Search bioRxiv⌕ Search

Biology subjects

Lu, G.

Publications and source records attributed to Lu, G..

At least 73 records · Page 4Linked to original sources

MoIsw2 activity could be the missing link between adaptation and mutation instrumental for natural adaptation-directed fast evolution (NADFE) of Magnaporthe oryzae

Isw2 proteins are conserved in eukaryotes and are known to bind to DNA and dynamically influence local chromosome condensation close to their DNA binding site in an ATP-dependent manner making genes close to the binding sites more accessible for transcription and repression. A putative MoISW2 gene was deleted with large effects on plant pathogenicity as a result. The gene was complemented and a ChIP-sec was performed to identify binding sites. RNAsec showed effects on the overall regulation of genes along the chromosomes for mutant and background strains and this was compared with RNAseq from 55 downloaded RNA-seq datasets from the same strain and found similar. MoIsw2 binding and activities create genomic regions affected by MoIsw2 with high gene expression variability close to the MoIsw2 binding sites while surrounding regions have lower gene expression variability. The genes affected by the MoIsw2 activity are niche-determinant genes (secreted proteins, secondary metabolites and stress-coping genes) and avirulence genes. We further show that MoIsw2 binding sites with the DNA binding motifs coincide with known transposable elements (TE) making it likely that TE-transposition at the binding sites can affect the transcription profile of M. oryze in a strain-specific manner. We conclude that MoIsw2 is a likely candidate for a master regulator, regulating the dynamic balance between biomass growth genes (like housekeeping genes) and nich-determinant genes important for ecological fitness. Stress-induced TE transposition is together with MoIsw2 activity a likely mechanism creating more mutations and faster evolution of the niche-determinant genes than for housekeeping genes.

evolutionary biology↗

Splicing factor BUD31 promotes ovarian cancer progression through sustaining the expression of anti-apoptotic BCL2L12

Dysregulated expression of splicing factors has important roles in cancer development and progression. However, it remains a challenge to identify the cancer-specific splicing variants. Here we demonstrated that spliceosome component BUD31 is increased in ovarian cancer, and its higher expression predicts worse prognosis. We characterized the BUD31 binding motif and found that BUD31 preferentially binds exon-intron regions near splicing sites by CLIP-seq. Further analysis revealed that BUD31 inhibition results in extensive exon skipping and decreased abundance of long CDS isoforms. In particular, we identified BCL2L12, an anti-apoptotic BCL2 family member, as a functional splicing target of BUD31. BUD31 stimulates the inclusion of exon 3 to generate full-length BCL2L12 and promotes ovarian cancer progression. Knockdown of BUD31 or splice-switching antisense oligonucleotide treatment promotes exon 3 skipping and results in a truncated isoform of BCL2L12 that undergoes nonsense-mediated mRNA decay, and the cells subsequently undergo apoptosis. Our findings reveal BUD31-regulated exon inclusion as a critical factor in ovarian cancer cell survival and progression.

cancer biology↗

The SARS-CoV-2 Omicron (B.1.1.529) variant exhibits altered pathogenicity, transmissibility, and fitness in the golden Syrian hamster model

The newly emerging SARS-CoV-2 Omicron (B.1.1.529) variant first identified in South Africa in November 2021 is characterized by an unusual number of amino acid mutations in its spike that renders existing vaccines and therapeutic monoclonal antibodies dramatically less effective. The in vivo pathogenicity, transmissibility, and fitness of this new Variant of Concerns are unknown. We investigated these virological attributes of the Omicron variant in comparison with those of the currently dominant Delta (B.1.617.2) variant in the golden Syrian hamster COVID-19 model. Omicron-infected hamsters developed significantly less body weight losses, clinical scores, respiratory tract viral burdens, cytokine/chemokine dysregulation, and tissue damages than Delta-infected hamsters. The Omicron and Delta variant were both highly transmissible (100% vs 100%) via contact transmission. Importantly, the Omicron variant consistently demonstrated about 10-20% higher transmissibility than the already-highly transmissible Delta variant in repeated non-contact transmission studies (overall: 30/36 vs 24/36, 83.3% vs 66.7%). The Delta variant displayed higher fitness advantage than the Omicron variant without selection pressure in both in vitro and in vivo competition models. However, this scenario drastically changed once immune selection pressure with neutralizing antibodies active against the Delta variant but poorly active against the Omicron variant were introduced, with the Omicron variant significantly outcompeting the Delta variant. Taken together, our findings demonstrated that while the Omicron variant is less pathogenic than the Delta variant, it is highly transmissible and can outcompete the Delta variant under immune selection pressure. Next-generation vaccines and antivirals effective against this new VOC are urgently needed. One Sentence SummaryThe novel SARS-CoV-2 Omicron variant, though less pathogenic, is highly transmissible and outcompetes the Delta variant under immune selection pressure in the golden Syrian hamster COVID-19 model.

