Biology subjects

Gao, X.

Publications and source records attributed to Gao, X..

At least 217 records · Page 12Linked to original sources

bioRxiv2022

Differential expression patterns and potential regulatory function of piRNAs in western honey bee larval response to Ascosphaera apis infestation

piRNAs are a class of small non-coding RNAs that play an essential part in genomic defense, as well as in modulation of gene expression and diverse biological processes such as host-pathogen interaction. However, little is known about the expression pattern of the regulatory function of piRNAs in interactions between honey bees and pathogens. In this work, on the basis of our previously obtained high-quality small RNA-seq datasets from western honey bee (Apis mellifera) larval guts, the differential expression profile of piRNAs in A. mellifera larval guts after Ascosphaera apis infestation was analyzed, followed by structural characterization, target prediction, and regulatory network investigation. The potential roles of differentially expressed piRNAs (DEpiRNAs) in regulating host response, especially immune response, were further discussed. In this study, 504, 657, and 587 piRNAs were respectively identified in the 4-, 5-, and 6-day-old larval guts infected by A. apis, with 411 (53.24%) piRNAs shared. The length distribution of these piRNAs ranged from 24 nt to 33 nt and their first base had a "C" bias, similar to piRNAs discovered in other mammals and insects. Additionally, 96, 103, and 143 DEpiRNAs were detected in the 4-, 5-, and 6-day-old comparison groups; among these, piR-ame-149736, piR-ame-1066173, and piR-ame-1125190 were the most up-regulated, while piR-ame-1202932 was the most down-regulated. There were 68 DEpiRNAs shared between these three comparison groups. The targets of the DEpiRNAs in the three comparison groups were engaged in a suite of crucial functions associated with biological processes, molecular function, and cellular components, including molecular transducer activity, biological regulation, and membrane part. These targets were also relevant to diverse vital pathways such as the phosphatidylinositol signaling system, inositol phosphate metabolism, and Wnt signaling pathway. Further investigation demonstrated that targets of DEpiRNAs were involved in three energy metabolism-related pathways, seven development-associated signaling pathways, and seven immune-relevant pathways, including lysosome and endocytosis, as well as the MAPK and Jak-STAT signaling pathways. The expression trends of five randomly selected DEpiRNAs were verified using a combination of RT-PCR and RT-qPCR. Moreover, the expression levels of six genes targeted by piR-ame-945760 were detected by RT-qPCR, with the results showing that their expression trends were the same as the expression trend of piR-ame-945760, indicative of the positive correlation between piR-ame-945760 and these targets. These results suggest that A.apis infestation increased the overall expression level of piRNAs and altered the expression pattern of piRNAs in A. mellifera larval guts. DEpiRNAs potentially participate in the A. apis response of the host by modulating the expression of target genes associated with energy metabolism and development, as well as cellular and humoral immune response. Our findings not only offer novel insights into A. mellifera larva-A. apis interaction, but also lay the groundwork for clarifying the mechanisms underlying DEpiRNA-regulated larval response.

molecular biology↗

bioRxiv2022

Effect of Ascosphaera apis infestation on the activities of four antioxidant enzymes in Asian honey bee larval guts

