Biology subjects

Bei, J.

Publications and source records attributed to Bei, J..

13 recordsLinked to original sources

bioRxiv2025

Mechanistic insights and in vivo HIV suppression by the BRD4-targeting small molecule ZL0580

Epigenetic suppression and durable silencing of HIV represent a promising strategy to achieve ART-free remission, consistent with the "block and lock" HIV cure paradigm. BRD4 is a host epigenetic reader and plays a critical role in HIV transcriptional regulation. We previously identified ZL0580, a first-in-class BRD4-selective small molecule distinct from the pan-BET inhibitor JQ1, which induces HIV epigenetic suppression. However, detailed molecular mechanisms, pharmacokinetics (PK), and in vivo HIV-suppressive efficacy of ZL0580 remain undefined. Here, we show that ZL0580 selectively targets BRD4 bromodomain 1 (BD1) through interaction with a key glutamic acid residue (E151), as determined by structural modeling and mutagenesis. Transcriptomic profiling by RNA-seq reveals that ZL0580 and JQ1 induce opposing gene expression programs, consistent with their distinct effects on HIV proviral transcription and latency. In a humanized mouse model of HIV infection, ZL0580 monotherapy, or in combination with ART, potently suppressed active HIV replication, reducing the plasma viremia to nearly undetectable levels, and delayed viral rebound following treatment interruption. Collectively, these findings establish ZL0580 as an epigenetic suppressor of HIV in vivo and provide proof-of-concept for its potential as a "block and lock" HIV cure candidate, warranting further optimization and development.

microbiology↗

bioRxiv2025

mRNA-delivered neutralizing antibodies confer protection against SARS-CoV-2 variant in the lower and upper respiratory tract

Monoclonal antibodies (mAbs) have been developed as effective biological countermeasures against a range of human diseases. The high cost of antibody production and manufacturing limits its clinical application and widespread use. The mRNA-lipid nanoparticle (mRNA-LNP) is a versatile platform for development of vaccines and protein-replacement therapeutics. Since the COVID-19 pandemic, a number of neutralizing mAbs against SARS-CoV-2 have been identified with several being used clinically under emergency authorization. Herein, we report the design and generation of mRNA-LNPs expressing two SARS-CoV-2 neutralizing mAbs, 76E1 and LY1404, which respectively target the viral spike proteins fusion peptide (FP) epitope within the S2 subunit and the receptor-binding domain (RBD) within the S1 subunit. We show a single intramuscular administration of mRNA-LNPs results in efficient LY1404 and 76E1 mAb production in mice which is sustained for 7-14 days. Further, we evaluate the protective efficacy of mRNA-LNP formulations encoding the two antibodies in mouse and hamster models challenged with different SARS-CoV-2 viral strains. The data demonstrate that a single administration of mRNA-LNP encoding the more broadly neutralizing antibody 76E1 confers significant protection against the immune-evasive SARS-CoV-2 Omicron variant BQ.1 in both the upper and lower respiratory tract of the hamsters, indicating its potential impact on limiting both viral disease and viral acquisition. Together, our study expands the potential of the mRNA-LNP platform to deliver therapeutic antibodies for rapid prevention or treatment of pathogenic infections.

microbiology↗

bioRxiv2025

Circulating extracellular vesicles from HIV-1 gp120-treated mice act as endogenous algogens, mediating and maintaining HIV-associated chronic pain

