Biology subjects

Cheng, C.

Publications and source records attributed to Cheng, C..

At least 91 records · Page 5Linked to original sources

bioRxiv2023

LINE-associated cryptic splicing induces dsRNA-mediated interferon response and tumor immunity

RNA splicing plays a critical role in post-transcriptional gene regulation. Exponential expansion of intron length poses a challenge for accurate splicing. Little is known about how cells prevent inadvertent and often deleterious expression of intronic elements due to cryptic splicing. In this study, we identify hnRNPM as an essential RNA binding protein that suppresses cryptic splicing through binding to deep introns, preserving transcriptome integrity. Long interspersed nuclear elements (LINEs) harbor large amounts of pseudo splice sites in introns. hnRNPM preferentially binds at intronic LINEs and represses LINE-containing pseudo splice site usage for cryptic splicing. Remarkably, a subgroup of the cryptic exons can form long dsRNAs through base-pairing of inverted Alu transposable elements scattered in between LINEs and trigger interferon immune response, a well-known antiviral defense mechanism. Notably, these interferon-associated pathways are found to be upregulated in hnRNPM-deficient tumors, which also exhibit elevated immune cell infiltration. These findings unveil hnRNPM as a guardian of transcriptome integrity. Targeting hnRNPM in tumors may be used to trigger an inflammatory immune response thereby boosting cancer surveillance.

genomics↗

bioRxiv2023

The AEGEAN-169 clade of bacterioplankton is synonymous with SAR11 subclade V (HIMB59) and metabolically distinct

Bacterioplankton of the SAR11 clade are the most abundant marine microorganisms and consist of numerous subclades spanning Order level divergence (Pelagibacterales). The assignment of the earliest diverging subclade V (a.k.a. HIMB59) to the Pelagibacterales is highly controversial, with multiple recent phylogenetic studies placing them completely separate from SAR11. Other than through phylogenomics, subclade V has not received detailed examination due to limited genomes from this group. Here, we assessed the ecogenomic characteristics of subclade V to better understand the role of this group in comparison to the Pelagibacterales. We used a new isolate genome, recently released single amplified genomes (SAGs) and metagenome-assembled genomes (MAGs), and previously established SAR11 genomes to perform a comprehensive comparative genomics analysis. We paired this analysis with recruitment of metagenomes spanning open ocean, coastal, and brackish systems. Phylogenomics, average amino acid identity, and 16S rRNA gene phylogeny indicate that SAR11 subclade V is synonymous with the ubiquitous AEGEAN-169 clade, and support the contention that this group represents a taxonomic Family. AEGEAN-169 shared many bulk genome qualities with SAR11, such as streamlining and low GC content, but genomes were generally larger. AEGEAN-169 had overlapping distributions with SAR11 but was metabolically distinct from SAR11 in its potential to transport and utilize a broader range of sugars as well as in transport of trace metals and thiamin. Thus, regardless of the ultimate phylogenetic placement of AEGEAN-169, these organisms have distinct metabolic capacities that likely allow them to differentiate their niche from canonical SAR11 taxa. IMPORTANCEOne goal of marine microbiologists is to uncover the roles various microorganisms are playing in biogeochemical cycles. Success in this endeavor relies on differentiating groups of microbes and circumscribing their relationships. An early-diverging group (subclade V) of the most abundant bacterioplankton, SAR11, has recently been proposed as a separate lineage that does not share a most recent common ancestor. But beyond phylogenetics, little has been done to evaluate how these organisms compare with SAR11. Our work leverages dozens of new genomes to demonstrate the similarities and differences between subclade V and SAR11. In our analysis, we also establish that subclade V is synonymous with a group of bacteria established from 16S rRNA gene sequences, AEGEAN-169. Subclade V/AEGEAN-169 has clear metabolic distinctions from SAR11 and their shared traits point to remarkable convergent evolution if they do not share a most recent common ancestor.

microbiology↗

bioRxiv2023

A complex three-dimensional microfluidic model that mimics the early stage events in the human atherosclerotic artery