Ascosphaera apis exclusively infects bee larvae and causes chalkbrood, a lethal fungal disease that results in the sharp reduction in adult bees and colony productivity. However, little is known about the effect of A. apis infestation on the activities of antioxidant enzymes in bee larvae. Here, A. apis spores were purified and used to inoculate Asian honey bee (Apis cerana) larvae, followed by detection of the host survival rate and evaluation of the activities of four major antioxidant enzymes. At 6 days post inoculation (dpi) with A. apis spores, white mycelia penetrated the posterior end of the larva, extended to the anterior end, and eventually covered the entire larval body surface, presenting an obvious symptom of chalkbrood disease similar to that occurs in Apis mellifera larvae. Additionally, PCR identification showed that the expected fragment was amplified from the A. apis-inoculated larval guts and the A. apis spores, verifying the A. apis infection of A. cerana larvae. The survival rate of larvae inoculated with A. apis was high at 1-2 dpi, sharply decreased to 4.16% at 4 dpi, and reached 0% at 5 dpi; whereas that of un-inoculated larvae was always high at 1~8 dpi, with an average survival rate of 95.37%, indicating the negative impact of A. apis infection on larval survival. Furthermore, in comparison with those in the corresponding un-inoculated groups, the superoxide dismutase (SOD) and catalase (CAT) activities in the 4-day-old larval gut in the A. apis-inoculated groups were reduced (p > 0.05), while those in the 5- and 6-day-old larval guts were significantly decreased (p < 0.05); the glutathione S-transferase (GST) activity in the 4- and 5-day-old larval guts was significantly increased (p < 0.05), while that in the 6-day-old larval gut was reduced (p > 0.05); the polyphenol oxidase (PPO) activity in 4-day-old larval gut was increased (p > 0.05) and that in the 5-day-old larval gut was significantly increased (p < 0.05), whereas that in the 6-day-old larval gut was significantly reduced (p < 0.01). These results together suggested that the activities of SOD and CAT in the larval guts were suppressed during the process of A. apis infestation, while the GST activity was induced to activation, and the PPO activity was first enhanced and then inhibited. Our findings not only unravel the response of A. cerana larvae to A. apis infestation from a biochemical perspective, but also offer a valuable insight into the interaction between Asian honey bee larvae and A. apis.

molecular biology↗

bioRxiv2022

The RNA m6A reader YTHDF1 promotes hematopoietic malignancy by maintaining oncogenic translation

N6-methyladenosine (m6A), the most abundant modification in mRNAs, has been defined as a crucial modulator in the progression of acute myeloid leukemia (AML), while the detailed mechanism remains elusive. Here we report that YTHDF1, an m6A reader protein, is overexpressed in human AML samples with enrichment in leukemia stem cells (LSCs). Whereas YTHDF1 is dispensable for normal hematopoiesis in mice, depletion of YTHDF1 attenuates self-renewal and proliferation of patient-derived LSCs, and impedes leukemia establishment in immunodeficient mice. Mechanistically, YTHDF1 promotes the translation of diverse m6A-modified oncogene mRNAs particularly cyclin E2. We applied a structure-based virtual screening of FDA-approved drugs and identified tegaserod as a potential YTHDF1 inhibitor. Tegaserod blocks the direct binding of YTHDF1 with m6A-modified mRNAs and inhibits YTHDF1-regulated mRNA translation. Moreover, tegaserod inhibits leukemogenesis in vitro and in mice, phenocopying the loss of YTHDF1. Together, our study defines YTHDF1 as an integral regulator of AML progression at the translational level and identifies tegaserod as a potential therapeutic agent for AML by targeting YTHDF1. Statement of SignificanceO_LIThe RNA m6A reader YTHDF1 is an oncoprotein in AML. C_LIO_LIYTHDF1 promotes the translation of m6A-modified oncogene mRNAs. C_LIO_LITegaserod, an FDA-approved drug for irritable bowel syndrome, inhibits YTHDF1-regulated mRNA translation and leukemogenesis. C_LI Graphical Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

cancer biology↗

bioRxiv2022

Flaver: mining transcription factors in genome-wide transcriptome profiling data using weighted rank correlation statistics

BackgroundMining key transcription factors (TFs) in genome-wide transcriptome profiling data has been an active research area for many years and it has been partially solved by mathematically modelling the ranking orders of genes in the target gene-set for the TF of interest in the gene-list ranked by expression values, called gene-set enrichment analysis (GSEA). However, in some application scenarios the gene-set itself also has a rank attribute, such as the putative target gene-set predicted by the Grit software and other alternatives like FIMO and Pscan. New algorithms must be developed to analyze these data properly. Methodology/Principal FindingsBy implementing the weighted Kendalls tau statistic, we proposed a method for genome-wide transcriptome profiling data mining that can identify the key TFs orchestrating a profile. Theoretical properties of the proposed method were established, and its advantages over the GSEA approach were demonstrated when analyzing the RNA-Atlas datasets. The results showed that the top-rated TFs by our method always have experimentally supported evidences in the literatures. Benchmarking using gene ontology (GO) annotations in the AmiGO database indicated that the geometry performance (SQR_P) of our method is higher than GSEA in more than 14% of the cases. SignificanceThe developed method is suitable for analyzing the significance of overrepresentation of ranked gene-sets in a ranked gene-list. A software implementing the method, called "Flaver", was developed and is publicly available at http://www.thua45.cn/flaver under an academic free license. Author SummaryIdentification of the regulation roles of TFs in the transcriptome is fundamental in understanding various biological processes. Improve the performance of the prediction tools is important because accurate TF-mining in transcriptome data can finely improve the efficiency of wet-lab experiments. Also, genome wide TF-mining can provide new target genes for transcriptome regulation analysis in system biology perspective. This study developed a new TF-mining tool based on weighted rank correlation statistical method. The tool has better performance in analyzing ranked gene-set and ranked gene-list than its competitor, the GSEA tool. It can help the researchers in identification of the most important TFs in transcriptional data.

