Biology subjects

Shi, Y.

Publications and source records attributed to Shi, Y..

At least 199 records · Page 11Linked to original sources

bioRxiv2024

Structural Dynamics of the Ubiquitin Specific Protease USP30 in Complex with a Cyanopyrrolidine-Containing Covalent Inhibitor

Inhibition of the mitochondrial deubiquitinating enzyme USP30 is neuroprotective and presents therapeutic opportunities for the treatment of idiopathic Parkinsons Disease and mitophagy-related disorders. We have integrated structural and quantitative proteomics with biochemical assays to decipher the mode of action of covalent USP30 inhibition by a small molecule containing a cyanopyrrolidine reactive group, USP30-I-1. The inhibitor demonstrated high potency and selectivity for endogenous USP30 in neuroblastoma cells. Enzyme kinetics and Hydrogen Deuterium eXchange mass spectrometry (HDX-MS) infers that the inhibitor binds tightly to regions surrounding the USP30 catalytic cysteine and positions itself to form a binding pocket along the thumb and palm domains of the protein, thereby interfering its interaction with ubiquitin substrates. A comparison to a non-covalent USP30 inhibitor containing a benzosulfonamide scaffold revealed a slightly different binding mode closer to the active site Cys77, which may provide the molecular basis for improved selectivity towards USP30 against other members of the DUB enzyme family. Our results highlight advantages in developing covalent inhibitors, such as USP30-I-1, for targeting USP30 as treatment of disorders with impaired mitophagy.

biochemistry↗

bioRxiv2024

Biophysical Modeling and Experimental Analysis of the Dynamics of C. elegans Body-Wall Muscle Cells

This study combines experimental techniques and mathematical modeling to investigate the dynamics of C. elegans body-wall muscle cells. Specifically, by conducting voltage clamp and mutant experiments, we identify key ion channels, particularly the L-type voltage-gated calcium channel (EGL-19) and potassium channels (SHK-1, SLO-2), which are crucial for generating action potentials. We develop Hodgkin-Huxley-based models for these channels and integrate them to capture the cells electrical activity. To ensure the model accurately reflects cellular responses under depolarizing currents, we develop a parallel simulation-based inference method for determining the models free parameters. This method performs rapid parallel sampling across high-dimensional parameter spaces, fitting the model to the responses of muscle cells to specific stimuli and yielding accurate parameter estimates. We validate our model by comparing its predictions against cellular responses to various current stimuli in experiments and show that our approach effectively determines suitable parameters for accurately modeling the dynamics in mutant cases. Additionally, we discover an optimal response frequency in body-wall muscle cells, which corresponds to a burst firing mode rather than regular firing mode. Our work provides the first experimentally constrained and biophysically detailed muscle cell model of C. elegans, and our analytical framework combined with robust and efficient parametric estimation method can be extended to model construction in other species. Author summaryDespite the availability of many biophysical neuron models of C. elegans, a biologically detailed model of its muscle cell remains lacking, which hampers an integrated understanding of the motion control process. We conduct voltage clamp and mutant experiments to identify ion channels that influence the dynamics of body-wall muscle cells. Using these data, we establish Hodgkin-Huxley-based models for these ion channels and integrate them to simulate the electrical activity of the muscle cells. To determine the free parameters of the model, we develop a simulation-based inference method with parallel sampling that aligns the model with the muscle cells responses to specific stimuli. Our method allows for swift parallel sampling of parameters in high dimensions, facilitating efficient and accurate parameter estimation. To validate the effectiveness of the determined parameters, we verify the cells responses under different current stimuli in wild type and mutant cases. Furthermore, we investigate the optimal response frequency of body-wall muscle cells and find that it exhibits a frequency consistent with burst firing mode rather than regular firing mode. Our research introduces the first experimentally validated and biophysically detailed model of muscle cells in C. elegans. Additionally, our modeling and simulation framework for efficient parametric estimation in high-dimensional dynamical systems can be extended to model constructions in other scenarios.

neuroscience↗

bioRxiv2024

Systematically investigating and identifying unmodified bacteriocins in the human gut microbiome

