Biology subjects

Luo, Y.

Publications and source records attributed to Luo, Y..

At least 181 records · Page 10Linked to original sources

bioRxiv2024

LncCE: Landscape of Cellular-Elevated LncRNAs in Single Cells Across Normal and Cancer Tissues

Long non-coding RNAs (lncRNAs) have emerged as significant players for maintaining the morphology and function of tissues or cells. The precise regulatory effectiveness of lncRNA is closely associated with the spatial expression patterns across tissues and cells. Here, we proposed the Cellular-Elevated LncRNA (LncCE) database to systematically explore cellular-elevated (CE) lncRNAs across normal and cancer tissues in single cells. LncCE encompasses 87,946 CE lncRNAs of 149 cell types by analyzing 181 single-cell RNA sequencing (scRNA-seq) datasets, involved in 20 fetal normal tissues, 59 adult normal tissues, as well as 32 adult and 5 pediatric cancer tissues.Two main search options were provided via a given lncRNA name or a cell type. The output results emphasize both qualitative and quantitative expression features of lncRNAs across different cell types, co-expression with protein-coding genes as well as their involved in biological functions. For cancers, LncCE particularly provided quantitative figures for exhibiting their expression changes compared to control samples and clinical associations with patient overall survivals. Together, LncCE offers an extensive, quantitative and user-friendly interface to investigate cellular-elevated expression atlas for lncRNAs across normal and cancers tissues at single-cell level. The LncCE database is available at http://bio-bigdata.hrbmu.edu.cn/LncCE.

cancer biology↗

bioRxiv2024

Mapping Endothelial-Macrophage Interactions in Diabetic Vasculature: Role of TREM2 in Vascular Inflammation and Ischemic Response

Diabetes mellitus (DM) significantly accelerates vascular diseases like peripheral arterial disease (PAD). Endothelial cells (ECs) and macrophages (M{Phi}s) singularly and synergistically are important contributors to DM-associated vascular dysfunction. Single-cell (sc) profiling technologies are revealing the true heterogeneity of ECs and M{Phi}s, but how this cellular diversity translates to cell-cell interactions, and consequentially vascular function, remains unknown. We leveraged scRNA sequencing and spatial transcriptome (ST) profiling to analyze human mesenteric arteries from non-diabetic (ND) and type 2 diabetic (T2D) donors. We generated a transcriptome and interactome map encompassing the major arterial cells and highlighted Triggering Receptor Expressed on Myeloid Cells 2 (TREM2) as a top T2D-induced gene in mononuclear phagocytes (MPs), with concomitant increases of TREM2 ligands in ECs. We verified DM-associated TREM2 induction in cell and mouse models, and found that TREM2 inhibition decreases pro-inflammatory responses in MPs and ECs, as well as increases EC migration in vitro. Furthermore, TREM2 inhibition using a neutralizing antibody enhanced ischemic recovery and flow reperfusion in DM mice subjected to hindlimb ischemia, suggesting that TREM2 promotes ischemic injury in DM. Finally, in human PAD, co-existing DM was associated with greater expression of TREM2 and its interaction with ECs, with a further increase in ischemic tissue compared to patient-matched non-ischemic tissue. Collectively, our study presents the first atlas of human diabetic vessels with single cell and spatial resolution, and identifies TREM2-EC interaction as a key driver of diabetic vasculopathies, the targeting of which may offer an opportunity to ameliorate vascular dysfunction associated with DM-PAD.

cell biology↗

bioRxiv2024

Comparative Analysis of the Techniques for the Determination of Binding Affinity between a Small Molecule Inhibitor and a Protein Target