HIV-associated chronic pain (HIV-PAIN) remains prevalent in the post combined antiretroviral therapy era, affecting 30-60% of HIV patients worldwide. The underlying mechanism responsible for the development and maintenance of chronic pain remains unclear. gp120 is a causal factor of the HIV-PAIN and functions as an exogenous algogen. The pain experienced by human HIV-PAIN has been modeled in mice (referred to as mHIV-PAIN) using intrathecal (i.t.) injections of gp120. gp120 is a relatively short-term, static, exogenous algogen that is exhaustible in vivo. In authentic infection, HIV virions serve as the primary source of exogenous gp120, which initiates the early phase of clinical HIV-PAIN. Interestingly, while the source of replenishing gp120 decreases after antiretroviral therapy by suppressing viremia, the prevalence of chronic HIV-PAIN remains stable. To induce chronic pain in mice, gp120 needs to be repeatedly applied by the i.t. route. This raises a key question: Is an endogenous inexhaustible algogen responsible for maintaining the chronicity of HIV-PAIN? In the present study, we isolated circulating small extracellular vesicles (sEV) from mice using our mHIV-PAIN model that is i.t. injected with gp120. We refer to such sEV as gp120-sEV herein. We observed that gp120 is absent in gp120-sEV. Following transfusion of gp120-sEV intrathecally, naive recipient mice exhibit an extensive pain phenotype, including cold pain tested with we newly invented dry ice vapor cold test. RNA-sequence analysis suggests that gp120-sEVs induced expression of genes related to nociception and neuroinflammation pathways. These findings provide direct evidence that circulating sEV function as endogenous long-term "dynamic" algogens that enhance initial pain and extend the chronification of HIV-PAIN in mice, suggesting that chronic HIV-PAIN requires an exogenous algogen (gp120) paired with endogenous algogen (gp120-sEV), and that these components work synchronically to initiate and extend pain chronification. This double algogen concept provides a new insight into the pathogenesis of HIV-PAIN chronification. Our new mechanistic understanding will also assist in identifying new therapeutics to alleviate HIV-PAIN by targeting pathological gp120-sEV.

neuroscience↗

bioRxiv2025

Global expression profile of Enterohemorrhagic Escherichia coli O157:H7 in phagosome of murine macrophages

EHEC O157:H7 is responsible for Hemolytic Uremic Syndrome (HUS) outbreaks in humans and its virulence is associated with Shiga toxin (Stx). During the pathogenesis of HUS, EHEC O157:H7 is phagocytized by the intestinal host macrophage into the phagosome. According to our study, a low percentage of phagocytized bacteria survived in the murine macrophage phagosome. To understand the initials mechanisms involved in bacterial persistence, we performed a global profile of bacterial RNAseq analysis in the early phagosome of murine macrophage and in vitro assays with more extreme stress conditions. The defense strategy of an early murine phagosome consisted on producing DNA damage, membrane damage, acid pH and nitric oxide (NO) agents. These hostile conditions triggering the bacterial SOS response, lipid biosynthesis and NO detoxification enzymes in response to DNA damage, membrane damage and nitric oxide agents, respectively. In turn, to survive, the bacterium conserves energy by downregulating flagellar biosynthesis, T3SS and T6SS virulence mechanisms. In contrast, stx2a expression is upregulated. At the same time, it increases the ribosomal levels and amino acid synthesis to react more effectively against adverse conditions. Under more extreme stress conditions, EHEC O157:H7 expressed genes related to an acidic environment (pH 4.5) upregulating the acid stress response pathways AR2-5, whereas, upon high concentrations of hydrogen peroxide, it transcribed higher levels of genes of the OxyR pathway and subsequently hydrogen peroxide oxidative stress-related genes.

microbiology↗

bioRxiv2023

Circulating exosomes from Alzheimer's disease suppress VE-cadherin expression and induce barrier dysfunction in recipient brain microvascular endothelial cell