Human gut microbiota produces unmodified bacteriocins, natural antimicrobial peptides that protect against pathogens and regulate host physiology. However, current bioinformatic tools limit the comprehensive investigation of bacteriocins biosynthesis, obstructing research into their biological functions. Here, we introduce IIBacFinder, a superior analysis pipeline for identifying unmodified class II bacteriocins. Through large-scale bioinformatic analysis and experimental validation, we demonstrate their widespread distribution across the bacterial kingdom, with being niche-specific and enriched in the human microbiome. Analyzing over 280,000 bacterial genomes, we reveal the diverse potential of human gut bacteria to produce these bacteriocins. Guided by meta-omics analysis, we synthesized 26 hypothetical bacteriocins from gut commensal species, with 16 showing antibacterial activities. Further ex vivo tests show minimal impact of narrow-spectrum bacteriocins on human fecal microbiota. Our study highlights the huge biosynthetic potential of unmodified bacteriocins in the human gut, paving the way for understanding their biological functions and health implications.

microbiology↗

bioRxiv2024

Dynamic Localization of Leukemia Stem Cells via CXCL12 Regulates Leukemia Progression

The processes that govern leukemia progression and remission are poorly understood. Our research reveals that the CXCL12 gradient, traditionally associated with LSC quiescence and survival, critically determines LSC localization and associated behavior. Specifically, CXCL12 guides LSCs to either the quiescent niche in the metaphysis, characterized by N-cadherin-expressing mesenchymal stromal cells (N-cad+ MSCs), or the proliferative niche in the central marrow (CM), marked by sinusoidal endothelial cells and associated stromal cells. We identified that the CXCL12 gradient is finely regulated by the interplay between dipeptidyl peptidase 4 (DPP4) on LSCs and glypican-3 (GPC3) on N-cad+ MSCs. DPP4 deactivates CXCL12, while GPC3 inhibits DPP4, resulting in a higher CXCL12 concentration in the metaphysis and a lower concentration in the CM. This differential gradient facilitates leukemia progression by promoting LSC quiescence and survival in the metaphysis versus relative proliferation and apoptosis in the CM. Depletion of Dpp4 from LSCs or Cxcl12 from N-cad+ MSCs disrupts this gradient, mobilizing LSCs from the metaphysis to the CM and significantly hindering leukemia development. Our findings redefine the role of CXCL12 in LSC behavior and provide a clearer understanding of leukemia progression. This novel insight highlights the potential for targeted therapeutic strategies that disrupt the CXCL12 gradient to treat minimal residual LSCs, offering a promising path toward a lasting cure for acute myeloid leukemia (AML).

cancer biology↗

bioRxiv2024

Genetic variation modulates susceptibility to aberrant DNA hypomethylation and imprint deregulation in naive pluripotent stem cells

Naive pluripotent stem cells (nPSC) frequently undergo pathological and not readily reversible loss of DNA methylation marks at imprinted gene loci. This abnormality poses a hurdle for using pluripotent cell lines in biomedical applications and underscores the need to identify the causes of imprint instability in these cells. We show that nPSCs from inbred mouse strains exhibit pronounced strain-specific susceptibility to locus-specific deregulation of imprinting marks during reprogramming to pluripotency and upon culture with MAP kinase inhibitors, a common approach to maintain naive pluripotency. Analysis of genetically highly diverse nPSCs from the Diversity Outbred (DO) stock confirms that genetic variation is a major determinant of epigenome stability in pluripotent cells. We leverage the variable DNA hypomethylation in DO lines to identify several trans-acting quantitative trait loci (QTLs) that determine epigenome stability at either specific target loci or genome-wide. Candidate factors encoded by two multi-target QTLs on chromosomes 4 and 17 suggest specific transcriptional regulators that contribute to DNA methylation maintenance in nPSCs. We propose that genetic variants represent candidate biomarkers to identify pluripotent cell lines with desirable properties and might serve as entry points for the targeted engineering of nPSCs with stable epigenomes. HighlightsO_LINaive pluripotent stem cells from distinct inbred mouse strains exhibit variable DNA methylation levels at imprinted gene loci. C_LIO_LIThe vulnerability of pluripotent stem cells to loss of genomic imprinting caused by MAP kinase inhibition strongly differs between inbred mouse strains. C_LIO_LIGenetically diverse pluripotent stem cell lines from Diversity Outbred mouse stock allow the identification of quantitative trait loci controlling DNA methylation stability. C_LIO_LIGenetic variants may serve as biomarkers to identify naive pluripotent stem cell lines that are epigenetically stable in specific culture conditions. C_LI