The binding affinity constant (KD) between a small molecule inhibitor and a protein target is pivotal parameter for target identification and early drug discovery. Despite the extensive applications of three major techniques, namely SPR, ITC and MST, the KD values for a specifically defined binding measured by these techniques are drastically different, which poses a remarkable difficulty to deal with these ambiguous data. Here, we report the evaluation of the accuracy of KD values from SPR, ITC and MST compared with enzyme kinetics. To enable an objective comparison, we theoretically proved that enzyme competitive inhibition constant (Ki) could directly reflect the binding affinity. Using purine nucleoside phosphorylase, its substrate inosine and competitive inhibitor immucillin-H, we determined respective KD and Ki values to make a direct comparison. Moreover, we found that the KD value measured by SPR is more relevant to its Ki value. This study highlights the urgent need on the development of new technologies for the determination of binding affinity between small molecule inhibitors and protein targets.

biophysics↗

bioRxiv2024

Pooled genetic screens identify breast cancer risk genes involved in evasion from T cell-mediated killing

Genome-wide association studies have identified more than 220 loci associated with breast cancer susceptibility. A major challenge is now to identify the effector genes with plausible functions in the context of breast cancer risk. We have previously performed pooled CRISPR screens to identify target genes at risk loci that drive cancer hallmarks including proliferation or modulating DNA damage response. We now extend these screens to identify genes involved in response to cytotoxic T lymphocyte (CTL) killing. We performed knockout and inhibition screens to identify genes that affect the response of the MCF7 human breast cancer cell line to CTL killing in an in vitro co-culture system. We identified 33 candidate risk genes associated with resistance or sensitisation to T cell-mediated killing. Using single gene perturbation, we showed that deletion of candidate risk genes IRF1, ATF7IP, CCDC170 and CASP8 induced resistance, while ablation of CFLAR, CREBBP, and PRMT7 sensitized cells to CTL killing. We used reporter assays to show that the risk-associated alleles at rs736801 and rs3769821 reduced transactivation of the IRF1 and CASP8 promoters, respectively. We showed that pharmacological inhibition of PRMT7 rendered breast cells sensitive to CTL killing and PRMT7 levels were negatively correlated with CD8+ infiltration and patient survival in luminal A breast cancer patient cohorts. Our results demonstrate that phenotypic pooled CRISPR screens are a useful approach for high throughput functional follow-up of GWAS findings, identifying genes which alter immune responses to breast cancer which offer opportunities to enhance immunotherapy.

genetics↗

bioRxiv2024

An ultra-short-acting benzodiazepine in thalamic nucleus reuniens undermines fear extinction via intermediation of hippocamposeptal circuits

Benzodiazepines, commonly used for anxiolytics, hinder conditioned fear extinction, and the underlying circuit mechanisms are unclear. Utilizing remimazolam, an ultra-short-acting benzodiazepine, we reveal its impact on the thalamic nucleus reuniens (RE) and interconnected hippocamposeptal circuits during fear extinction. Systemic or RE-specific administration of remimazolam impedes fear extinction by reducing RE activation through A type GABA receptors. Remimazolam enhances long-range GABAergic inhibition from lateral septum (LS) to RE, underlying the compromised fear extinction. RE projects to ventral hippocampus (vHPC), which in turn sends projections characterized by feed-forward inhibition to the GABAergic neurons of the LS. This is coupled with long-range GABAergic projections from the LS to RE, collectively constituting an overall positive feedback circuit construct that promotes fear extinction. RE-specific remimazolam negates the facilitation of fear extinction by disrupting this circuit. Thus, remimazolam in RE disrupts fear extinction caused by hippocamposeptal intermediation, offering mechanistic insights for the dilemma of combining anxiolytics with extinction-based exposure therapy.

neuroscience↗

bioRxiv2024

Linkage: an interactive shiny app and R package for linking of DNA regulatory peaks to genes