BackgroundBlood-brain barrier (BBB) breakdown is a component of the progression and pathology of Alzheimers disease (AD). BBB dysfunction is primarily caused by reduced or disorganized tight junction or adherens junction proteins of brain microvascular endothelial cell (BMEC). While there is growing evidence of tight junction disruption in BMECs in AD, the functional role of adherens junctions during BBB dysfunction in AD remains unknown. Exosomes secreted from senescent cells have unique characteristics and contribute to modulating the phenotype of recipient cells. However, it remains unknown if and how these exosomes cause BMEC dysfunction in AD. ObjectivesThis study aimed to investigate the potential roles of AD circulating exosomes and their RNA cargos in brain endothelial dysfunction in AD. MethodsWe isolated exosomes from sera of five cases of AD compared with age- and sex-matched cognitively normal controls using size-exclusion chromatography technology. We validated the qualities and particle sizes of isolated exosomes with nanoparticle tracking analysis and atomic force microscopy. We measured the biomechanical natures of the endothelial barrier of BMECs, the lateral binding forces between live BMECs, using fluidic force miscopy. We visualized the paracellular expressions of the key adherens junction protein VE-cadherin in BMEC cultures and a 3D BBB model that employs primary human BMECs and pericytes with immunostaining and evaluated them using confocal microscopy. We also examined the VE-cadherin signal in brain tissues from five cases of AD and five age- and sex-matched cognitively normal controls. ResultsWe found that circulating exosomes from AD patients suppress the paracellular expression levels of VE-cadherin and impair the barrier function of recipient BMECs. Immunostaining analysis showed that AD circulating exosomes damage VE-cadherin integrity in a 3D model of microvascular tubule formation. We found that circulating exosomes in AD weaken the BBB depending on the RNA cargos. In parallel, we observed that microvascular VE-cadherin expression is diminished in AD brains compared to normal controls. ConclusionUsing in vitro and ex vivo models, our study illustrates that circulating exosomes from AD patients play a significant role in mediating the damage effect on adherens junction of recipient BMEC of the BBB in an exosomal RNA-dependent manner. This suggests a novel mechanism of peripheral senescent exosomes for AD risk.

neuroscience↗

bioRxiv2023

Identification of common sequence motifs shared exclusively among selectively packed exosomal pathogenic microRNAs during rickettsial infections

We previously reported that microRNA (miR)23a and miR30b are selectively sorted into rickettsia-infected, endothelial cell-derived exosomes (R-ECExos). Yet, the mechanism remains unknown. The number of cases of spotted fever rickettsioses has been increasing in recent years, and infections with these bacteria cause life-threatening diseases by targeting brain and lung tissues. Therefore, the aim of the present study is to continue to dissect the molecular mechanism underlying R-ECExos-induced barrier dysfunction of normal recipient microvascular endothelial cells (MECs), depending on their exosomal RNA cargos. Rickettsiae are transmitted to human hosts by the bite of an infected tick into the skin. In the present study we demonstrate that treatment with R-ECExos, which were derived from spotted fever group R parkeri infected human dermal MECs, induced disruptions of the paracellular adherens junctional protein VE-cadherin and breached the paracellular barrier function in recipient pulmonary MECs (PMECs) in an exosomal RNA-dependent manner. Similarly, we did not detect different levels of miRs in parent dermal MECs following rickettsial infections. However, we demonstrated that the microvasculopathy-relevant miR23a-27a-24 cluster and miR30b are selectively enriched in R-ECExos. Bioinformatic analysis revealed that common sequence motifs are shared exclusively among the exosomal, selectively-enriched miR23a cluster and miR30b at different levels. Taken together, these data warrant further functional identification and characterization of a single, bipartition, or tripartition among ACA, UCA, and CAG motifs that guide recognition of microvasculopathy-relevant miR23a-27a-24 and miR30b, and subsequently results in their selective enrichments in R-ECExos.

molecular biology↗

bioRxiv2022

Exosomally targeting microRNA23a ameliorates microvascular endothelial barrier dysfunction following rickettsial infection

Spotted fever group rickettsioses caused by Rickettsia (R) are devastating human infections, which mainly target microvascular endothelial cells (EC) and can induce lethal EC barrier dysfunction in the brain and lungs. Our previous evidence reveals that exosomes (Exos) derived from rickettsial-infected ECs, namely R-ECExos, can induce disruption of the tight junctional (TJ) protein ZO-1 and barrier dysfunction of human normal recipient brain microvascular endothelial cells (BMECs). However, the underlying mechanism remains elusive. Given that we have observed that microRNA23a (miR23a), a negative regulator of endothelial ZO-1 mRNA, is selectively sorted into R-ECExos, the aim of the present study was to characterize the potential functional role of exosomal miR23a delivered by R-ECExos in normal recipient BMECs. We demonstrated that EC-derived Exos (ECExos) have the capacity to deliver oligonucleotide RNAs to normal recipient BMECs in an RNase-abundant environment. miR23a in ECExos impairs normal recipient BMEC barrier function, directly targeting TJ protein ZO-1 mRNAs. In separate studies using a traditional in vitro model and a novel single living-cell biomechanical assay, our group demonstrated that miR23a anti-sense oligonucleotide-enriched ECExos ameliorate R-ECExo-provoked recipient BMEC dysfunction in association with stabilization of ZO-1 in a dose-dependent manner. These results suggest that Exo-based therapy could potentially prove to be a promising strategy to improve vascular barrier function during bacterial infection and concomitant inflammation.