developmental biology↗

bioRxiv2024

Microsporidian Nosema bombycis secretes serine protease inhibitor to suppress host cell apoptosis via caspase BmICE

Microsporidia are a group of intracellular pathogens that actively manipulate host cell biological processes to facilitate their intracellular niche. Apoptosis is an important defense mechanism by which host cell control intracellular pathogens. Microsporidia modulating host cell apoptosis has been reported previously, however the molecular mechanism is not yet clear. In this report, we describe that the microsporidia Nosema bombycis inhibits apoptosis of Bombyx mori cells through a secreted protein NbSPN14, which is a serine protease inhibitor (Serpin). An immunofluorescent assay demonstrated that upon infection with N. bombycis, NbSPN14 was initially found in the B. mori cell cytoplasm and then became enriched in the host cell nucleus. Overexpression and RNA-interference (RNAi) of NbSPN14 in B. mori embryo cells confirmed that NbSPN14 inhibited host cell apoptosis. Immunofluorescent and Co-IP assays verified the co-localization and interaction of NbSPN14 with the BmICE, the caspase 3 homolog in B. mori. Knocking out of BmICE or mutating the BmICE-interacting P1 site of NbSPN14, eliminated the localization of NbSPN14 into the host nucleus and prevented the apoptosis-inhibiting effect of NbSPN14, which also proved that the interaction between BmICE and NbSPN14 occurred in host cytoplasm and the NbSPN14 translocation into host cell nucleus is dependent on BmICE. These data elucidate that N. bombycis secretory protein NbSPN14 inhibits host cell apoptosis by directly inhibiting the caspase protease BmICE, which provides an important insight for understanding pathogen-host interactions and a potential therapeutic target for N. bombycis proliferation. Author SummaryMicrosporidia constitute a class of eukaryotic pathogens that exclusively reside within host cells. The species Nosema bombycis is the first microsporidian identified as the pathogen of silkworm Pebrine disease. In our research, we discovered how N. bombycis cleverly evades the hosts defenses. It has developed a strategy to survive inside host cells by manipulating host cell apoptosis, disarming the host cells self-destruct mechanism. In this study, we discovered that the N. bombycis secretes a serine protease inhibitor named NbSPN14, which infiltrates the cytoplasm of the host cell. The NbSPN14 interacts with the executioner Caspase protease BmICE within the silkworms apoptotic pathway, effectively neutralizing its apoptoic activity and thus curbing the apoptosis of the host cells.

microbiology↗

bioRxiv2024

Filament structures unveil the dynamic organization of human acetyl-CoA 1 carboxylase

Human acetyl-CoA carboxylases (ACCs) catalyze the carboxylation of acetyl-CoA, which is the rate-limiting step in fatty acid synthesis. The molecular mechanism underlying the dynamic organization of ACCs is largely unknown. Here, we determined the cryo-EM structure of human ACC1 in its inactive state, which forms a unique filament structure and is in complex with acetyl-CoA. We also determined the cryo-EM structure of human ACC1 activated by dephosphorylation and citrate treatment, at a resolution of 2.55 [A]. Notably, the covalently linked biotin binds to a site that is distant from the acetyl-CoA binding site when acetyl-CoA is absent, suggesting a potential coordination between biotin binding and acetyl-CoA binding. These findings provide insights into the structural dynamics and regulatory mechanisms of human ACCs. TeaserCryo-EM structures reveal how human acetyl-CoA carboxylase 1 changes shape to control its activity.