BackgroundAtherosclerosis is a complex inflammatory vascular disease characterized by lipid and immune cells accumulation in the vessel wall, leading to lumen narrowing. Although several 3D in vitro microfluidic systems were previously described, a realistic reconstruction of the in vivo human atherosclerotic environment requires co-culture of different cell types arranged in atherosclerotic vessel-like structures with exposure to flow and circulating cells, creating challenges for disease modelling. In this study we developed a 3D tubular microfluidic model with quadruple coculture of human aortic smooth muscle cells (hAoSMCs), human umbilical cord vein endothelial cells (HUVECs) and foam cells to re-create a complex human atherosclerotic vessel in vitro to study the effect of flow and circulating immune cells. Methods & ResultsOur new co-culture protocol with BFP-labelled hAoSMCs, GFP-labelled HUVECs and THP-1 macrophages-derived, Dil-labelled Oxidized Low-Density Lipoprotein (Dil-Ox-LDL) foam cells in a fibrinogen-collagen-I based 3D extracellular matrix (ECM) resulted in vessels with an early lesion morphology, showing a layered vessel-like composition with an endothelium and media, with foam cells accumulating in the sub-endothelial space. Perfusion for 24 hours of atherosclerotic and "healthy" vessels (BFP hAoSMCs and GFP HUVECs without foam cells) showed that the layered wall composition remained stable. Perfusion with circulating THP-1 monocytes demonstrated cell extravasation into the atherosclerotic vessel wall and recruitment of THP-1 cells to the foam cell core. QPCR analysis revealed increased expression of atherosclerosis markers in the atherosclerotic vessels and adaptation in VSMCs migration to flow and the plaque microenvironment, compared to control vessels. ConclusionWe present a 3D tubular microfluidic model of a complex early atherosclerotic human vessel that can be exposed to flow and circulating THP-1 monocytes to study hemodynamic changes and immune cell recruitment under live confocal imaging. This novel atherosclerosis-on-a-chip model offers a humanized platform for in-depth mechanistic in vitro studies and drug testing.

cell biology↗

bioRxiv2023

ADO-MEDIATED SYNTHESIS OF TAURINE ALTERS THE CHROMATIN LANDSCAPE OF INGUINAL ADIPOSE TISSUE TO ENHANCE NON-SHIVERING THERMOGENESIS

Non-shivering thermogenesis (NST) has strong potential to combat obesity, however, a safe molecular approach to activate this process has not yet been identified. The sulfur amino acid taurine has the ability to safely activate NST and confer protection against obesity and metabolic disease in both mice and humans, but the mechanism of action is unknown. In this study, we discover that a suite of taurine biosynthetic enzymes, especially that of cysteamine dioxygenase (ADO), significantly increases in response to {beta}3 adrenergic signaling in inguinal tissues (IWAT) in order increase intracellular concentrations of taurine. We further show that ADO is critical for thermogenic mitochondrial function as its ablation in thermogenic adipocytes significantly reduces taurine levels which lead to declines in mitochondrial oxygen consumption rates. Finally, we demonstrate via assay for transposase-accessible chromatin with sequencing (ATAC-Seq) that taurine supplementation has the ability to remodel the chromatin landscape to increase the chromatin accessibility and transcription of genes, such as glucose-6-phosphate isomerase 1 (Gpi1), that are critical for NST. Taken together, our studies highlight a potential mechanism for taurine in the activation of NST that can be leveraged toward the treatment of obesity and metabolic disease.

cell biology↗

bioRxiv2022

TimiGP: inferring inter-cell functional interactions and clinical values in the tumor immune microenvironment through gene pairs

Determining how immune cells functionally interact in the tumor microenvironment and identifying their biological roles and clinical values are critical for understanding cancer progression and developing new therapeutic strategies. Here we introduce TimiGP, a computational method to infer inter-cell functional interaction networks and annotate the corresponding prognostic effect from bulk gene expression and survival statistics data. When applied to metastatic melanoma, TimiGP overcomes the prognostic bias caused by immune co-infiltration and identifies the prognostic value of immune cells consistent with their anti- or pro-tumor roles. It reveals the functional interaction network in which the interaction X[->]Y indicates a more positive impact of cell X than Y on survival. This network provides immunological insights to facilitate the development of prognostic models, as evidenced by our computational-friendly, biologically interpretable, independently validated models. By leveraging single-cell RNA-seq data for specific immune cell subsets, TimiGP has the flexibility to delineate the tumor microenvironment at different resolutions and is readily applicable to a wide range of cancer types.