microbiology↗

Single-cell analysis of microglial transcriptomic diversity in subarachnoid hemorrhage

BackgroundSubarachnoid hemorrhage (SAH) is a severe stroke and the advanced treatment for SAH is still limited. Recent studies have shown that microglia-mediated neuroinflammation plays a critical role in the pathogenesis of SAH. Microglia can transform their states in response to central nervous system injury. However, the transcriptomic features of microglia remained unknown in SAH. Recent developed single-cell RNA sequencing (scRNA-seq) provides a possible way to solve this problem. MethodsEndovascular perforation (EVP) murine SAH model was established to reproduce experimental SAH. Microglia states are examined with immune staining and quantitate analysis. Post-SAH microglial single-cell suspension were harvest and sequenced using 10X scRNA-seq platform. Then, the detailed single-cell transcriptomic characterization of post-SAH microglia were analyzed with bioinformatics. ResultsTranscriptional analysis revealed at least ten diverse microglial subgroups, including SAH-associated microglia (SAM), inflammatory-associated microglia (IAM) and proliferation-associated microglia (PAM), which all exhibit distinct marker gene expression patterns. Microglia subsets interaction reveals the functional relationship between elevated signaling pathways and microglial sub-populations in SAH. Receptor-ligand pair analysis revealed that complex inter-cellular interactions exist between the microglia subsets and other cell types, and indicated that microglia are important mediators of neuroinflammation after SAH. Integrated analysis with normal microglia further proved the existence of these microglia subpopulations and different gene markers associated with SAH were clarified. ConclusionsCollectively, we first report the single-cell transcriptome of post-SAH microglia and found specific biomarkers related to the neuroinflammation in SAH. These results enhanced our understanding of the pathological mechanisms of microglial response to SAH, and may guide future development of SAH monitoring methods and therapeutics.

neuroscience↗

Metformin abrogates pathological TNF-α-producing B cells through mTOR-dependent metabolic reprogramming in polycystic ovary syndrome

B cells contribute to the pathogenesis of polycystic ovary syndrome (PCOS). Clinically, metformin is used to treat PCOS, but it is unclear whether metformin exerts its therapeutic effect by regulating B cells. Here, we showed that the expression level of TNF- in peripheral blood B cells from PCOS patient was increased. Metformin used in vitro and in vivo was able to reduce the production of TNF- in B cells from PCOS patient. Administration of metformin improved mouse PCOS phenotypes induced by dehydroepiandrosterone (DHEA) and also inhibited TNF- expression in splenic B cells. Further, metformin induced metabolic reprogramming of B cells in PCOS patients, including the alteration in mitochondrial morphology, the decrease in mitochondrial membrane potential, ROS production and glucose uptake. In DHEA-induced mouse PCOS model, metformin altered metabolic intermediates in splenic B cells. Moreover, the inhibition of TNF- expression and metabolic reprogramming in B cells of PCOS patients and mouse model by metformin were associated with decreased mTOR phosphorylation. Together, TNF--producing B cells are involved in the pathogenesis of PCOS, and metformin inhibits mTOR phosphorylation and affects metabolic reprogramming, thereby inhibiting TNF- expression in B cells, which may be a new mechanism of metformin in the treatment of PCOS.