biochemistry↗

bioRxiv2024

Two-hit mouse model of heart failure with preserved ejection fraction combining diet-induced obesity and renin-mediated hypertension

Heart failure with preserved ejection fraction (HFpEF) is increasingly common but its pathogenesis is poorly understood. The ability to assess genetic and pharmacologic interventions is hampered by the lack of robust preclinical mouse models of HFpEF. We have developed a novel "2-hit" model, which combines obesity and insulin resistance with chronic pressure overload to recapitulate clinical features of HFpEF. C57BL6/NJ mice fed a high fat diet for >10 weeks were administered an AAV8-driven vector resulting in constitutive overexpression of mouse Renin1d. Control mice, HFD only, Renin only and HFD-Renin (aka "HFpEF") littermates underwent a battery of cardiac and extracardiac phenotyping. HFD-Renin mice demonstrated obesity and insulin resistance, a 2-3-fold increase in circulating renin levels that resulted in 30-40% increase in left ventricular hypertrophy, preserved systolic function, and diastolic dysfunction indicated by altered E/e, IVRT, and strain measurements; increased left atrial mass; elevated natriuretic peptides; and exercise intolerance. Transcriptomic and metabolomic profiling of HFD-Renin myocardium demonstrated upregulation of pro-fibrotic pathways and downregulation of metabolic pathways, in particular branched chain amino acid catabolism, similar to findings in human HFpEF. Treatment of these mice with the sodium-glucose cotransporter 2 inhibitor empagliflozin, an effective but incompletely understood HFpEF therapy, improved exercise tolerance, left heart enlargement, and insulin homeostasis. The HFD-Renin mouse model recapitulates key features of human HFpEF and will enable studies dissecting the contribution of individual pathogenic drivers to this complex syndrome. Addition of HFD-Renin mice to the preclinical HFpEF model platform allows for orthogonal studies to increase validity in assessment of interventions. NEW & NOTEWORTHYHeart failure with preserved ejection fraction (HFpEF) is a complex disease to study due to limited preclinical models. We rigorously characterize a new two-hit HFpEF mouse model, which allows for dissecting individual contributions and synergy of major pathogenic drivers, hypertension and diet-induced obesity. The results are consistent and reproducible in two independent laboratories. This high-fidelity pre-clinical model increases the available, orthogonal models needed to improve our understanding of the causes and assessment treatments for HFpEF.

physiology↗

bioRxiv2024

CLCC1 Governs Bilayer Equilibration at the Endoplasmic Reticulum to Maintain Cellular and Systemic Lipid Homeostasis

The intricate orchestration of lipid production, storage, and mobilization is vital for cellular and systemic homeostasis1,2. Dysfunctional plasma lipid control represents the major risk factor for cardio-metabolic diseases, the leading cause of human mortality3,4. Within the cellular landscape, the endoplasmic reticulum (ER) is the central hub of lipid synthesis and secretion, particularly in metabolically active hepatocytes in the liver or enterocytes in the gut5,6. Initially assembled in the ER lumen, lipid-ferrying lipoproteins necessitate the cross-membrane transfer of both neutral and phospho-lipids onto the lumenal apolipoprotein B (APOB), in a poorly-defined process7-10. Here we show that trans-bilayer equilibration of phospholipids, regulated by the ER protein CLCC1, determines lipid partition across the ER membrane and consequently systemic lipid homeostasis. CLCC1 partners with the phospholipid scramblase TMEM41B11,12 to recognize imbalanced bilayers and promote lipid scrambling, thereby licensing lipoprotein biogenesis and the subsequent bulk lipid transport. Strikingly, loss of CLCC1 or TMEM41B leads to the emergence of giant lumenal lipid droplets enclosed by extensively imbalanced ER bilayers, and consequently drastically accelerated pathogenesis of metabolic-dysfunction-associated liver steatohepatitis (MASH). The above results establish phospholipid scrambling at the ER as the lynchpin to maintain a dynamic equilibrium. Considering the requirement of trans-bilayer phospholipid equilibration in numerous biological processes, ranging from catabolic autophagy to viral infection13-16, our study may enable further elucidation of a previously under-appreciated homeostatic control mechanism intrinsic to the ER function in lipid biogenesis and distribution.