cell biology↗

bioRxiv2021

Host EPAC1 modulates rickettsial adhesion to vascular endothelial cells via regulation of ANXA2 Y23 phosphorylation

Recently we have identified that endothelial surface annexin A2 (ANAX2) functions as a receptor for spotted fever group rickettsial adhesin outer membrane protein B (OmpB), which binds to the endothelial cell (EC) surface. Moreover, we reported that intracellular cAMP receptor EPAC1 modulates ANXA2 tyrosine (Y) 23 phosphorylation, and inactivation of EPAC1 suppresses ANXA2 expression on the EC luminal surface by downregulating Y23 phosphorylation. Since we reported that EPAC1 plays a critical role in the initial step to successfully establish rickettsial infection of ECs, this work aims to answer the following: (a) What is the mechanism underlying the regulatory role of EPAC1 in ECs during the initial step of bacterial infection? (b) Is the EPAC1-ANXA2 signaling pathway involved in the regulation of rickettsial adhesion to ECs? In the present study, an established system that is anatomically-based and quantifies bacterial adhesion to ECs in vivo was combined with novel fluidic force microscopy (FluidFM) to dissect the functional role of the EPAC1-ANXA2 signaling pathway in rickettsiae-EC adhesion. We reveal that the deletion of the EPAC1 gene impedes rickettsial binding to endothelium in vivo. In addition, single living brain microvascular EC study that employs FluidFM and site-directed mutagenesis provides evidence that supports our finding that EPAC1 governs rickettsial adhesion to EC surfaces via regulation of ANXA2 Y23 phosphorylation.

pathology↗

bioRxiv2021

Full-length Genome of the Ogataea polymorpha strain HU-11 reveals large duplicated segments in subtelomeic regions

BackgroundCurrently, methylotrophic yeasts (e.g., Pichia pastoris, Ogataea polymorpha, and Candida boindii) are subjects of intense genomics studies in basic research and industrial applications. In the genus Ogataea, most research is focused on three basic O. polymorpha strains--CBS4732, NCYC495, and DL-1. However, the relationship between CBS4732, NCYC495, and DL-1 remains unclear, as the genomic differences between them have not be exactly determined without their high-quality complete genomes. As a nutritionally deficient mutant derived from CBS4732, the O. polymorpha strain CBS4732 ura3{Delta} (named HU-11) is being used for high-yield production of several important proteins or peptides. HU-11 has the same reference genome as CBS4732 (noted as HU-11/CBS4732), because the only genomic difference between them is a 5-bp insertion. ResultsIn the present study, we have assembled the full-length genome of O. polymorpha HU-11/CBS4732 using high-depth PacBio and Illumina data. Long terminal repeat (LTR) retrotransposons, rDNA, 5 and 3 telomeric, subtelomeric, low complexity and other repeat regions were curated to improve the genome quality. Particularly, we detected large duplicated segments (LDSs) in the subtelomeric regions and exactly determined all the structural variations (SVs) between CBS4732 and NCYC495. New findings mainly include(1) the genomic differences between HU-11/CBS4732 and NCYC495 include single nucleotide polymorphisms, small insertions and deletions, and only three SVs; (2) six genes were incorporated into CBS4732 from Cyberlindnera jadinii by horizontal gene transfer and may bring HU-11/CBS4732 new biological functions or physiological properties; (3) many recombination events may have occurred on chromosome 4 and 5 of CBS4732 and NCYC495 ancestors and two large segments were acquired by CBS4732 and NCYC495 from chromosome 6 and C. jadinii during recombination, respectively; and (4) the genome expansion in methylotrophic yeasts is mainly driven by large segment duplication in subtelomeric regions. ConclusionsThe present study preliminarily revealed the complex relationship between CBS4732, NCYC495, and DL-1. The new findings provide new opportunities for in-depth understanding of genome evolution in methylotrophic yeasts and lay the foundations for the industrial applications of O. polymorpha CBS4732, NCYC495, DL-1, and their derivative strains. The full-length genome of the O. polymorpha strain HU-11/CBS4732 should be included into the NCBI RefSeq database for future studies of Ogataea spp..