SummaryExisting studies have demonstrated that the integration analysis of transcriptomic and epigenomic data can be used to better understand the onset and progression of many diseases, as well as identify new diagnostic and prognostic biomarkers. However, such investigations on large-scale sequencing data remain challenging for researchers or clinicians with limited bioinformatics knowledge. To facilitate the interpretation of gene regulatory landscape, we developed an R Shiny application and R package [Linking of atac-seq to gene expression data (Linkage)] for exploring and visualizing potential cis-regulatory elements (CREs) of genes based on ATAC-seq and RNA-seq data. Linkage offers six modules to systematically identify, annotate, and interpret potential gene regulatory elements from the whole genome step by step. Linkage can provide interactive visualization for the correlation between chromatin accessibility and gene expression. More than that, Linkage identifies transcription factors (TFs) that potentially drive the chromatin changes through identifying TF binding motifs within the CREs and constructing trans-regulatory networks of the target gene set. This powerful tool enables researchers to conduct extensive multiomics integration analysis and generate visually appealing visualizations that effectively highlight the relationship between genes and corresponding regulatory elements. With Linkage, users can obtain publishable results and gain deeper insights into the gene regulatory landscape. Availability and implementation Linkage is freely available as a Shiny web application (https://xulabgdpu.org.cn/linkage) and an R package (https://github.com/XuLab-GDPU/Linkage). The documentation is available at (https://aicplane.github.io/Linkage-tutorial/).

bioinformatics↗

bioRxiv2024

Generative Modelling of Oncogene-carrying Extrachromosomal Circular DNA Biogenesis and Dynamics in Cells

Extrachromosomal circular DNAs (ecDNA) are focal gene amplifications frequently associated with cancer development and often indicating a poor prognosis. To understand the early dynamics of oncogene-carrying ecDNAs, we previously developed CRISPR-C, a tool for precise ecDNA generation by deleting specific chromosomal regions. Here, we adapted CRISPR-C to recreate tumor ecDNAs. This method also allowed us to enhance ecDNA generation efficiency by directly delivering Cas9 protein and sgRNAs as a ribonucleoprotein complex. By using the modified CRISPR-C, we successfully generated ecDNAs carrying oncogenes (EGFR, CDK4, MDM2, MYC, MYCN, FGFR2, ABCB1, and DHFR) in various human cell types. Furthermore, we demonstrated that our method could generate chimeric ecDNAs composed of target sequences from distant intra or inter-chromosomal regions. Using these generative ecDNA cell models, we studied the oncogene ecDNA expression and stability. The MDM2 expression was increased after CRISPR-C, while CDK4 was decreased indicating genomic-context dependent effect. The copy number of CRISPR-C generated CDK4 was ecDNA increased in cells after a long period of treatment with the CDK4 inhibitor palbociclib. Unlike CDK4, the CRISPR-C generated ABCB1 ecDNA was unstable in cells under normal growth conditions, but is stably retained when the cells were treated with colcemid, a recognized substrate for ABCB1. We thus provide valuable tools and an attractive platform for studying ecDNA biogenesisy and in vitro drug screening on ecDNA stability.

cancer biology↗

bioRxiv2024

Complementing Muscle Regeneration: Fibro-Adipogenic Progenitor and Macrophage-Mediated Repair of Elderly Human Skeletal Muscle

The capacity to regenerate skeletal muscle function after injury requires a complex and well-coordinated cellular response. Here, we unravel the intricate dynamics of human skeletal muscle regeneration by combining spatial, temporal, and single cell transcriptomics. Using spatial RNA sequencing, we profiled the expression of human protein-coding genes in elderly human skeletal muscle biopsies before as well as 2-, 8-, and 30-days post injury. Single Cell-Spatial deconvolution analysis highlights monocytes/macrophages and fibro-adipogenic progenitors (FAPs) as pivotal players in human muscle regeneration. By utilizing flow cytometry and cell sorting we confirmed increased cellular content and activity during regeneration. Spatial correlation analysis unveils FAPs and monocytes/macrophages co-localization and intercellular communication, mediated by complement factor C3. Immunostaining confirms C3 expression in FAPs and FAP secretion of C3, suggesting a role in phagocytosis. Functional assays demonstrate C3s impact on monocyte metabolism, survival and phagocytosis, unveiling its involvement in skeletal muscle regeneration. These insights elucidate FAP-macrophage interplay with perspectives for future therapeutic interventions.