cell biology↗

Annealing synchronizes the 70S ribosome into a minimum-energy conformation

Researchers commonly anneal metals, alloys, and semiconductors to repair defects and improve microstructures via recrystallization. Theoretical studies indicate simulated annealing on biological macromolecules helps predict the final structures with minimum free energy. Experimental validation of this homogenizing effect and further exploration of its applications are fascinating scientific questions that remain elusive. Here, we chose the apo-state 70S ribosome from Escherichia coli as a model, wherein the 30S subunit undergoes a thermally driven inter-subunit rotation and exhibits substantial structural flexibility as well as distinct free energy. We experimentally demonstrate that annealing at a fast cooling rate enhances the 70S ribosome homogeneity and improves local resolution on the 30S subunit. After annealing, the 70S ribosome is in a nonrotated state with respect to corresponding intermediate structures in unannealed or heated ribosomes, and exhibits a minimum energy in the free energy landscape. One can readily crystallize these minimum-energy ribosomes, which have great potential for synchronizing proteins on a single-molecule level. Our experimental results are consistent with theoretical analysis on the temperature-dependent Boltzmann distribution, and offer a facile yet robust approach to enhance protein stability, which is ideal for high-resolution cryogenic electron microscopy. Beyond structure determination, annealing can be extended to study protein folding and explore conformational and energy landscape. Significance statementIn metallurgy, annealing heats a metal or alloy to a predetermined temperature, holding for a certain time, and then cooling to room temperature to change the physical and sometimes also the chemical properties of the material. Researchers introduce the similar concept as simulated annealing to predict minimum-energy conformations of biological macromolecules. In this work, we experimentally verify that annealing at a fast cooling rate can synchronize the 70S ribosome into a nonrotated state with a minimum energy in the free energy landscape. Our results not only offer a facile yet robust approach to stabilize proteins for high-resolution structural analysis, but also contribute to the understanding of protein folding and temperature adaptation.

biophysics↗

A human antibody with blocking activity to RBD proteins of multiple SARS-CoV-2 variants including B.1.351 showed potent prophylactic and therapeutic efficacy against SARS-CoV-2 in rhesus macaques

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), which causes coronavirus disease-2019 (COVID-19), interacts with the host cell receptor angiotensin-converting enzyme 2 (hACE2) via its spike 1 protein for infection. After the virus sequence was published, we identified two potent antibodies against SARS-CoV-2 RBD from antibody libraries using a phage-to-yeast (PtY) display platform in only 10 days. Our lead antibody JMB2002, now in a phase I clinical trial, showed broad-spectrum in vitro blocking activity against hACE2 binding to the RBD of multiple SARS-CoV-2 variants including B.1.351 that was reportedly much more resistant to neutralization by convalescent plasma, vaccine sera and some clinical stage neutralizing antibodies. Furthermore, JMB2002 has demonstrated complete prophylactic and potent therapeutic efficacy in a rhesus macaque disease model. Prophylactic and therapeutic countermeasure intervention of SARS-CoV-2 using JMB2002 would likely slow down the transmission of currently emerged SARS-CoV-2 variants and result in more efficient control of the COVID-19 pandemic.

microbiology↗

Essential role of CFAP53 in sperm flagellum biogenesis

The sperm flagellum is essential for male fertility. Despite vigorous research progress towards understanding the pathogenesis of flagellum-related diseases, much remains unknown about the mechanisms underlying the flagellum biogenesis itself. Here, we show that the cilia and flagella associated protein 53 (Cfap53) gene is predominantly expressed in testes, and it is essential for sperm flagellum biogenesis. The knockout of this gene resulted in complete infertility in male mice but not in the females. CFAP53 localized to the manchette and sperm tail during spermiogenesis, the knockout of this gene impaired flagellum biogenesis. Furthermore, we identified two manchette and sperm tail-associated proteins that interacted with CFAP53 during spermiogenesis. The disruption of Cfap53 decreased the expression level of these two proteins and disrupted their localization in spermatids. Together, our results suggest that CFAP53 is an essential protein for sperm flagellum biogenesis, and its mutations might be associated with MMAF.

developmental biology↗

Single-cell transcriptome lineage tracing of human pancreatic development identifies distinct developmental trajectories of alpha and beta cells

In comparison to mouse, the developmental process of human islets has not been properly elucidated. The advancement of single cell RNA-seq technology enables us to study the properties of alpha and beta cells at single cell resolution. By using mitochondrial genome variants as endogenous lineage-tracing markers, we found that human alpha and beta cells have different lineage features. This finding suggests specific endocrine progenitors for alpha and beta cells, which is different from mouse islet cells. This strategy was also applied to a study of chemically-induced islet cell reprogramming and was used to help identify artemether-induced alpha-to-beta trans-differentiation in human islets. The computational results of this study will inspire future studies to establish, maintain, and expand beta cell-specific progenitors in vitro and in vivo.