bioinformatics↗

bioRxiv2021

How the replication and transcription complex of SARS-CoV-2 functions in leader-to-body fusion

BackgroundCoronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Although unprecedented efforts are underway to develop therapeutic strategies against this disease, scientists have acquired only a little knowledge regarding the structures and functions of the CoV replication and transcription complex (RTC) and 16 non-structural proteins, named NSP1-16. ResultsIn the present study, we proposed a two-route model to answer how the RTC functions in the jumping transcription of CoVs. The key step leading to this model was that the motif AAACH for METTL3 recognition flanking the transcription regulatory sequence (TRS) motif was discovered to determine the m6A methylation of SARS-CoV-2 RNAs, by reanalyzing public Nanopore RNA-seq data. As the most important finding, TRS hairpins were reported for the first time to interpret NSP15 cleavage, RNA methylation of CoVs and their association at the molecular level. In addition, we reported canonical TRS motifs of all CoVs to prove the importance of our findings. ConclusionsThe main conclusions are: (1) TRS hairpins can be used to identify recombination regions in CoV genomes; (2) RNA methylation of CoVs participates in the determination of the RNA secondary structures by affecting the formation of base pairing; and (3) The eventual determination of the CoV RTC global structure needs to consider METTL3 in the experimental design. Our findings enrich fundamental knowledge in the field of gene expression and its regulation, providing a crucial basis for future studies.

bioinformatics↗

bioRxiv2020

Endothelial exosome plays functional role during rickettsial infection

Spotted fever group rickettsioses (SFRs) are devastating human infections. Vascular endothelial cells (ECs) are the primary targets of infection. Edema resulting from EC barrier dysfunction occurs in the brain and lungs in most cases of lethal SFR, but the underlying mechanisms remain unclear. The aim of the study is to explore the potential role of Rickettsia (R)-infected, EC-derived exosomes (Exos) during infection. Using size-exclusion chromatography (SEC), we purified Exos from conditioned, filtered, bacteria-free media collected from R-infected human umbilical vein ECs (HUVECs) (R-ECExos) and plasma of R-infected mice (R-plsExos). We observed that rickettsial infection increases the release of heterogeneous plsExos, but endothelial exosomal size, morphology, and production were not significantly altered following infection. Compared to normal plsExos and ECExos, both R-plsExos and R-ECExos induced dysfunction of recipient normal brain microvascular Ecs (BMECs). The effect of R-plsExos on mouse recipient BMEC barrier function is dose-dependent. The effect of R-ECExos on human recipient BMEC barrier function is dependent on exosomal RNA cargo. Next-generation sequencing analysis and stem-loop quantitative reverse transcription PCR (RT-qPCR) validation revealed that R infection triggered the selective enrichment of endothelial exosomal mir-23a and mir-30b, which target the endothelial barrier. To our knowledge, this is the first report on the functional role of extracellular vesicles following infection by obligately intracellular bacteria. ImportanceSpotted fever group rickettsioses are devastating human infections. Vascular endothelial cells are the primary targets of infection. Edema resulting from endothelial barrier dysfunction occurs in the brain and lungs in most cases of lethal rickettsioses, but the underlying mechanisms remain unclear. The aim of the study is to explore the potential role of Rickettsia-infected, endothelial cell-derived exosomes during infection. We observed that rickettsial infection increases the release of heterogeneous plasma Exos, but endothelial exosomal size, morphology, and production were not significantly altered following infection. Rickettsia-infected, endothelial cell-derived exosomes induced dysfunction of recipient normal brain microvascular endothelial cells. The effect is dependent on exosomal RNA cargo. Next-generation sequencing analysis revealed that rickettsial infection triggered the selective enrichment of endothelial exosomal mir-23a and mir-30b, which target the endothelial barrier. To our knowledge, this is the first report on the functional role of extracellular vesicles following infection by obligately intracellular bacteria.