bioinformatics↗

bioRxiv2022

Evaluating the analytical validity of mutation calling pipeline for tumor whole exome sequencing

Detecting somatic mutations from the patients tumor tissues has the clinical impacts in medical decision making. Library preparation methods, sequencing platforms, read alignment tools and variant calling algorithms are the major factors to influence the data analysis results. Understanding the performance of the tool combinations of the somatic variant calling pipelines has become an important issue in the use of the whole exome sequences (WES) analysis in clinical actions. In this study, we selected four state-of-the-art sequence aligners including BWA, Bowtie2, DRAGMAP, DRAGEN aligner (DragenA) and HISAT2. For the variant callers, we chose GATK Mutect2, Sentieon TNscope, DRAGEN caller (DragenC) and DeepVariant. The benchmarking tumor whole exome sequencing data released from the FDA-led Sequencing and Quality Control Phase 2 (SEQC2) consortium was applied as the true positive variants to evaluate the overall performance. Multiple combinations of the aligners and variant callers were used to assess the variation detection capability. We measured the recall, precision and F1-score for each combination in both single nucleotide variants (SNVs) and short insertions and deletions (InDels) variant detections. We also evaluated their performances in different variant allele frequencies (VAFs) and the base pair length. The results showed that the top recall, precision and F1-score in the SNVs detection were generated by the combinations of BWA+DragenC(0.9629), Bowtie2+TNscope(0.9957) and DRAGMAP+DragenC(0.9646), respectively. In the InDels detection, BWA+DragenC(0.9546), Hisat2+TNscope(0.7519) and DragenA+DragenC(0.8081) outperformed the other combinations in the recall, precision and F1-Score, respectively. In addition, we found that the variant callers could bias the variant calling results. Finally, although some combinations yielded high accuracies of variant detection, but some variants still could not be detected by these outperformed combinations. The results of this study provided the vital information that no single combination could achieve superior results in detecting all the variants of the benchmarking dataset. In conclusion, applying both merged-based and ensemble-based variants detection approaches is encouraged to further detect variants comprehensively.

bioinformatics↗

bioRxiv2022

Functional Decomposition of Metabolism allows a system-level quantification of fluxes and protein allocation towards specific metabolic functions

Quantifying the contribution of individual molecular components to complex cellular processes is a grand challenge in systems biology. Here we establish a general theoretical framework (Functional Decomposition of Metabolism, FDM) to quantify the contribution of every metabolic reaction to metabolic functions, e.g. the biosynthesis of metabolic building blocks such as amino acids. This allows us to obtain a plethora of results for E. coli growing in different conditions. A detailed quantification of energetic costs for biosynthesis and biomass growth on glucose shows that ATP generated during de novo biosynthesis of building blocks almost balances the ATP costs of peptide chain polymerization, the single largest energy expenditure for growing cells. This leaves the bulk of energy generated by fermentation and respiration (consuming 1/3 of the glucose intake) during aerobic growth unaccounted for. FDM also enabled the quantification of protein allocated towards each metabolic function, unveiling linear enzyme-flux relations for biosynthesis. These results led us to derive a function-based coarse-grained model to capture global protein allocation and overflow metabolism, without relying on curated pathway annotation or clustering of gene expression data.

systems biology↗

bioRxiv2022

A genome wide association study identifies a new variant associated with word reading fluency in Chinese children