bioinformatics↗

bioRxiv2022

Structure of human phagocyte NADPH oxidase in the resting state

Phagocyte oxidase plays an essential role in the first line of host defense against pathogens. It oxidizes intracellular NADPH to reduce extracellular oxygen to produce superoxide anions for pathogen killing. The resting phagocyte oxidase is a heterodimeric complex formed by two transmembrane proteins NOX2 and p22. Despite the functional importance of this complex, its structure remains elusive. Here we reported the cryo-EM structure of the human NOX2-p22 complex in nanodisc in the resting state. The structure shows that p22 is formed by four transmembrane helices and interacts with NOX2 through its M1 and M4 helices. Hydrophobic residues on M3, M4, and M5 of NOX2 contribute to the complex formation. Structural analysis suggests that the cytosolic factors activate the NOX2-p22 complex by stabilizing the dehydrogenase domain (DH) in a productive docked conformation which is efficient for electron transfer between DH and the transmembrane domain.

biochemistry↗

bioRxiv2022

Dual control of lysogeny and phage defense by a phosphorylation-based toxin/antitoxin system

Regulatory systems that maintain prophage quiescence integrate phage and host gene expression with environmental conditions1,2. In the opportunistic bacterial pathogen Pseudomonas aeruginosa, Pf filamentous bacteriophages play critical roles in biofilm formation and virulence3-5, but mechanisms governing Pf prophage activation in biofilms are largely unknown. Here, we report a new type of prophage regulatory module in a widely-distributed P. aeruginosa lineage that not only controls virion production of co-resident Pf prophages, but also mediates defense against diverse lytic phages. By comparing two lineages of the prototype P. aeruginosa strain PAO1 that harbor different Pf prophages, we identified a prophage-encoded kinase-kinase-phosphatase (KKP) system that controls Pf production in biofilms. KKP components exhibit dynamic stoichiometry, where high kinase levels in planktonic conditions maintain phosphorylation of the host H-NS protein MvaU, repressing prophage activation. During biofilm formation, phosphatase expression is heightened, leading to MvaU dephosphorylation and alleviating repression of prophage gene expression. KKP clusters are present in hundreds of diverse temperate prophages and other mobile elements across Gram-negative bacteria. Characterization of KKP modules from different species revealed that, in addition to regulating Pf phage lysogeny, KKP functions as a tripartite toxin-antitoxin system that mediates host defense from predatory lytic phages. KKP represents a new phosphorylation-based mechanism for prophage regulation and for phage defense. The dual function of this module raises the question of whether other newly described phage defense systems6-9 also regulate intrinsic prophage biology in diverse hosts.

microbiology↗

bioRxiv2022

ZEB2 regulates the development of CD11c+ atypical B cells

CD11c+ atypical B cells (ABC) are an alternative memory B cell lineage identified both in normal immune responses as well as pathogenic responses in autoimmunity. While it is clear that ABCs have a distinct transcriptional program, the factors that direct this program have not been identified. Here, we generated a human tonsil single-cell RNA-seq dataset and identified candidate transcription factors associated with the ABC population. We selected 8 of these transcription factors for further analysis based on their conserved expression in mouse ABC bulk RNA-seq datasets. Using an optimized CRSPR-Cas9 knockdown method we found that only zinc finger E-box binding homeobox 2 (Zeb2) knock-out impaired ABC formation. To assess the role of Zeb2 in ABC formation in vivo we used Zeb2fl/fl mice crossed to a CD23Cre line. Germinal center and plasma cell responses in these mice after Plasmodium sporozoite immunization were largely unaltered but we observed a specific defect in ABC formation. We further determined that ZEB2 haploinsufficient Mowat Wilson syndrome patients also have decreased circulating ABCs in the blood, supporting a role for this transcription factor in humans as well as mice. In sum, we identified Zeb2 as a key TF governing the formation of ABCs.