microbiology↗

bioRxiv2020

Full-length genome sequence of segmented RNA virus from ticks was obtained using small RNA sequencing data

In 2014, A novel tick-borne virus of the genus Flavivirus was first reported from the Mogiana region in Brazil. This virus was named the Mogiana tick virus (MGTV). Later, MGTV was also named as Jingmen tick virus (JMTV), Kindia tick virus (KDTV), Guangxi tick virus (GXTV) etc. In the present study, we used small RNA sequencing (sRNA-seq) to detect viruses in ticks and detected MGTV in Amblyomma testudinarium ticks, which had been captured in Yunnan province of China in the year of 2016. The full-length genome sequence of a new MGTV strain Yunnan2016 (GenBank: MT080097, MT080098, MT080099 and MT080100) was obtained and recommended to be included into the NCBI RefSeq database for the future studies of MGTV. Our phylogenetic analyses showed that viruses named MGTV, JMTV, KDTV and GXTV are monophyletic: the MGTV group (lineage) of viruses. We show, for the first time, that 5' and 3' sRNAs can be used to obtain full-length sequences of the 5 and 3 ends of, but not limited to genome sequences of RNA viruses. And we proved the feasibility of using the sRNA-seq based method for the detection of viruses in a sample containing miniscule RNA.

bioinformatics↗

bioRxiv2018

Quantification of Enterohemorrhagic Escherichia coli O157:H7 proteome using TMT-Based Analysis

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a human pathogen responsible for diarrhea, hemorrhagic colitis and hemolytic uremic syndrome (HUS). EHEC infection is distributed worldwide and numerous outbreaks of diseases caused by enterohemorrhagic have been reported. To promote a comprehensive insight into the molecular basis of EHEC O157:H7 physiology and pathogenesis, the combined proteome of EHEC O157:H7 strains, Clade 8 and Clade 6 isolated from cattle in Argentina, and the standard EDL933 (clade 3) strain has been analyzed. TMT (Tandem Mass Tags)-based quantitative proteomic and emPAI analyses were performed to estimate the protein abundance in EHEC proteome. 2,234 non-redundant proteins of EHEC O157:H7 were identified. A comparison of this result with in silico data of EHEC O157:H7 genome showed that approximately 40% of the predicted proteome of this pathogen were covered. According to the emPAI analysis, 85 proteins were among the most abundant (e.g. GAPDH, FliC H-antigen, Enolase, and GroEL). Tellurite resistance proteins were also highly abundant. COG analysis showed that although most of the identified proteins are related to cellular metabolism, the majority of the most abundant proteins are associated with translation processes. A KEGG enrichment analysis revealed that Glycolysis / Gluconeogenesis was the most significant pathway. On the other hand, the less abundant detected proteins are those related to DNA processes, cell respiration and prophage. Among the proteins that composed the Type III Secretion System, the most abundant protein was EspA. Altogether, the results show a subset of important proteins that contribute to physiology and pathogenicity of EHEC O157:H7.\n\nIMPORTANCEThe study of the abundance of proteins present within a complex mixture of proteins in a cell, under different conditions, can provide important information about the activities of individual protein components and protein networks that are cornerstones for the comprehension of physiological adaptations in response to biological demands promoted by environmental changes. We generated a comprehensive and accurate quantitative list of EHEC O157:H7 proteome, which provides a description of the most abundant proteins produced by this pathogen that were related to physiology and pathogenesis of EHEC. This study provides information and extends the understanding on functional genomics and the biology of this pathogen.

microbiology↗