Reading disability exhibited defects in different cognitive domains, including word reading fluency, word reading accuracy, phonological awareness, rapid automatized naming, and morphological awareness. To identify the genetic basis of Chinese reading disability, we conducted a genome wide association study (GWAS) of the cognitive traits related to Chinese reading disability in 2284 unrelated Chinese children. Among the traits analyzed in the present GWAS, we detected one genome wide significant association (p<5x10-8) on reading fluency for one SNP on 4p16.2, within EVC genes (rs6446395, p=7.55x10-10). Rs6446395 also showed significant association with word reading accuracy (p=3.39x10-4), phonological awareness (p=7.12x10-3), and rapid automatized naming (p=4.71x10-3), implying multiple effects of this variant. Gene-based analyses identified a gene to be associated with reading fluency at the genome-wide level. The eQTL data showed that rs6446395 affected EVC expression in the cerebellum. Our study discovered a new candidate susceptibility variant for reading ability and provide new insights into the genetics of development dyslexia in Chinese Children.

genetics↗

bioRxiv2022

A genome-wide association study identified new variants associated with mathematical abilities in Chinese children

Mathematical ability is moderately heritable, and it is a complex trait which can be evaluated in several different categories. A few genetic studies have been published on general mathematical ability. However, no genetic study focused on specific mathematical ability categories. In this study, we separately performed genome-wide association studies (GWASs) on 11 mathematical ability categories in 1 146 students from Chinese elementary schools. We identified 7 genome-wide significant single nucleotide polymorphisms (SNPs) with strong linkage disequilibrium (LD) among each other (all r2>0.8) associated with mathematical reasoning ability (top SNP: rs34034296, p=2.01x10-8, nearest gene: CUB and Sushi multiple domains 3, CSMD3). We replicated one SNP (rs133885) from 585 SNPs previously reported to be associated with general mathematical ability associated with division ability in our data (p=1.053x10-5). In the gene- and gene-set enrichment analysis by MAGMA, we found three significant enrichments of associations with three mathematical ability categories for three genes. We also observed four significant enrichments of associations with four mathematical ability categories for three gene sets. Our results suggest new candidate genetic loci for the genetics of mathematical ability.

genetics↗

bioRxiv2022

Chemical Imaging Reveals Diverse Functions of Tricarboxylic Acid Metabolites in Root Growth and Development

Understanding how plants grow is critical for agriculture and fundamental for illuminating principles of multicellular development 1. Here, we apply chemical mapping of the developing maize root using desorption electrospray ionization mass spectrometry imaging (DESI-MSI) 2. This technique reveals a range of small molecule distribution patterns across the gradient of stem cell differentiation in the root. To understand the developmental logic of these patterns, we examined tricarboxylic acid (TCA) cycle metabolites. In both Arabidopsis and maize, TCA metabolites are enriched in developmentally opposing regions, suggesting that stem-cell specific TCA metabolite localization may be conserved in evolutionarily divergent species. We find that these metabolites, particularly succinate, aconitate, citrate, and -ketoglutarate, control root development in diverse and distinct ways. Critically, the effects of metabolites on stem cell behavior can be independent of their canonical role in ATP production. These results present new insights into development and suggest practical means for controlling plant growth.

plant biology↗

bioRxiv2022

Characterizing control of memory CD8 T cell differentiation by BTB-ZF transcription factor Zbtb20

Members of the BTB-ZF transcription factor family regulate the immune system. Our lab identified that family member Zbtb20 contributes to the differentiation, recall responses and metabolism of CD8 T cells. Here, we report a characterization of the transcriptional and epigenetic signatures controlled by Zbtb20 at single-cell resolution during the effector and memory phases of the CD8 T cell response. Without Zbtb20, transcriptional programs associated with memory CD8 T cell formation were upregulated throughout the CD8 T response. A signature of open chromatin was associated with genes controlling T cell activation, consistent with the known impact on differentiation. Additionally, memory CD8 T cells lacking Zbtb20 were characterized by open chromatin regions with overrepresentation of AP-1 transcription factor motifs and elevated RNA- and protein-level expression of the corresponding AP-1 components. Finally, we describe motifs and genomic annotations from the DNA targets of Zbtb20 in CD8 T cells identified by CUT&RUN. Together, these data establish the transcriptional and epigenetic networks contributing to the control of CD8 T cell responses by Zbtb20.