immunology↗

bioRxiv2022

Defective Ca2+- dependent activation of TRPM4 channels contributes to age-related cerebral small vessel disease in Col4a1 mutant mice

Humans and mice with mutations in COL4A1 and COL4A2 manifest hallmarks of cerebral small vessel disease (cSVD), but the pathogenic mechanisms are largely unknown. Mice with a missense mutation in Col4a1 at amino acid 1344 (Col4a1+/G1344D) exhibited age-dependent intracerebral hemorrhage (ICH) and brain lesions. Here we report that this pathology was associated with the loss of myogenic vasoconstriction, an intrinsic vascular response essential for the autoregulation of cerebral blood flow. Electrophysiological analyses showed that the loss of myogenic constriction resulted from blunted pressure-induced smooth muscle cell (SMC) membrane depolarization. Further, we found that dysregulation of membrane potential was associated with impaired Ca2+-dependent activation of large-conductance Ca2+-activated K+ (BK) and transient receptor potential melastatin 4 (TRPM4) cation channels linked to disruptions in sarcoplasmic reticulum (SR) Ca2+ signaling. Treating Col4a1+/G1344D mice with 4-phenylbutyrate, a compound that promotes the trafficking of misfolded proteins and alleviates SR stress, restored SR Ca2+ signaling, BK and TRPM4 channel activity, prevented loss of myogenic tone, and reduced ICH. We conclude that alterations in SR Ca2+ handling that impair membrane potential regulating ion channel activity result in dysregulation of SMC membrane potential and loss of myogenic tone contributing to age-related cSVD in Col4a1+/G1344D mice. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=85 SRC="FIGDIR/small/505186v3_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@11a8cb9org.highwire.dtl.DTLVardef@d00b8org.highwire.dtl.DTLVardef@1aa5b24org.highwire.dtl.DTLVardef@145a9c0_HPS_FORMAT_FIGEXP M_FIG C_FIG

pathology↗

bioRxiv2022

Targeting Epsins by nanotherapy regulates lipid metabolism and promotes 1 ABCG1-mediated cholesterol efflux to fortify atheroma regression

