioRxiv2020

Nonsense mediated RNA decay factor UPF1 is critical for post-transcriptional and translational gene regulation in Arabidopsis

Nonsense mediated RNA decay (NMD) is an evolutionary conserved RNA control mechanism that has also been implicated in the broader regulation of gene expression. Nevertheless, a role for NMD in genome regulation has not been fully assessed, partially because NMD inactivation is lethal in many organisms. Here, we performed in depth comparative analysis of Arabidopsis mutants lacking key proteins involved in different steps of NMD. We observed that UPF3, UPF1, and SMG7 have different impacts on NMD and the Arabidopsis transcriptome, with UPF1 having the biggest effect. Transcriptome assembly using stringent pipeline in UPF1-null plants revealed genome wide changes in alternative splicing, including switches in mRNA variants, suggesting a role for UPF1 in splicing. We further found that UPF1 inactivation leads to translational repression, manifested by a global shift in mRNAs from polysomes to monosomes and a downregulation of genes involved in translation and ribosome biogenesis. Despite this global change, NMD targets and low-expressed mRNAs with short half-lives were enriched in polysomes, indicating that UPF1 specifically suppresses the translation of aberrant RNAs. Particularly striking was an increase in the translation of TIR domain-containing, nucleotide-binding, leucine-rich repeat (TNL) immune receptors. The regulation of TNLs via UPF1/NMD-mediated mRNA stability and translational de-repression offers a dynamic mechanism for the rapid activation of TNLs in response to pathogen attack.

molecular biology↗

bioRxiv2020

Latent learning drives sleep-dependent plasticity in distinct CA1 subpopulations

Latent learning allows the brain the transform experiences into cognitive maps, a form of implicit memory, without reinforced training. Its mechanism is unclear. We tracked the internal states of the hippocampal neural ensembles and discovered that during latent learning of a spatial map, the state space evolved into a low-dimensional manifold that topologically resembled the physical environment. This process requires repeated experiences and sleep in-between. Further investigations revealed that a subset of hippocampal neurons, instead of rapidly forming place fields in a novel environment, remained weakly tuned but gradually developed correlated activity with other neurons. These weakly spatial neurons bond activity of neurons with stronger spatial tuning, linking discrete place fields into a map that supports flexible navigation.

neuroscience↗