developmental biology↗

Regulating enzymatic reactions in Escherichia coli utilizing light-responsive cellular compartments based on liquid-liquid phase separation

Enzymatic reactions in cells are well organized into different compartments, among which protein-based membraneless compartments formed through liquid-liquid phase separation (LLPS) are believed to play important roles1,2. Hijacking them for our own purpose has promising applications in metabolic engineering3. Yet, it is still hard to precisely and dynamically control target enzymatic reactions in those compartments4. To address those problems, we developed Photo-Activated Switch in E. coli (PhASE), based on phase separating scaffold proteins and optogenetic tools. In this system, a protein of interest (POI) can be enriched up to 15-fold by LLPS-based compartments from cytosol within only a few seconds once activated by light, and become fully dispersed again within 15 minutes. Furthermore, we explored the potentiality of the LLPS-based compartment in enriching small organic molecules directly via chemical-scaffold interaction. With enzymes and substrates co-localized under light induction, the overall reaction efficiency could be enhanced. Using luciferin and catechol oxidation as model enzymatic reactions, we found that they could accelerate 2.3-fold and 1.6-fold, respectively, when regulated by PhASE. We anticipate our system to be an extension of the synthetic biology toolkit, facilitating rapid recruitment and release of POIs, and reversible regulation of enzymatic reactions.

synthetic biology↗

PD-L1 lncRNA splice promotes lung adenocarcinoma progression via enhancing c-Myc activity

Although blockade of programmed death-ligand 1 (PD-L1) to enhance T cell immune responses shows great promise in tumor immunotherapy, the efficacy of such immune-checkpoint inhibition strategy is limited for patients with solid tumors. The mechanism underlying the limited efficacy of PD-L1 inhibitors remains unclear. Here, we show that human lung adenocarcinoma, regardless of PD-L1 protein positive or negative, all produce a long non-coding RNA isoform of PD-L1 (PD-L1-lnc) via alternative splicing, which promotes lung adenocarcinoma proliferation and metastasis. PD-L1-lnc in various lung adenocarcinoma cells is significantly upregulated by IFN{gamma} in a manner similar to PD-L1 mRNA. Both in vitro and in vivo studies demonstrate that PD-L1-lnc increases proliferation and invasion but decreases apoptosis of lung adenocarcinoma cells. Mechanistically, PD-L1-lnc directly binds to c-Myc and enhances c-Myc transcriptional activity downstream in lung adenocarcinoma cells. Our results provide targeting PD-L1-lnc-c-Myc axis as a novel strategy for lung cancer therapy.

cancer biology↗

Development of a simple in vitro assay to identify and evaluate nucleotide analogs against SARS-CoV-2 RNA-dependent RNA polymerase

Nucleotide analogs targeting viral RNA polymerase have been approved to be an effective strategy for antiviral treatment and are attracting antiviral drugs to combat the current SARS-CoV-2 pandemic. In this report, we develop a robust in vitro nonradioactive primer extension assay to evaluate the incorporation efficiency of nucleotide analog by SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) quantitively. Our results show that many nucleotide analogs can be incorporated into RNA by SARS-CoV-2 RdRp, and that the incorporation of some of them leads to chain termination. The discrimination values of nucleotide analog over those of natural nucleotide were measured to evaluate the incorporation efficiency of nucleotide analog by RdRp. We found that the incorporation efficiency of Remdesivir-TP is higher than ATP, and we did not observe chain termination or delayed chain termination caused by single Remdesivir-TP incorporation, while multiple incorporations of Remdesivir-TP caused chain termination in our assay condition. The incorporation efficiency of Ribavirin-TP and Favipiravir-TP is very low either as ATP or GTP analogs, which suggested that mutagenesis may not be the mechanism of action of those two drugs against SARS-CoV-2. Incorporation of Sofosbuvir-TP is also very low suggesting that sofosbuvir may not be very effective in treating SARS-CoV-2 infection. As a comparison, 2-C-Methyl-GTP can be incorporated into RNA efficiently, and the derivative of 2-C-Methyl-GTP may have therapeutic application in treating SARS-CoV-2 infection. This report provides a simple screening method that should be useful in evaluating nucleotide-based drugs targeting SARS-CoV-2 RdRp, and for studying the mechanism of action of selected nucleotide analog.