BACKGROUNDExcess cholesterol accumulation in lesional macrophages elicits complex responses in atherosclerosis. Epsins, a family of endocytic adaptors, fuel the progression of atherosclerosis; however, the underlying mechanism and therapeutic potential of targeting Epsins remains unknown. In this study, we determined the role of Epsins in macrophage-mediated metabolic regulation. We then developed an innovative method to therapeutically-target macrophage Epsins with specially-designed S2P-conjugated lipid nanoparticles (NPs), which encapsulate small interfering RNAs to suppress Epsins. METHODSWe used single cell RNA sequencing (scRNA-seq) with our newly developed algorithm MEBOCOST to study cell-cell communications mediated by metabolites from sender cells and sensor proteins on receiver cells. Biomedical, cellular and molecular approaches were utilized to investigate the role of macrophage Epsins in regulating lipid metabolism and transport. We performed this study using myeloid-specific Epsin double knockout (LysM-DKO) mice and mice with a genetic reduction of ABCG1 (LysM-DKO-ABCG1fl/+). The NPs targeting lesional macrophages were developed to encapsulate interfering RNAs to treat atherosclerosis. RESULTSWe revealed that Epsins regulate lipid metabolism and transport in atherosclerotic macrophages. Inhibiting Epsins by nanotherapy halts inflammation and accelerates atheroma resolution. Harnessing lesional macrophage-specific NP delivery of Epsin siRNAs, we showed that silencing of macrophage Epsins markedly diminished atherosclerotic plaque size and promoted plaque regression. Mechanistically, we demonstrated that Epsins bound to CD36 to facilitate lipid uptake by enhancing CD36 endocytosis and recycling. Conversely, Epsins promoted ABCG1 degradation via lysosomes and hampered ABCG1-mediated cholesterol efflux and reverse cholesterol transport. In a LysM-DKO-ABCG1fl/+ mouse model, enhanced cholesterol efflux and reverse transport due to Epsin deficiency was suppressed by the reduction of ABCG1. CONCLUSIONSOur findings suggest that targeting Epsins in lesional macrophages may offer therapeutic benefits for advanced atherosclerosis by reducing CD36-mediated lipid uptake and increasing ABCG1-mediated cholesterol efflux. Novelty and SignificanceO_ST_ABSWHAT IS KNOWN?C_ST_ABSO_LIEpsin endocytic adaptor proteins are upregulated in human and mouse atherosclerotic lesions C_LIO_LILesional macrophages internalize lipids primarily through scavenger receptor-mediated endocytosis such as CD36 and SR-A C_LIO_LIMacrophage-mediated cholesterol efflux and reverse cholesterol transport is crucial to atheroma resolution C_LI WHAT NEW INFORMATION DOES THIS ARTICLE CONTRIBUTE?O_LIScRNA-seq combined with the newly-developed algorithm MEBOCOST reveals that Epsins are involved in macrophage-mediated lipid metabolic regulation C_LIO_LIMacrophage epsins promote lipid uptake by targeting CD36 endocytosis and membrane recycling via the Epsin ENTH domain C_LIO_LIEpsins bind ubiquitinated ABCG1--resulting in the endocytosis and lysosomal degradation of this cholesterol transporter, which reduces cholesterol efflux. C_LIO_LIMacrophage-specific, nanoparticle-mediated RNAi delivery exhibits a therapeutic benefit for the treatment of atherosclerosis C_LIO_LIAtherosclerotic plaque regression using this nanoparticle delivery platform represents a clinically-relevant approach for the treatment of advance atherosclerosis C_LI BRIEF SUMMARYDespite the development of new cholesterol-lowering therapies, including the recently approved PCSK9 small interfering RNA (siRNA) antagonists, patients still face a tremendous risk of developing major acute cardiovascular events resulting from chronic inflammation in the plaque. We employed a novel nanomedicine platform containing a stabilin-2 targeting peptide (S2P) to deliver Epsin-specific siRNAs to lesional macrophages. We discovered that inhibition of these adaptor proteins in lesional macrophages significantly diminished plaque size and necrotic core area, increased fibrous cap thickness, and promoted plaque regression.

cell biology↗

bioRxiv2022

Rapid Color Categorization in the Brain Revealed by Frequency-tagging-based EEG

The origin of color categories has been debated extensively. Historically, linguistic relativists claim that color categories are shaped by the language we speak and that color terms subsequently affect our perception of color, while universalists postulate that color categories are independent of language and formed based on perceptual mechanisms. A recent hypothesis suggests that the original fine-grained color space in the visual cortex may be transformed into categorical encoding due to top-down modulation. To test the nature of color categorization, our study adopted a sensitive frequency-tagging-based EEG paradigm where the color stimuli were presented sequentially at a fast speed of 10 Hz (SOA: 100 ms) to probe fast, implicit processing of color categories. This SOA was supposed to disrupt top-down feedbacks in visual processing. We showed that EEG responses to cross-category oddball colors at the frequency where the oddball stimuli were presented was significantly larger than the responses to within-category oddball colors. This finding suggested that the brain encodes color categories automatically when top-down feedbacks from frontoparietal areas are blocked. Our study supports the view that the categorical processing of color emerges at the early perceptual stage.