physiology↗

bioRxiv2024

FLASH Irradiation Regulates IFN-β induction by mtDNA via Cytochrome c Leakage

Ultrahigh dose rate radiotherapy (FLASH-RT) is under intensive investigation for its biological benefits. The mechanisms underlying its ability to spare normal tissues while suppress tumor growth still remain controversial. Here we reveal that compared to the low dose rate electron irradiation (0.36 Gy/s), FLASH electron irradiation at 61 or 610 Gy/s enhances the cytochrome c leakage from mitochondria in human breast cells MCF-10A, which elicits substantial caspase activation, suppresses both the cytosolic mitochondrial DNA (mtDNA) accumulation and IFN-{beta} secretion. Besides, the deletion of mtDNA severely decreases the radiation-induced cGAS-STING activation. Conversely, the cytochrome c leakage in carcinoma cells MDA-MB-231 post electron irradiation is limited, especially for the case of FLASH irradiation, resulting in less cytosolic cytochrome c but stronger cGAS-STING activation than those in MCF-10A cells. The enhanced difference of cytochrome c leakage between cancer cells and normal cells post FLASH irradiation indicates a potential mechanism of FLASH effect by regulating the apoptotic and inflammatory pathway.

biophysics↗

bioRxiv2024

Caspase-8-mediated CYLD cleavage boots LPS-induced endotoxic shock.

Caspase-8, a pivotal protease intricately involved in various cellular signaling pathways related to cell death and inflammation, has been identified as a contributor to cytokine production during septic shock. However, the mechanisms governing this regulatory role remain enigmatic. In this study, we uncovered that mice harboring a specific mutation in CYLD at its D215 position (CyldD215A/D215A mutant mice), rendering CYLD resistant to caspase8 cleavage, exhibited marked protection against lethal endotoxic shock. Moreover, the removal of Cyld in Caspase8-/-Mlkl-/- mice restored their sensitivity to endotoxic shock, indicating Caspase8 promotes LPS-induced endotoxic shock by cleaving its substrate CYLD and maintaining CYLD stability confers resistance to endotoxic shock. Mechanistically, CYLD was found to catalyze the removal of LUBAC-mediated M1-linked ubiquitination of NF-kB p65 at K301/K303, thereby suppressing the nuclear translocation and activation of p65 for subsequent cytokines production. Notably, the cleaved N-terminal fragment of CYLD (named CP25) is secreted in a TRIF/Caspase8-dependent manner. Moreover, CP25 can be detected in the serum of septic mice, and its levels show a strong correlation with corresponding serum IL-1{beta}, suggesting its potential utility as an inflammatory biomarker. Overall, these findings highlight the significance of CYLD cleavage as a promising therapeutic target and diagnostic marker for endotoxic shock.

immunology↗

bioRxiv2024

Advancing CAR T-Cell Therapy: Simultaneously Attack Tumor and Immunosuppressive Cells in the Tumor Microenvironment

Chimeric antigen receptor (CAR) T-cell therapy has encountered limited success in solid tumors. The lack of dependable antigens and the immunosuppressive tumor microenvironment (TME) are major challenges. Within the TME, tumor cells along with immunosuppressive cells employ an immune-evasion mechanism that upregulates programmed death ligand 1 (PD-L1) to deactivate effector T cells; this makes PD-L1 a reliable, universal target for solid tumors. We developed a novel PD-L1 CAR (MC9999) using our humanized anti-PD-L1 monoclonal antibody, designed to simultaneously target tumor and immunosuppressive cells. The antigen-specific antitumor effects of MC9999 CAR T-cells were observed consistently across four solid tumor models: breast cancer, lung cancer, melanoma, and glioblastoma multiforme (GBM). Notably, intravenous administration of MC9999 CAR T-cells eradicated intracranially established LN229 GBM tumors, suggesting penetration of the blood-brain barrier. The proof-of-concept data demonstrate the cytolytic effect of MC9999 CAR T-cells against immunosuppressive cells, including microglia HMC3 cells and M2 macrophages. Furthermore, MC9999 CAR T-cells elicited cytotoxicity against primary tumor-associated macrophages within GBM tumors. The concept of targeting both tumor and immunosuppressive cells with MC9999 was further validated using CAR T-cells derived from cancer patients. These findings establish MC9999 as a foundation for the development of effective CAR T-cell therapies against solid tumors.