immunology↗

bioRxiv2022

Ecophysiology and genomics of the brackish water adapted SAR11 subclade IIIa

The Order Pelagibacterales (SAR11) is the most abundant group of heterotrophic bacterioplankton in global oceans and comprises multiple subclades with unique spatiotemporal distributions. Subclade IIIa is the primary SAR11 group in brackish waters and shares a common ancestor with the dominant freshwater IIIb (LD12) subclade. Despite its dominance in brackish environments, subclade IIIa lacks systematic genomic or ecological studies. Here, we combine closed genomes from new IIIa isolates, new IIIa MAGS from San Francisco Bay (SFB), and 466 high-quality publicly available SAR11 genomes for the most comprehensive pangenomic study of subclade IIIa to date. Subclade IIIa represents a taxonomic family containing three genera (denoted as subgroups IIIa.1, IIIa.2, and IIIa.3) that had distinct ecological distributions related to salinity. The expansion of taxon selection within subclade IIIa also established previously noted metabolic differentiation in subclade IIIa compared to other SAR11 subclades such as glycine/serine prototrophy, mosaic glyoxylate shunt presence, and polyhydroxyalkanoate synthesis potential. Our analysis further shows metabolic flexibility among subgroups within IIIa. Additionally, we find that subclade IIIa.3 bridges the marine and freshwater clades based on its potential for compatible solute transport, iron utilization, and bicarbonate management potential. Pure culture experimentation validated differential salinity ranges in IIIa.1 and IIIa.3 and provided the first IIIa cell size and volume data. This study is an important step forward for understanding the genomic, ecological, and physiological differentiation of subclade IIIa and the overall evolutionary history of SAR11.

microbiology↗

bioRxiv2022

Mapping PTBP splicing in human brain identifies targets for therapeutic splice switching including SYNGAP1

Alternative splicing of neuronal genes is controlled in part by the coordinated action of the polypyrimidine tract binding proteins (PTBP1 and PTBP2). While PTBP1 is ubiquitously expressed, PTBP2 is predominantly neuronal, controlling the expression of such targets as DLG4, which encodes PSD95, a protein important in synaptic function whose deficiency causes neurodevelopmental disorders. Here, we fully define the PTBP2 footprint in the human transcriptome using both human brain tissue and neurons derived from human induced pluripotent stem cells (iPSC-neurons). We identify direct PTBP2 binding sites and define PTBP2-dependent alternative splicing events, finding novel targets such as STXBP1 and SYNGAP1, which are synaptic genes also associated with neurodevelopmental disorders. The resultant PTBP2 binding and splicing maps were used to test if PTBP2 binding could be manipulated to increase gene expression in PTBP-targeted genes that cause disease when haploinsufficient. We find that PTBP2 binding to SYNGAP1 mRNA promotes alternative splicing and non-sense mediated decay. Antisense oligonucleotides that disrupt PTBP binding sites on SYNGAP1 redirect splicing and increase gene and protein expression. Collectively, our data provide a comprehensive view of PTBP2-dependent alternative splicing in human neurons and human cerebral cortex, guiding the development of novel therapeutic tools that may benefit a range of neurodevelopmental disorders.

molecular biology↗

bioRxiv2022

Protein Lactylation and Metabolic Regulation of the Zoonotic Parasite Toxoplasma gondii