neuroscience↗

bioRxiv2022

Type 17 Follicular Helper T (Tfh17) Cells are Superior for Memory Maintenance

A defining feature of successful vaccination is the ability to induce long-lived antigen- specific memory cells. Follicular helper T (Tfh) cells specialize in providing help to B cells in mounting protective humoral immunity in infection and after vaccination. Memory Tfh cells that retain the CXCR5 expression can confer protection through enhancing humoral response upon antigen re-exposure but how they are maintained is poorly understood. CXCR5+ memory Tfh cells in human blood are divided into Tfh1, Tfh2 and Tfh17 cells by the expression of chemokine receptors CXCR3 and CCR6 associated with Th1 and Th17 respectively. Here, we developed a new method to induce Tfh1, Tfh2 and Tfh17-like (iTfh1, iTfh2 and iTfh17) cells in vitro. Although all three iTfh subsets efficiently support antibody responses in recipient mice with immediate immunization, iTfh17 cells are superior to iTfh1 and iTfh2 cells in supporting antibody response to a later immunization after extended resting in vivo to mimic memory maintenance. Notably, the counterpart human Tfh17 cells are selectively enriched in CCR7+ central memory Tfh (TfhCM) with survival and proliferative advantages. Furthermore, the analysis of multiple human cohorts that received different vaccines for HBV, influenza virus, tetanus toxin or measles revealed that vaccine-specific Tfh17 cells outcompete Tfh1 or Tfh2 cells for the persistence in memory phase. Therefore, the complementary mouse and human results showing the advantage of Tfh17 cells in maintenance and memory function supports the notion that Tfh17-induced immunization might be preferable in vaccine development to confer long-term protection.

immunology↗

bioRxiv2022

Utilizing a Cell-free Protein Synthesis Platform for Natural Product Synthesis

Natural products are a great source of pharmaceuticals, providing a majority of all small molecule drugs that exist today. However, creating natural products through organic synthesis or in heterologous hosts can be difficult and time-consuming. Therefore, to allow for easier screening and production of natural products, we demonstrated the use of a cell-free protein synthesis (CFPS) system to partially assemble natural products in vitro using coupled enzyme reactions. The tea caffeine synthase TCS1 was utilized to synthesize caffeine within a CFPS system. Cell-free systems also provide the benefit of allowing the use of substrates that would normally be toxic in a cellular environment to synthesize novel products. The automation and reduced metabolic engineering requirements of CFPS systems combined with other synthesis methods can allow for the efficient generation of new compounds.

synthetic biology↗

bioRxiv2022

Erythrocyte signalling is critical for Plasmodium falciparum invasion

Successful Plasmodium falciparum merozoite invasion requires the activation of red blood cell (RBC) signalling pathways. The binding of parasite ligand reticulocyte binding protein homologue 5 (RH5) to its host receptor Basigin is essential for merozoite invasion and triggers a Ca2+ influx in RBCs. Here we observed that RH5-bound RBCs form a multimeric protein complex containing Basigin, CD44 and {beta}2-adrenergic receptor ({beta}2AR), suggesting that RH5-Basigin interaction is functionally associated with the host cAMP signalling pathway. Interestingly, we detected a characteristic rise in cAMP levels in the RBC upon RH5-Basigin interaction, which can be blocked by G protein and cAMP-synthesising adenylyl cyclase (AC) inhibitors. Furthermore, we demonstrated that RBC L-type Ca2+ channel inhibitor and cAMP signalling inhibitors are able to block merozoite invasion. Checkerboard invasion inhibition assay containing different combinations of signalling inhibitors also exhibited a drastic amplification of inhibition levels, indicating that these signalling proteins are functioning in a common signalling cascade to activate the L-type Ca2+ channels. Taken together, this study provides new insights into the role of a host cAMP-Ca2+ signalling pathway during merozoite invasion and sheds new light on antimalarial therapeutic strategies to tackle the high infection rate and growing threat of drug resistant parasites. Key PointsO_LIA pre-existing Basigin-associated membrane protein complex undergoes increased protein assembly upon RH5 binding on the RBC surface. C_LIO_LIPlasmodium falciparum merozoite exploits host cAMP signalling to initiate Ca2+ influx in the RBC. C_LI

microbiology↗

bioRxiv2022

Adrenal sympathetic nerve mediated the anti-inflammatory effect of electroacupuncture at ST25 acupoint on a rat model of sepsis