cell biology↗

bioRxiv2024

Genomic and Genetic Insights into Mendel's Pea Genes

Pea, Pisum sativum, is an excellent model system through which Gregor Mendel established the foundational principles of inheritance. Surprisingly, till today, the molecular nature of the genetic differences underlying the seven pairs of contrasting traits that Mendel studied in detail remains partially understood. Here, we present a genomic and phenotypic variation map, coupled with haplotype-phenotype association analyses across a wide range of traits in a global Pisum diversity panel. We focus on a genomics-enabled genetic dissection of each of the seven traits Mendel studied, revealing many previously undescribed alleles for the four characterized genes, R, Le, I and A, and elucidating the gene identities and mutations for the remaining three uncharacterized traits. Notably, we identify: (1) a ca. 100kb deletion upstream of the Chlorophyll synthase (ChlG) gene, which generates aberrant transcripts and confers the yellow pod phenotype of gp mutants; (2) an in-frame premature stop codon mutation in a Dodeca-CLE41/44 signalling peptide which explains the parchmentless mutant phenotype corresponding to p; and (3) a 5bp in-frame deletion in a CIK-like receptor kinase gene corresponding to the fasciated stem phenotype fa, which Mendel described in terms of flower position, and we postulate the existence of a Modifier of fa (Mfa) locus that masks this meristem defect. Mendel noted the pleiotropy of the a mutation, including inhibition of axil ring anthocyanin pigmentation, a trait we found to be controlled by allelic variants of the gene D within an R2R3-MYB gene cluster. Furthermore, we characterize and validate natural variation of a quantitative genetic locus governing both pod width and seed weight, characters that Mendel deemed were not sufficiently demarcated for his analyses. This study establishes a cornerstone for fundamental research, education in biology and genetics, and pea breeding practices.

genetics↗

bioRxiv2024

Luminescent ingestible electronic capsules for in vivo regulation of optogenetic engineered bacteria

AbstractsThe ideal engineered microbial smart-drug should be capable of functioning on demand at specific sites in vivo. However, precise regulation of engineered microorganisms poses challenges in the convoluted and elongated intestines. Despite the promising application potential of optogenetic regulation strategies based on light signals, the poor tissue penetration of light signals limits their application in large experimental animals. Given the rapid development of ingestible electronic capsules in recent years, taking advantage of them as regulatory devices to deliver light signals in situ to engineered bacteria within the intestines has become feasible. In this study, we established an electronic-microorganism signaling system, realized by two Bluetooth-controlled luminescent electronic capsules were designed. The "Manager" capsule is equipped with a photosensor to monitor the distribution of engineered bacteria and to activate the optogenetic function of the bacteria by emitting green light. The other capsule, "Locator", can control the in situ photopolymerization of hydrogels in the intestines via ultraviolet light, aiding in the retention of engineered bacteria at specific sites. These two electronic capsules are expected to work synergistically to regulate the distribution and function of engineered bacteria in vivo, and their application in the treatment of colitis in pigs is currently being investigated, with relevant results to be updated subsequently.

synthetic biology↗

bioRxiv2024

Establishing remote communication between human operator and engineered intestinal microbe via bio-electronic optical language

Synthetic biology enables microbes to be engineered to detect biomarkers and deliver therapies in situ, offering advantages such as self-replication and localized response and treatment. However, the engineered microbes in the intestine remains a "black box", the status of microbial functionality is inaccessible, and the activation of microbial functions is uncontrollable. There is a demand of a "language" between microbes and human for real-time and in-situ monitoring and controlling engineered microbes in the intestine. Light, due to its advantages of specificity and accuracy in the body, was chosen as the language to establish an interactive communication path between human and the microbes with the features of sophisticated monitoring and controlling. For effectively remote operation of this "language" in the intestine, we introduced an electronic capsule as the intermedia, translating information and instructions understandable for human operator into bacterial regulatory and detectable signals in vivo. In this paper, the electronic capsule and the engineered Escherichia coli Nissle 1917 (EcN) strains were designed in a collaborative manner. An EcN-to-capsule bioluminescent signal transmission was established and testified as the language to report the status of engineered EcN, especially the response of EcN to intestinal environment. Similarly, another capsule-to-EcN optogenetic signal transmission was established and testified as the language to control the function of engineered EcN, especially the secretion of therapeutic substances. On this basis, bidirectional communication between human operator and intestinal microbes was achieved via bidirectional bio-electronic optical language. The communication was established to monitor and regulate engineered EcN community in the intestine in 50-90 kg live pigs. The in vitro and in vivo estimations demonstrated the capabilities of monitoring and regulating the engineered microbes, offering a remotely controllable language-based communication between human operator and internal intestinal microbes.