The biology of Toxoplasma gondii, the causative pathogen of one of the most wide-spread parasitic diseases remains poorly understood. Lactate, which is derived from glucose metabolic pathways, is considered to be not only an energy source in a variety of organisms including Toxoplasma gondii, but also a regulatory molecule that participates in gene activation and protein functioning. Lysine lactylation is a type of posttranslational modifications (PTMs) that was recently associated with chromatin remodeling, but lysine lactylation of histone and non-histone proteins has not yet been studied in T. gondii. To examine the prevalence and function of lactylation in T. gondii parasites, we mapped the lactylome of proliferating tachyzoite cells and found 1964 lactylation sites on 955 proteins in the T. gondii RH strain. The lactylated proteins were distributed in multiple subcellular compartments and were closely related to a wide variety of biological processes, including mRNA splicing, glycolysis, aminoacyl-tRNA biosynthesis, RNA transport, and multiple signaling pathways. We also performed chromatin immunoprecipitation sequencing (ChIP-seq) analysis with a lactylation specific antibody, the results revealed that histone H4K12la and H3K14la were enriched in the promoter and exon regions of Toxoplasma gondii genes associated with microtubule-based movement and cell invasion. We further confirmed the de-lactylase activity of histone deacetylase TgHDACs 2, 3, and 4, and that treatment with anti-histone acetyltransferase (TgMYST-A) antibodies profoundly reduced protein lactylation in the parasites. This study offers the first dataset of the global lactylation proteome and provides a basis for further dissection of the functional biology of Toxoplasma gondii.

microbiology↗

bioRxiv2022

Nipsnap1- A Regulatory Factor Required for Long-Term Maintenance of Non-Shivering Thermogenesis

The molecular activation of non-shivering thermogenesis (NST) has strong potential to combat obesity and metabolic disease. However, the mechanisms surrounding the maintenance of NST once it is fully activated, remain unexplored. Here, we present 4-Nitrophenylphosphatase Domain and Non-Neuronal SNAP25-Like 1 (Nipsnap1) as a critical regulator of long-term thermogenic maintenance in brown adipose tissue (BAT). Nipsnap1 localizes to the mitochondrial matrix and increases its transcript and protein levels in response to both chronic cold and {beta}3 adrenergic signaling. Through the generation of BAT-specific Nipsnap1 knockout mice (N1-KO), we show that these mice are unable to sustain activated energy expenditure and fail to protect their body temperature in the face of an extended cold challenge. Mechanistically, we demonstrate that Nipsnap1 integrates with lipid metabolism and BAT-specific ablation of Nipsnap1 leads to severe defects in {beta}-oxidation capacity. Our findings identify Nipsnap1 as a potent regulator of long-term NST maintenance.

molecular biology↗

bioRxiv2022

Prefusion-Stabilized Lassa Virus Trimer Identifies Neutralizing Nanobodies and Reveals an Apex-Situated Site of Vulnerability

Lassa virus (LASV) is responsible for 100,000-300,000 zoonotic infections annually and poses a threat to public health. Development of antibody-based therapeutics or vaccines has been challenging because neutralizing antibodies - even among Lassa hemorrhagic fever survivors - are generally of low titer, and the target of neutralizing antibodies, the trimeric glycoprotein complex (GPC), a type 1-fusion machine with GP1 and GP2 subunits, has been difficult to produce. Here, we use structure-based design to obtain a soluble LASV GPC by engineering an inter-protomer disulfide (R207GCGP1-L326CGP2) and appending the T4-fibritin trimerization domain. We verified the antigenicity of this prefusion-stabilized LASV GPC against a panel of human antibodies and used electron microscopy (EM) to confirm its trimeric association. We panned the prefusion-stabilized LASV GPC against single domain nanobody libraries and identified one of camel origin, which we named D5, which bound GPC with 27 nM affinity and neutralized the Josiah strain of LASV with an IC50 of 12 {micro}g/ml when formatted into a bivalent IgG2a context. The cryo-EM structure of a ternary complex of the D5 nanobody, the antigen-binding fragment of human antibody 8.11G, and LASV GPC revealed D5 to recognize a site-of-vulnerability at the trimer apex. The recognized site appeared to be specific to GPC lacking cleavage of between GP1 and GP2 subunits. Collectively, our findings suggest that GPC-cleavage intermediates may be targets for LASV neutralization and define an apex-situated site of vulnerability for vaccine development. SignificanceLassa virus (LASV) infection is expanding outside its traditionally endemic areas in West Africa, posing a biothreat to the world. LASV-neutralizing antibodies, moreover, have proven difficult to elicit. To gain insight into requirements for antibody-mediated neutralization of LASV, we developed a prefusion-stabilized LASV glycoprotein trimer (GPC), panned it against phage libraries comprised single-domain antibodies or nanobodies from shark and camel, and identified one, D5, which - when placed into bivalent IgG2a context - could neutralize LASV. Cryo-EM analysis revealed D5 to recognize a cleavage-dependent site-of-vulnerability at the trimer apex. We propose this apex-situated site to be an attractive target for LASV vaccine and therapeutic development.