cancer biology↗

bioRxiv2024

Evolutionary trajectory of TRPM2 channel activation by adenosine diphosphate ribose and calcium

Ion channel activation upon ligand gating triggers a myriad of biological events and, therefore, evolution of ligand gating mechanism is of fundamental importance. TRPM2, a typical ancient ion channel, is activated by adenosine diphosphate ribose (ADPR) and calcium and its activation has evolved from a simple mode in invertebrates to a more complex one in vertebrates, but the evolutionary process is still unknown. Molecular evolutionary analysis of TRPM2s from more than 280 different animal species has revealed that, the C-terminal NUDT9-H domain has evolved from an enzyme to a ligand binding site for activation, while the N-terminal MHR domain maintains a conserved ligand binding site. Calcium gating pattern has also evolved, from one Ca2+-binding site as in sea anemones to three sites as in human. Importantly, we identified a new group represented by olTRPM2, which has a novel gating mode and fills the missing link of the channel gating evolution. We conclude that the TRPM2 ligand binding or activation mode evolved through at least three identifiable stages in the past billion years from simple to complicated and coordinated. Such findings benefit the evolutionary investigations of other channels and proteins.

evolutionary biology↗

bioRxiv2024

CORACLE (COVID-19 liteRAture CompiLEr): A platform for efficient tracking and extraction of SARS-CoV-2 and COVID-19 literature, with examples from post-COVID with respiratory involvement

BackgroundDuring the COVID-19 pandemic there emerged a need to efficiently monitor and process large volumes of scientific literature on the subject. Currently, as the pandemic is winding down, the clinicians encountered a novel syndrome - Post-acute Sequelae of COVID- 19 (PASC) - that affects over 10% of those who contract SARS-CoV-2 and presents a significant and growing challenge in the medical field. The continuous influx of new research publications underscores a critical need for efficient tools for navigating the literature. ObjectivesWe aimed to develop an application which will allow monitoring and categorizing COVID-19-related literature through building publication networks and medical subject headings (MeSH) maps to be able to quickly identify key publications and publication networks. MethodsWe introduce CORACLE (COVID-19 liteRAture CompiLEr), an innovative web application designed for the analysis of COVID-19-related scientific articles and the identification of research trends. CORACLE features three primary interfaces: The "Search" interface, which displays research trends and citation links; the "Citation Map" interface, allowing users to create tailored citation networks from PubMed Identifiers (PMIDs) to uncover common references among selected articles; and the "MeSH" interface, highlighting current MeSH trends and associations between MeSH terms. ResultsOur web application, CORACLE, leverages regularly updated PubMed data to aggregate and categorize the extensive literature on COVID-19 and PASC, aiding in the identification of relevant research publication hubs. Using lung function in PASC patients as a search example, we demonstrate how to identify and visualize the interactions between the relevant publications. ConclusionCORACLE proves to be an effective tool for the extraction and analysis of literature. Its functionalities, including the MeSH trends and customizable citation mapping, facilitate the discovery of relevant information and emerging trends in COVID-19 and PASC research.

bioinformatics↗

bioRxiv2024

Lipophilic compounds restore wt function of neurodevelopmental-associated KCNQ3 mutations.