synthetic biology↗

bioRxiv2024

Targeting PRMT3 Impairs Methylation and Oligomerization of HSP60 to Boost Anti-Tumor Immunity by Activating cGAS/STING Signaling

Hepatocellular carcinoma (HCC) is a highly lethal cancer for which current available treatment options have limited efficacy. Immunotherapy has emerged as a promising therapeutic option for HCC, yet resistance to immunotherapy is a major challenge. Here, we uncover protein arginine methyltransferase 3 (PRMT3) as a novel driver of immunotherapy resistance in HCC. We show that PRMT3 expression is induced by activated T cells in response to immune checkpoint blockade (ICB) via an interferon-gamma (IFN{gamma})-STAT1 signaling pathway, and that high PRMT3 expression inversely correlates with tumor-infiltrating CD8+ T cells and predicts poor response to ICB in HCC patients. We demonstrate that genetic depletion and pharmacological inhibition of PRMT3 induce a profound influx of T cells into tumors and activate anti-tumor immunity to suppress HCC progression. Mechanistically, we demonstrate that PRMT3 methylates HSP60, a mitochondrial chaperone protein, at R446, and that this novel post-translational modification is required for HSP60 oligomerization and maintaining mitochondrial homeostasis. We reveal that targeting PRMT3-dependent HSP60-R446 methylation boosts anti-tumor immunity by disrupting mitochondrial function, increasing mitochondrial DNA (mtDNA) leakage, and activating the cGAS/STING pathway, a key innate immune sensor of cytosolic DNA. Importantly, we provide genetic and pharmacologic evidence that targeting PRMT3 enhances anti-PD1 efficacy and anti-tumor immunity in HCC mouse models. Our study identifies PRMT3 as a potential biomarker and therapeutic target to overcome immunotherapy resistance in HCC.

cancer biology↗

bioRxiv2024

Maladaptation in cereal crop landraces following a soot-producing climate catastrophe

Aerosol-producing global catastrophes such as nuclear war, super-volcano eruption, or asteroid strike, although rare, pose a serious threat to human survival. Light-absorbing aerosols would sharply reduce temperature and solar radiation reaching the earths surface, decreasing crop productivity including for locally adapted traditional crop varieties, i.e. landraces. Here, we test post-catastrophic climate impacts on barley, maize, rice, and sorghum, four crops with extensive landrace cultivation, under a range of nuclear war scenarios that differ in the amount of black carbon aerosol (soot) injected into the climate model. We used a crop growth model to estimate gradients of environmental stressors that drive local adaptation. We then fit genotype environment associations using high density genomic markers with gradient forest offset (GF offset) methods and predicted maladaptation through time. As a validation, we found that our GF models successfully predicted local adaptation of maize landraces in multiple common gardens across Mexico. We found strong concordance between GF offset and disruptions in climate, and landraces of all tested crop species were predicted to be the most maladapted across space and time where soot-induced climate change was the greatest. We further used our GF models to identify landrace varieties best matched to specific post-catastrophic conditions, indicating potential substitutions for agricultural resilience. We found the best landrace genotype was often far away or in another nation, though countries with more climatic diversity had better within-country substitutions. Our results highlight that a soot-producing catastrophe would result in the global maladaptation of landraces and suggest that current landrace adaptive diversity is insufficient for agricultural resilience in the case of the soot scenarios with the greatest change to climate.