immunology↗

bioRxiv2022

Single-cell transcriptomics reveal hyperacute cytokine and immune checkpoint axis in patients with poor neurological outcomes after cardiac arrest

Neurological injury is a major driver of mortality among patients hospitalized after cardiac arrest (CA). The early systemic inflammatory response after CA is associated with neurological injury and mortality but remains poorly defined. We determine the innate immune network induced by clinical CA at single-cell resolution. Immune cell states diverge as early as 6h post-CA between patients with good or poor neurological outcomes at hospital discharge. Nectin-2+ monocyte and Tim-3+ natural killer (NK) cell subpopulations associate with poor outcomes, and interactome analysis highlights their crosstalk via cytokines and immune checkpoints. Ex vivo studies on peripheral blood cells from CA patients demonstrate that immune checkpoints are a compensatory mechanism against inflammation after CA. IFN{gamma}/IL-10 induce Nectin-2 on monocytes; in a negative feedback loop, Nectin-2 suppresses IFN{gamma} production by NK cells. The initial hours after CA may represent a window for therapeutic intervention in the resolution of inflammation via immune checkpoints.

immunology↗

bioRxiv2022

SdiA, a Quorum Sensing Transcriptional Regulator, Enhanced the Drug Resistance of Cronobacter sakazakiiand Suppressed its Motility, Adhesion and Biofilm Formation

Cronobacter sakazakii is a common foodborne pathogen, and the mortality rate of its infection is as high as 40-80%. Quorum sensing is a regulation system of bacterial density-dependent multigene expression and is an important regulatory mechanism involved in adhesion, biofilm formation and virulence. C. sakazakii contains a QS signal molecular receiver, which is the LuxR receptor homolog SdiA, but its regulatory mechanism in C. sakazakii QS has not been defined. Here, we further determined the effect of SdiA on the QS system of C. sakazakii. The SdiA gene in C. sakazakii was knocked out by gene editing technology, and the biological characteristics of the {Delta}sdiA gene deletion strain of C. sakazakii were studied, followed by transcriptome analysis to elucidate its effects. The results suggested that SdiA enhanced the drug resistance of C. sakazakii but diminished its motility, adhesion and biofilm formation ability and had no effect on its growth. Transcriptome analysis showed that the deletion of the SdiA gene upregulated the expression levels of D-galactose operon genes (including dgoR, dgoK, dgoA, dgoD and dgoT) and flagella-related genes (FliA and FliC) in C. sakazakii and downregulated the expression levels of related genes in the type VI secretion system (VasK gene was downregulated by 1.53-fold) and ABC transport system (downregulated by 1.5-fold), indicating that SdiA was related to the physiological metabolism of C. sakazakii. The results of this study may be useful for clarifying the pathogenic mechanism of C. sakazakii and provide a theoretical basis for controlling bacterial infection. IMPORTANCECronobacter sakazakii, as an emerging opportunistic foodborne pathogen, was associated with sepsis, meningitis and necrotizing enterocolitis in neonates and infants, with a mortality rate of 40-80%. Quorum sensing plays an important regulatory role in the pathogenicity of C. sakazakii. Nevertheless, the regulatory mechanism of QS in C. sakazakii remains unknown. Here, we studied the QS transcriptional regulator SdiA of C. sakazakii. We revealed the regulatory mechanisms of SdiA in C. sakazakii cell adhesion, motility, biofilm formation and drug resistance. It was helpful to further explore the function of the SdiA gene, revealing the pathogenic mechanism of C. sakazakii. It will also provide a new target for therapeutic interventions targeting the pathogenicity of C. sakazakii and developing quorum-sensing inhibitors.

molecular biology↗