biochemistry↗

Fusarium verticillioides peroxins associated with peroxisomal docking/translocation module serve important roles in fumonisin biosynthesis and virulence

Peroxisomes are ubiquitous organelles in eukaryotic cells that fulfill various important metabolic functions. In this study, we investigated the role of Docking/Translocation Module (DTM) peroxins, mainly FvPex8, FvPex13, FvPex14, and FvPex33, in Fusarium verticillioides virulence and fumonisin B1 (FB1) biosynthesis. Protein interaction experiments suggested that FvPex13 serves as the core subunit of F. verticillioides DTM. When we generated gene deletion mutants ({Delta}Fvpex8, {Delta}Fvpex13, {Delta}Fvpex14, {Delta}Fvpex33, {Delta}Fvpex33/14) and examined whether the expression of other peroxin genes were affected in the DTM mutants, {Delta}Fvpex8 strain showed most drastic changes to PEX gene expression profiles. Deletion mutants exhibited disparity in carbon source utilization and defect in cell wall integrity when stress agents were applied. Under nutrient starvation, mutants also showed higher levels of lipid droplet accumulation. Notably, {Delta}Fvpex8 mutant showed significant FB1 reduction and altered expression of FUM1 and FUM19 genes. However, FvPex13 was primarily responsible for virulence, while {Delta}Fvpex33/14 double mutant also showed virulence defect. In summary, our study suggests that FvPex13 is the core component of DTM, regulating peroxisome membrane biogenesis as well as PTS1- and PTS2-mediated transmembrane cargo transportation. Importantly, we predict FvPex8 as a key component in DTM that affects peroxisome function in FB1 biosynthesis in F. verticillioides.

microbiology↗

Mico-Community associated with ectomycorrhizal Russula symbiosis and sporocarp-producing Russula in Fagaceae dominant nature areas in southern China

Russula griseocarnosa, an ectomycorrhizal (ECM) fungus, is a species of precious wild edible mushrooms with very high market value in southern China. Its yield is affected by many factors including the tree species and environmental conditions such as soil microbiome, humidity. How the microbiome promotes the ECM fungus symbiosis with Fagaceae plants and sporocarp-producing has never been studied. In this study, we collected rhizosphere samples from Fujian province, the microbiota in the root and mycorrhizal rhizosphere were identified by Illumina MiSeq high-throughput sequencing. First, we compared three types of fungal communities: root tips infected with ECM Russula (type 1), tips with Russula sporocarp (type 2) and tips without ECM (type 3). Our results showed that the fungal richness was negatively correlated with Russula. Russula, Tomentella and Lactarius were common in Fagaceae ECM roots. As to the mycorrhizal interactions, Boletus may be considered as an indicator species for sporocarp-producing Russula, and Acremonium, Cladophialophora were associated with Russula symbiosis. Second, we analyzed the fungal and bacterial communities in rhizosphere soils from the corresponding to previously three types (type 1, 2, 3). Dacryobolus and Acidocella may be considered as an indicator species for sporocarp-producing Russula. Fungi Tomentella, Saitozyma, Elaphomyces and bacteria Acidicaldus, Bryobacter, Sorangium and Acidobacterium occurred more frequently in the ECM Russula rhizosphere. Furthermore, the indicators Elaphomyces, Tomentella, Sorangium had a positive correlation with Russula symbiosis by network analyses. Overall, our results suggest a relationship between micro-community and ECM Russula formation and Russula sporocarp, which may provide new strategies for improving Russula symbiosis rate and sporocarp production.

ecology↗

Focused ultrasound stimulates ER localized mechanosensitive PANNEXIN-1 to mediate intracellular calcium release in invasive cancer cells