evolutionary biology↗

bioRxiv2024

Structural insights into human propionyl-CoA carboxylase (PCC) and 3-methylcrotonyl-CoA carboxylase (MCC)

Propionyl-CoA carboxylase (PCC) and 3-methylcrotonyl-CoA carboxylase (MCC) are biotin-dependent carboxylases (BDCs) that catalyze the metabolism of odd-chain fatty acids, cholesterol, and specific amino acids. For human PCC and MCC, only a low-resolution (15 [A]) three-dimensional structure of human PCC has been reported. Here, we report high-resolution (2.29-3.38 [A]) cryo-EM structures of human PCC and MCC holoenzymes in their apo and acetyl-CoA and propionyl-CoA-bound states. Propionyl-CoA and acetyl-CoA bind to PCC with almost identical binding modes, indicating that the acyl-CoA specificity of PCC is largely attributed to minor differences in interactions mediated by the acyl groups. In MCC, biotin is relocated from an exo-site to an endo-site upon acetyl-CoA binding, suggesting coordination between biotin binding and acyl-CoA binding. Our work provides insights into the substrate specificity and catalytic process of BDCs.

biochemistry↗

bioRxiv2024

IL-13 and IL-17A Activate β1 Integrin through an NF-κB/Rho kinase/PIP5K1γ pathway to Enhance Force Transmission in Airway Smooth Muscle

Integrin activation resulting in enhanced adhesion to the extracellular matrix plays a key role in fundamental cellular processes. Although G-protein coupled receptor-mediated integrin activation has been extensively studied in non-adherent migratory cells such as leukocytes and platelets, much less is known about the regulation and functional impact of integrin activation in adherent stationary cells such as airway smooth muscle. Here we show that two different asthmagenic cytokines, IL-13 and IL-17A, activate type I and IL-17 cytokine receptor families respectively, to enhance adhesion of muscle to the matrix. These cytokines also induce activation of {beta}1 integrins as detected by the conformation-specific antibody HUTS-4. Moreover, HUTS-4 binding is significantly increased in the smooth muscle of patients with asthma compared to healthy controls, suggesting a disease-relevant role for aberrant integrin activation. Indeed, we find integrin activation induced by a {beta}1 activating antibody, the divalent cation manganese, or the synthetic peptide {beta}1-CHAMP, dramatically enhances force transmission in collagen gels, mouse tracheal rings, and human bronchial rings even in the absence of cytokines. We further demonstrate that cytokine-induced activation of {beta}1 integrins is regulated by a common pathway of NF-O_SCPLOWKC_SCPLOWB-mediated induction of RhoA and its effector Rho kinase, which in turn stimulates PIP5K1{gamma}-mediated synthesis of PIP2 resulting in {beta}1 integrin activation. Taken together, these data identify a previously unknown pathway by which type I and IL-17 cytokine receptor family stimulation induces functionally relevant {beta}1 integrin activation in adherent smooth muscle and help explain the exaggerated force transmission that characterizes chronic airways diseases such as asthma. SIGNIFICANCE STATEMENTIntegrin activation plays a central role in regulating cellular adhesion and migration. While chemokine-mediated integrin activation has been extensively studied in circulating cells, the role and impact of other cytokine families on non-migratory cells remains incompletely characterized. Here, we demonstrate in airway smooth muscle that asthmagenic cytokines IL-13 and IL-17A stimulate type I and IL-17 cytokine receptor families to induce {beta}1 integrin activation and enhance adhesion. We also identify a common pathway linking NF-O_SCPLOWKC_SCPLOWB/RhoA/Rho kinase with PIP5K1{gamma}/PIP2/{beta}1 integrin activation. We show that airway biopsies from asthmatic patients have increased active {beta}1 integrin staining in the muscle, and furthermore that {beta}1 integrin activation alone dramatically enhances force transmission, underscoring the disease-relevant impact of cytokine-mediated integrin activation in adherent muscle.