Acupuncture plays a vital anti-inflammatory action on sepsis through activating autonomic nerve anti-inflammatory pathways, such as sympathoadrenal medullary pathway, but the mechanism remains unclear. This study aims to explore the optimum parameter of electroacupuncture (EA) stimulation in regulating sympathoadrenal medullary pathway and evaluate EAs anti-inflammatory effect on sepsis. To explore the optimum parameter of EA at homosegmental acupoint on adrenal sympathetic activity, the left adrenal sympathetic nerve firing rate evoked by different intensities of single shock electrical stimulation (ES) at ST25 in healthy male Sprague-Dawley (SD) rats were evaluated by in vivo electrophysiological recording, and the levels of norepinephrine (NE) and its metabolites were also examined using mass spectrometry. To verify the role of EA at ST25 in sepsis, the rat was given intraperitoneal injection lipopolysaccharide to induce sepsis model, and survival rate, clinical score, and the level of interleukin (IL)-6, IL-1{beta}, and IL-10 were evaluated after EA application. We observed that 3 mA is the optimal intensity on activating adrenal sympathetic nerve, which significantly elevated the level of NE in the peripheral blood. For LPS-treated rats, EA at the ST25 apparently increased the survival rate and improved the clinical score compared to the control group. Furthermore, 3 mA EA at ST25 significantly decreased pro-inflammatory cytokines IL-6 and IL-1{beta} and upregulated anti-inflammatory cytokine IL-10 compared to the Lipopolysaccharide (LPS)-treated group. Overall, these data suggest that 3 mA is the optimal EA intensity at ST25 to activate the sympathoadrenal medullary pathway and exert an anti-inflammatory effect on sepsis. HighlightsO_LI3 mA single shock ES at ST25 drove adrenal sympathetic nerve reflexs. C_LIO_LI3 mA EA at ST25 upregulated NE concentration in peripheral blood. C_LIO_LIEA at ST25 improved the survival rate in a rat of LPS-induced sepsis model. C_LIO_LIEA at the ST25 upregulated anti-inflammatory IL10 and downregulated pro-inflammatory IL1{beta} and IL-6 in LPS-treated rats. C_LI

neuroscience↗

bioRxiv2022

Regulation of sleep quantity and intensity by long and short isoforms of SLEEPY kinase

In Sleepy (Sik3Slp) or Sik3S551A mice, deletion or mutation of inhibitory phosphorylation site serine551 (S551) from salt-inducible kinase 3 (SIK3) markedly increases daily non-rapid eye movement sleep (NREMS) amount, accompanied with constitutively elevated NREMS delta power density-a measure of sleep intensity or quality. Multiple SLP/SIK3 isoforms are expressed in mouse brain neurons, however, their respective roles in sleep regulation remain to be elucidated. Here, we identified a new and most abundant short isoform of SLP/SIK3 and examined sleep phenotypes resulted from isoform-specific expression of SLP-short (S) and long (L) isoforms. Adeno-associated virus-mediated adult brain chimeric (ABC)-expression of SLP-S in neurons, but not in astrocytes, significantly and constitutively elevates NREMS delta power, whereas slightly increases NREMS amount. The ability of SLP-S to regulate sleep quantity/intensity is abrogated by kinase-inactivating mutations, suggesting that the sleep-promoting activity of SLP-S is dependent on its kinase activity. In Sik3S551A-L knock-in mice, isoform-specific expression of SIK3S551A-L (or SLP-L) significantly increases NREMS amount with a modest effect on NREMS delta power. ABC-expression of SLP-S complements the sleep phenotypes of heterozygous Sik3S551A-L mice by further increasing NREMS amount and NREMS delta power to levels of Sik3Slp or Sik3S551A mice. Taken together, these results indicate that both SLP-L and SLP-S isoforms contribute critically to the increases of sleep quantity and intensity in Sik3Slp or Sik3S551A mice. Statement of SignificancePrevious studies have identified SIK3 as a key sleep regulatory kinase, of which gain-of-function mutations markedly increase sleep quantity and intensity in Sleepy mice. Multiple SLEEPY isoforms are expressed in mouse brain neurons, but their respective contributions to the hypersomnia phenotypes of Sleepy mice remain unclear. Here, we identified a new short isoform of SLP/SIK3 and examined sleep phenotypes resulted from specific expression of SLEEPY-short or long isoform. Our results indicate that both long and short isoforms of SLEEPY kinase contribute critically to increases of sleep quantity and intensity in Sleepy mice. Future studies are needed to identify downstream substrates of SLEEPY kinase that mediate regulation of sleep quantity/intensity, which should facilitate development of new treatments for human sleep disorders.