Focused ultrasound (FUS) is a rapidly developing stimulus technology with the potential to uncover novel mechanosensory dependent cellular processes. Since it is noninvasive, it holds great promise for future therapeutic applications in patients used either alone or as a complement to boost existing treatments. For example, FUS stimulation causes invasive but not noninvasive cancer cell lines to exhibit marked activation of calcium signaling pathways. Here, we identify the membrane channel PANNEXIN1 (PANX1) as a mediator for activation of calcium signaling in invasive cancer cells. Knockdown of PANX1 decreases calcium signaling in invasive cells, while PANX1 overexpression enhances calcium elevations in non-invasive cancer cells. We demonstrate that FUS may directly stimulate mechanosensory PANX1 localized in endoplasmic reticulum to evoke calcium release from internal stores. This process does not depend on mechanosensory stimulus transduction through an intact cytoskeleton and does not depend on plasma membrane localized PANX1. Plasma membrane localized PANX1 however plays a different role in mediating the spread of intercellular calcium waves via ATP release. Additionally, we show that FUS stimulation evokes cytokine/chemokine release from invasive cancer cells, suggesting that FUS could be an important new adjuvant treatment to improve cancer immunotherapy.

cell biology↗

qKW9 encodes a pentatricopeptide repeat protein affecting photosynthesis and grain filling in maize

Kernel weight is an important yield component in maize that was selected during domestication. Many kernel weight genes have been identified through mutant analysis, and are mostly involved in the biogenesis and functional maintenance of organelles or other fundamental cellular activities. However, only a limited number of loci underlying quantitative variation in kernel weight have been cloned. Here we characterize a maize kernel weight QTL, qKW9 and find that it encodes a DYW motif pentatricopeptide repeat protein involved in C-to-U editing of NdhB, a subunit of the chloroplast NADH dehydrogenase-like complex. In a null qKW9 background, C-to-U editing of NdhB was abolished, and photosynthesis was reduced, suggesting that qKW9 regulates kernel weight by controling the maternal source of photosynthate for grain filling. Characterization of qKW9 highlights the importance of optimizing photosynthesis on maize grain yield production.

plant biology↗

TET1-mediated Oligodendrocyte Homeostasis Regulates Myelination and Synaptic Functions

Ten-eleven translocation (TET) proteins, the dioxygenase for DNA hydroxymethylation, are important players in nervous system development and diseases. However, their role in myelination and remyelination after injury remains elusive. Here, we identify a genome-wide and locus-specific DNA hydroxymethylation landscape shift during differentiation of oligodendrocyte-progenitor cells (OPC). Ablation of Tet1 results in stage-dependent defects in oligodendrocyte (OL) development and myelination in the mouse brain. The mice lacking Tet1 in the oligodendrocyte lineage develop behavioral deficiency. We also show that TET1 is required for remyelination in adulthood. Transcriptomic, genomic occupancy, and DNA hydroxymethylation profiling reveal a critical TET1-regulated epigenetic program for oligodendrocyte differentiation that includes genes associated with myelination, cell division, and calcium transport. Tet1-deficient OPCs exhibit reduced calcium activity, increasing calcium activity rescues the differentiation defects in vitro. Deletion of a TET1-5hmC target gene, Itpr2 impairs the onset of OPC differentiation. Together, our results suggest that stage-specific TET1-mediated epigenetic programming and intracellular signaling are important for proper myelination and remyelination in mice.

neuroscience↗

Bayogenin 3-O-Cellobioside is a novel non-cultivar specific anti-blast metabolite produced in rice in response to Pyricularia oryzae infection

Rice cultivars from japonica and indica lineage possess differential resistance against blast fungus on an account genetic divergence. Whether different rice cultivars also show distinct metabolomic changes in response to P. oryzae, and their role in host resistance, are poorly understood. Here, we examine the responses of six different rice cultivars from japonica and indica lineage challenged with P. oryzae. Both susceptible and resistant rice cultivars expressed several metabolites exclusively during P. oryzae infection, including the saponin Bayogenin 3-O-cellobioside. Bayogenin 3-O-cellobioside level in infected rice directly correlated with their resistant attributes. These findings reveal, for the first time to our knowledge that besides oat, other grass plants including rice produces protective saponins. Our study provides insight into the role of pathogen-mediated metabolomics-reprogramming in host immunity. The correlation between Bayogenin 3-O-Cellobioside levels and blast resistance suggests that engineering saponin expression in cereal crops represents an attractive and sustainable disease control strategy.

biochemistry↗