molecular biology↗

bioRxiv2024

Cyclical transcription factor AP2XII-9 is a key activator for asexual division and apicoplast inheritance in Toxoplasma gondii tachyzoite

Toxoplasma gondii is an intracellular parasitic protozoan that poses a significant risk to pregnant women and immunocompromised individuals. T. gondii tachyzoites duplicate rapidly in host cells during acute infection through endodyogeny. This highly regulated division process is accompanied by complex gene regulation networks. TgAP2XII-9 is a cyclical transcription factor, but its specific role in the parasite cell cycle is not fully understood. Here, we demonstrate that TgAP2XII-9 is identified as a nuclear transcription factor and is dominantly expressed during the S/M phase of the tachyzoite cell cycle. CUT&Tag results indicate that TgAP2XII-9 targets key genes for the moving junction machinery (RON2, 4, 8) and daughter cell inner membrane complex (IMC). TgAP2XII-9 deficiency resulted in a significant downregulation of rhoptry proteins and rhoptry neck proteins, leading to a severe defect in the invasion and egress efficiency of tachyzoites. Additionally, the loss of TgAP2XII-9 correlated with a substantial downregulation of multiple IMC and apicoplast proteins, leading to disorders of daughter bud formation and apicoplast inheritance, and further contributing to the inability of cell division and intracellular proliferation. Our study reveals that TgAP2XII-9 acts as a critical S/M-phase regulator that orchestrates the endodyogeny and apicoplast division in T. gondii tachyzoite. This study contributes to a broader understanding of the complexity of the parasites cell cycle and its key regulators. Significance: The intracellular apicoplast parasite Toxoplasma gondii posts great threat to the public health. The acute infection of T. gondii tachyzoite relies on efficient invasion by forming a moving junction structure and also fast replication by highly regulated endodyogeny. This study shows that an ApiAP2 transcription factor TgAP2XII-9 acts as an activator for the S/M-phase gene expression, including genes related to daughter buds and moving junction formation. Loss of TgAP2XII-9 results significant growth defects and disorders in endodyogeny and apicoplast inheritance of the parasites. Our results provide valuable insights into the transcriptional regulation of parasite cell cycle and invading machinery in T. gondii.

microbiology↗

bioRxiv2024

CCR7+ Dendritic Cells Define a Type 17 Module in Psoriasis

Interleukin (IL)-23 is the master pathogenic cytokine in psoriasis and neutralization of IL-23 alleviates psoriasis. Psoriasis relapses after the withdrawal of IL-23 antibodies, and the persistence of IL-23-producing cells probably contributes to such recurrence. However, the cellular source of IL-23 was unclear, which hinders the development of targeted therapies focusing on modulating IL-23 expression aimed at resolving relapse. Here, we showed that IL4I1+CD200+CCR7+ dendritic cells (CCR7+ DC) dominantly produced IL-23 by concomitantly expressing the IL-23A and IL-12B subunits in human psoriatic skin. Deletion of CCR7+ DC completely abrogated IL-23 production in a mouse model of psoriasis and enforced expression of IL-23a in CCR7+ DC elicited not only {beta}T cell-driven psoriasis-like skin disease, but also arthritis. CCR7+ DC co-localized with CD161+ IL-17-producing T cells and KRT17+ keratinocytes, which were located in the outermost layers of psoriatic epidermis and strongly exhibited IL-17 downstream signatures. Based on these data, we identified CCR7+ DC as the source of IL-23 in psoriasis, which paves the way for the design of therapies focused on manipulating IL-23 production that may resolve the relapse of chronic inflammatory disorders like psoriasis. HIGHLIGHTSO_LIIL4I1+CD200+CCR7+ DC are dominant IL-23 producers in psoriasis and its mouse model. C_LIO_LIPsoriatic CCR7+ DC likely arise from cDC2. C_LIO_LICD161 marks all IL-17-producing T cells in psoriatic skin. C_LIO_LIIL-23a overexpression in CCR7+ DC elicits an {beta}T cell-driven mouse model of psoriasis and arthritis. C_LIO_LICCR7+ DC spatially define a type 17 module in psoriatic epidermis. C_LI

immunology↗