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The competence regulon in Streptococcus mutans biofilms

变形链球菌生物膜的能力调节子

基本信息

批准号：
7995205
负责人：
Dennis G. Cvitkovitch
金额：
$ 47.37万
依托单位：
UNIVERSITY OF TORONTO
依托单位国家：
美国
项目类别：
财政年份：
1999
资助国家：
美国
起止时间：
1999-09-01 至 2012-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7995205
关键词：
Acids Address Affect Amino Acid Sequence Architecture Area Attenuated Bacteria Base Pairing Binding Binding Sites Biochemical Biological Assay Cell Count Cell Density Cell Separation Cell Survival Cells Competence Complex Consensus Sequence DNA DNA Binding DNA Footprint DNA-Binding Proteins Dental Plaque Dental caries Detection Dietary Carbohydrates Environment Gene Expression Gene Expression Profile Gene Fusion Gene Proteins Genes Genetic Genetic Transcription Genome Goals Growth In Vitro Individual LacZ Genes Laser Scanning Confocal Microscopy Lead Measures Mediating Microarray Analysis Microbial Biofilms Molecular Profiling Mutagenesis Mutate Operator Regions Pathway interactions Peptide Receptor Peptide Signal Sequences Peptides Phenotype Phosphorylation Physiological Process Promoter Regions Regulon Relative (related person)Reporter Genes Role Scanning Electron Microscopy Signal Transduction Signal Transduction Inhibition Signal Transduction Pathway Site Streptococcus mutans System Techniques Testing Time base design extracellular gel mobility shift assay in vitro Assay inhibitor/antagonist insight mRNA Expression mutant novel pathogen peptide analog promoter protein protein interaction protein-histidine kinase quorum sensing receptor research study response stress tolerance uptake

项目摘要

Streptococcus mutans resides in the biofilm of dental plaque where it produces acid from dietary carbohydrate to cause caries. S. mutans uses a quorum sensing signaling system in biofilms to activate genetic competence, acid tolerance, and influence biofilm architecture. This proposal addresses mechanisms by which cell-cell and environmental signals activate this 'biofilm phenotype'. S. mutans quorum sensing system is encoded by the comCDE genes that encode a competence-stimulating peptide (CSP) precursor, a histidine kinase and a response regulator. We expect to answer 1)Which phenotypic changes does CSP invoke through its various putative receptors and regulators and how do these changes facilitate optimized growth/survival in a biofilm environment? 2) How are these pathways regulated? 3) Can we inhibit these pathways to attenuate the biofilm phenotype of S. mutans? DMA microarray analysis will be used to identify genes and proteins that are activated by CSP. A number of mutants defective in various components of the system will be used to elucidate these complex signaling networks. Genes encoding products with altered expression in a biofilm context will be cloned and mutated using a novel allelic exchange technique; the mutants' genetic competence and biofilm forming abilities will be tested. Cell segregation and the 3D architecture of the mutant biofilms will be assessed by SEM and Confocal Scanning Laser Microscopy (CSLM). Reporter gene fusions (gfp, lacZ and luc) constructed in selected genes will be used to measure gene expression temporally in response to CSP using fluorimetric and luminetric analysis and CSLM to give insight into how the CSP signaling cascade functions to influence biofilm phenotype in real time. CSP analogs will be tested for their inhibition of the signal transduction process. The regulatory processes involved at modulating this network will be investigated using DNA-protein binding assays, protein-protein interactions and phosphorylation assays. These experiments will give insight into the signaling mechanisms used by S. mutans and potentially other biofilm-forming pathogens for optimal survival in biofilms and will hopefully lead to the design of means to control them.

变形链球菌存在于牙菌斑的生物膜中，它从饮食中产生酸。碳水化合物引起龋齿。S.变形杆菌利用生物膜中的群体感应信号系统来激活遗传能力、耐酸性和影响生物膜结构。该提案涉及细胞-细胞和环境信号激活这种“生物膜表型”的机制。S.变异法定人数传感系统由comCDE基因编码，comCDE基因编码感受态刺激肽（CSP），前体、组氨酸激酶和应答调节剂。我们期望回答1）哪些表型变化 CSP是否通过其各种假定的受体和调节剂调用，以及这些变化如何促进在生物膜环境中优化生长/存活？2)这些途径是如何调节的？3)我们能抑制这些途径减弱了S.变种人？DMA微阵列分析将用于识别由CSP激活的基因和蛋白质。许多突变体在各种成分上都有缺陷将用于阐明这些复杂的信号网络。基因编码产物，将使用新的等位基因交换技术克隆和突变生物膜环境中改变的表达; 将测试突变体的遗传能力和生物膜形成能力。细胞分离和3D 突变生物膜的结构将通过SEM和共聚焦扫描激光显微镜进行评估（CSLM）。在选定的基因中构建的报告基因融合体（gfp、lacZ和luc）将用于测量使用荧光和发光分析以及CSLM，深入了解CSP信号级联如何发挥作用以真实的时间影响生物膜表型。CSP 将测试类似物对信号转导过程的抑制。规管程序将使用DNA-蛋白质结合测定、蛋白质-蛋白质结合测定和蛋白质-蛋白质结合测定来研究参与调节该网络的分子。相互作用和磷酸化测定。这些实验将使我们深入了解使用S。变形菌和潜在的其他生物膜形成病原体，以在生物膜中获得最佳存活，希望能设计出控制它们的方法。

项目成果

期刊论文数量（34）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Cell death in Streptococcus mutans biofilms: a link between CSP and extracellular DNA.

DOI：
10.1111/j.1574-6968.2009.01758.x
发表时间：
2009-10
期刊：
FEMS microbiology letters
影响因子：
2.1
作者：
Perry JA;Cvitkovitch DG;Lévesque CM
通讯作者：
Lévesque CM

A novel function for the competence inducing peptide, XIP, as a cell death effector of Streptococcus mutans.

诱导肽XIP作为链球菌突变的细胞死亡效应的能力的新功能。

DOI：
10.1111/j.1574-6968.2012.02660.x
发表时间：
2012-11
期刊：
FEMS microbiology letters
影响因子：
2.1
作者：
Wenderska IB;Lukenda N;Cordova M;Magarvey N;Cvitkovitch DG;Senadheera DB
通讯作者：
Senadheera DB

An intimate link: two-component signal transduction systems and metal transport systems in bacteria.

DOI：
10.2217/fmb.14.87
发表时间：
2014
期刊：
Future microbiology
影响因子：
3.1
作者：
Singh K;Senadheera DB;Cvitkovitch DG
通讯作者：
Cvitkovitch DG

Involvement of Streptococcus mutans regulator RR11 in oxidative stress response during biofilm growth and in the development of genetic competence.

DOI：
10.1111/j.1472-765x.2008.02455.x
发表时间：
2008-11
期刊：
Letters in applied microbiology
影响因子：
2.4
作者：
Perry JA;Lévesque CM;Suntharaligam P;Mair RW;Bu M;Cline RT;Peterson SN;Cvitkovitch DG
通讯作者：
Cvitkovitch DG

Acid tolerance mechanisms utilized by Streptococcus mutans.