genetics↗

bioRxiv2022

Functional characterization of multidomain protein Vip3Aa from Bacillus thuringiensis reveals a strategy to increase its insecticidal potency

Microbially derived, protein-based biopesticides offer a more sustainable pest management alternative to synthetic pesticides. Vegetative insecticidal proteins (Vip3), multidomain proteins secreted by Bacillus thuringiensis, represent a second-generation insecticidal toxin that have been used in transgenic crops. However, the molecular mechanism underlying Vip3s toxicity is poorly understood. Here, we determine the distinct functions and contributions of the domains of the Vip3Aa protein to its toxicity against Spodoptera frugiperda larvae. Vip3Aa domains II and III (DII-DIII) bind the midgut epithelium, while DI maintains the tetrameric state of the Vip3Aa protoxin, which is essential for its stability and toxicity. DI-DIII can be activated by midgut proteases, and exhibits ex vivo cytotoxicity similar to full-length Vip3Aa. We also determine that DV binds the peritrophic matrix via its glycan-binding activity, which is essential for Vip3Aa insecticidal activity. We further show that Vip3Aa has multiple protease activation sites and that introducing additional cleavage sites between DI and DII can increase the proteolysis efficiency and boost Vip3Aa insecticidal potency. This study provides insights into Vip3Aas mode-of-action and demonstrates a proof-of-concept strategy to enhance the insecticidal potency of Vip3Aa, which should significantly improve its application and development as a biopesticide.

microbiology↗

bioRxiv2022

Kynurenic acid promotes A1 to A2 astrocyte conversion against HIV-associated neurocognitive disorder

Although extensive astrocyte activation is present in the CNS of patients suffering from HIV-associated neurocognitive disorder (HAND), little is known about the contribution of reactive astrocytes to HAND neuropathology. Here, we report that the imbalance between neurotoxic (A1 astrocytes) and protective astrocytes (A2 astrocytes) occurring in HIV-1 gp120 transgenic mice impedes the resolution of inflammation in the CNS, perpetuating neuron damage and cognitive deficits. Kynurenic acid (KYNA), a tryptophan metabolite with 7 nicotinic acetylcholine receptors (7nAChR) inhibitory property, blunts gp120-induced A1 astrocyte activation while promoting A2 astrocyte formation through blockade of 7nAChR/JAK2/STAT3 signaling activation. Meanwhile, we provide evidence that 7nAChR deletion and tryptophan administration convert astrocyte phenotypes, ultimately facilitating neuronal and cognitive improvement in the gp120tg mice. These initial and determinant findings mark a turning point in our understanding of the role of 7nAChR in gp120-mediated astrocyte activation, which opens new opportunities to control reactive astrocyte generation through KYNA and tryptophan administration as a pharmacological approach for treating HAND.

microbiology↗

bioRxiv2022

Predicting the antigenic evolution of SARS-COV-2 with deep learning

The severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) antigenic profile evolves in response to the vaccine and natural infection-derived immune pressure, resulting in immune escape and threatening public health. Exploring the possible antigenic evolutionary potentials improves public health preparedness, but it is limited by the lack of experimental assays as the sequence space is exponentially large. Here we introduce the Machine Learning-guided Antigenic Evolution Prediction (MLAEP), which combines structure modeling, multi-task learning, and genetic algorithm to model the viral fitness landscape and explore the antigenic evolution via in silico directed evolution. As demonstrated by existing SARS-COV-2 variants, MLAEP can infer the order of variants along antigenic evolutionary trajectories, which is also strongly correlated with their sampling time. The novel mutations predicted by MLAEP are also found in immunocompromised covid patients and newly emerging variants, like XBB1.5. The predictions of MLAEP were validated by conducting in vitro neutralizing antibody binding assay, which demonstrated that the model-generated variants displayed significantly increased immune evasion ability compared with the controls. In sum, our approach enables profiling existing variants and forecasting prospective antigenic variants, thus may help guide the development of vaccines and increase preparedness against future variants. Our model is available at https://mlaep.cbrc.kaust.edu.sa.

evolutionary biology↗