A major driver of neuronal hyperexcitability is dysfunction of K+ channels, including voltage-gated KCNQ2/3 channels. Their slow activation and deactivation kinetics produces a current that regulates membrane potential and impedes repetitive firing. Mutations in KCNQ2 and KCNQ3 lead to a wide spectrum of neurodevelopmental disorders (NDDs), ranging from benign familial neonatal seizures to severe epileptic encephalopathies and autism spectrum disorders. However, the impact of these mutations on KCNQ channel function remains poorly understood and existing treatments have unpleasant side effects. Here we use voltage clamp fluorometry and molecular dynamic simulations to investigate how R227Q and R236C, two novel NDD-causing mutations in the voltage sensor of KCNQ3, impair channel function. We show that the two mutations perturb channel gating by two distinct mechanisms: R227Q altering voltage sensor movement and R236C altering voltage sensor-to-gate coupling. Our study further shows that polyunsaturated fatty acids (PUFAs), a novel class of ion channel modulators, primarily target the voltage sensor domain in its activated conformation and yield partial and complete restoration of wt function in R227Q- and R236C-containing channels, respectively. Our results reveal the potential of PUFAs to be developed into therapies for diverse KCNQ3-based channelopathies.

biophysics↗

bioRxiv2024

Emodin: an alveolar macrophage protector in acute pancreatitis induced lung injury

Emodin (EMO), an anthraquinone derivative from roots and leaves of various plants, has been widely used in many inflammatory diseases. Alveolar macrophages (AMs) play a critical role in maintaining alveolar homeostasis in the lung. To investigate the pathophysiological mechanism of AMs in acute pancreatitis-associated lung injury (AP-ALI) and the potential protective therapeutic of EMO for AP-ALI, AMs isolated from AP-ALI mice, murine cell line MH-S and RAW264.7 were pre-treated with EMO to assess its protective roles on regulating inflammation, pyroptosis, and mitophagy of macrophages in AP-ALI. The results showed that 1) in vivo, the relative quantity of AMs was significantly decreased across the time in API-ALI; however, the mitochondria flux presented earlier changes than the resident AMs alteration in our experimental system. EMO pretreatment significantly alleviated the severity of lung injury and improved the damaged alveolar structure, and further reversed the reduction of mitochondria impairment in AMs in mice. 2) In vitro, EMO significantly suppressed LPS/ATP associated NLRP3 inflammasome activation, enhanced mitophagy, and protected mitochondria damage. Furthermore, the data of mitophagy inhibition by 3-MA demonstrated that EMO protective effects were partially via manipulating mitophagy-mitochondria-alveolar macrophage axis. Our data shed light on the comprehensive understanding on EMO therapeutics in AP-ALI.

molecular biology↗

bioRxiv2024

Transcriptional Regulation of Protein Synthesis by Mediator Kinase in MYC-driven Medulloblastoma

MYC-driven medulloblastoma (MB) is a highly aggressive cancer type with poor prognosis and limited treatment options. Through CRISPR-Cas9 screening across MB cell lines, we identified the Mediator-associated kinase CDK8 as the top dependence for MYC-driven MB. Loss of CDK8 markedly reduces MYC expression and impedes MB growth. Mechanistically, we demonstrate that CDK8 depletion suppresses ribosome biogenesis and mRNA translation. CDK8 regulates occupancy of phospho-Polymerase II at specific chromatin loci facilitating an epigenetic alteration that promotes transcriptional regulation of ribosome biogenesis. Additionally, CDK8-mediated phosphorylation of 4EBP1 plays a crucial role in initiating eIF4E-dependent translation. Targeting CDK8 effectively suppresses cancer stem and progenitor cells, characterized by increased ribosome biogenesis activity. We also report the synergistic inhibition of CDK8 and mTOR in vivo and in vitro. Overall, our findings establish a connection between transcription and translation regulation, suggesting a promising therapeutic approach targets multiple points in the protein synthesis network for MYC-driven MB.

cancer biology↗

bioRxiv2024

Technical and biological variations in the purification of extrachromosomal circular DNA (eccDNA) and the finding of more eccDNA in the plasma of lung adenocarcinoma patients compared with healthy donors

Human plasma DNA originates from all tissues and organs, holding the potential as a versatile marker for diseases such as cancer, as fragments of cancer-specific alleles can be found circulating in the blood. While linear DNA has been studied intensely as a liquid biomarker, the role of circular circulating DNA in cancer is more unknown due, in part, to a lack of comprehensive testing methods. Our developed method profiles extrachromosomal circular DNA (eccDNA) in plasma, integrating Solid-Phase Reversible Immobilization (SPRI) bead purification, the removal of linear DNA and mitochondrial DNA, and DNA sequencing. As an initial assessment, we examined the method, biological variations, and technical variations using plasma samples from four patients with lung adenocarcinoma and four healthy and physically fit individuals. Despite the small sample group, we observed a significant eccDNA increase in cancer patients in two independent laboratories and that eccDNA covered up to 0.4 % of the genome/mL plasma. We found a subset of eccDNA from recurrent genes present in cancer samples but not in every control. In conclusion, our data reflect the large variation found in eccDNA sequence content and show that the variability observed among replicates in eccDNA stems from a biological source and can cause inconclusive findings for biomarkers. This suggests the need to explore other biological markers, such as epigenetic features on eccDNA.

molecular biology↗

bioRxiv2024

The Bacillus subtilis yqgC-sodA operon protects magnesium-dependent enzymes by supporting manganese efflux

Microbes encounter a myriad of stresses during their life cycle. Dysregulation of metal ion homeostasis is increasingly recognized as a key factor in host-microbe interactions. Bacterial metal ion homeostasis is tightly regulated by dedicated metalloregulators that control uptake, sequestration, trafficking, and efflux. Here, we demonstrate that deletion of the Bacillus subtilis yqgC-sodA (YS) complex operon, but not deletion of the individual genes, causes hypersensitivity to manganese (Mn). YqgC is an integral membrane protein of unknown function and SodA is a Mn-dependent superoxide dismutase (MnSOD). The YS strain has reduced expression of two Mn efflux proteins, MneP and MneS, consistent with the observed Mn sensitivity. The YS strain accumulated high levels of Mn, had increased reactive radical species (RRS), and had broad metabolic alterations that can be partially explained by the inhibition of Mg-dependent enzymes. Although the YS operon deletion strain and an efflux-deficient mneP mneS double mutant both accumulate Mn and have similar metabolic perturbations they also display phenotypic differences. Several mutations that suppressed Mn intoxication of the mneP mneS efflux mutant did not benefit the YS mutant. Further, Mn intoxication in the YS mutant, but not the mneP mneS strain, was alleviated by expression of Mg-dependent, chorismate-utilizing enzymes of the menaquinone, siderophore, and tryptophan (MST) family. Therefore, despite their phenotypic similarities, the Mn sensitivity in the mneP mneS and the yqgC-sodA deletion mutants results from distinct enzymatic vulnerabilities. ImportanceBacteria require multiple trace metal ions for survival. Metal homeostasis relies on the tightly regulated expression of metal uptake, storage, and efflux proteins. Metal intoxication occurs when metal homeostasis is perturbed and often results from enzyme mis-metalation. In Bacillus subtilis, MnSOD is the most abundant Mn-containing protein and is important for oxidative stress resistance. Here, we report novel roles for MnSOD and a co-regulated membrane protein, YqgC, in Mn homeostasis. Loss of both MnSOD and YqgC (but not the individual mutations) prevents the efficient expression of Mn efflux proteins and leads to a large-scale perturbation of the metabolome due to inhibition of Mg-dependent enzymes, including key chorismate-utilizing MST (menaquinone, siderophore, and tryptophan) family enzymes.

microbiology↗