Lysine Acetylation in N. gonorrhoeae Quorum Sensing and Biofilm Formation

淋病奈瑟菌群体感应和生物膜形成中的赖氨酸乙酰化

• 批准号: 8705141
• 负责人: Michael A. Apicella
• 金额: $ 83.12万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-05 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8705141
• 关键词: Acetylation; Acetyltransferase; Affect; Antibiotic Therapy; Antimicrobial Resistance; Azithromycin; Bioinformatics; Biology; Bulla; Ceftriaxone; Cell division; Cell surface; Centers for Disease Control and Prevention (U.S.); Cervical; Cervix Uteri; Chromosomes; Collaborations; DNA; Data; Deacetylase; Development; Emergency Situation; Environment; Epithelial Cells; Exhibits; Gene Expression; Gene Expression Profile; Gene Proteins; Genes; Genetic Transcription; Gonorrhea; Grant; Growth; Health; Histone Deacetylase; Human; Human Papillomavirus; Immune; In Vitro; Infection; Laboratories; Lysine; Mass Spectrum Analysis; Membrane; Messenger RNA; Microbial Biofilms; Mind; Mutation; Neisseria gonorrhoeae; Nitric Oxide; Operon; Oral; Organism; Pathogenesis; Pathway interactions; Peptides; Peptidoglycan; Pharmaceutical Preparations; Phosphotransferases; Play; Post-Translational Protein Processing; Protein Family; Proteins; Proteome; Regulator Genes; Regulon; Reporting; Resistance; Role; Signal Transduction; Surface; System; Techniques; Testing; base; design; effective therapy; enzyme biosynthesis; mutant; novel; nuclease; pathogen; public health relevance; quorum sensing; respiratory; response; sensor; vaccine development

DESCRIPTION (provided by applicant): The Apicella, Gibson, Kirby and Wolfe labs have made the unique observations that Neisseria gonorrhoeae proteins undergo extensive N?-lysine acetylation and that quorum sensing coordinates this post-translational modification. Our studies have also shown that the degree of N-lysine acetylation effects survival in biofilms. Utilizing novel mass spectrometry techniques, Dr. Gibson's lab has shown that mutants of the luxS operon and ackA exhibit a significant increase in acetylated proteins. One such acetylated protein is the response regulator, MisR, which partners with its cognate sensor kinase MisS to form a two-component signal transduction system (TCS). Our data strongly suggest that MisRS senses the LuxS product. Our preliminary results suggest that the luxS operon functions as a classic quorum sensing system through MisRS and that this TCS controls genes that modulate cell division and N?-lysine acetylation. qRT-PCR studies with Ng grown in broth cultures and continuous flow biofilms demonstrate that the product of luxS significantly influences the transcription of a number of genes important in biofilm formation, including the gonococcal nuclease that remodels biofilm matrix, cell division proteins (minD and ftsZ), peptidoglycan biosynthesis enzymes (murD-murF), regulatory genes (metR), a SIR-2 family protein deacetylase and misRS. The hypothesis guiding this proposal is that quorum sensing and acetylation coordinately regulate biofilm formation to affect pathogenesis. To test this hypothesis, we propose the following 3 specific aims: 1) to determine the transcriptome of key Ng genes in the LuxS-regulated network of genes and the genes associated with reversible acetylation. These studies will be performed on mRNA isolated from organisms in the biofilm and planktonic states, 2) To understand the factors controlling Ng biofilm development, we will carry out comprehensive and targeted analyses designed to identify changes in the transcriptome, proteome and acetylome of Ng strain 1291 grown under defined in biofilms and planktonically and 3) Using N. gonorrhoeae grown over HPV E6/E7 transformed human cervical epithelial cells (HCEC), we will carry out an analogous study of the transcriptome, proteome and acetylome to interrogate the possible role of bacterial/eukaryotic signaling on N. gonorrhoeae biofilm development. These studies have the potential to open a new range of novel targets for antibiotics for the treatment of gonorrhea.

描述（由申请人提供）：Apicella，Gibson，Kirby和Wolfe Labs已经做出了独特的观察结果，即淋病的奈瑟氏菌蛋白质经历了广泛的N？ - 雷氏乙酰基乙酰化，而法定人的感觉会协调这种后经过的变质后修饰。我们的研究还表明，N-赖氨酸乙酰化的程度在生物膜中影响生存。吉布森博士的实验室利用新型的质谱技术，表明Luxs操纵子和ACKA的突变体显着增加乙酰化蛋白质。一种这样的乙酰化蛋白是响应调节剂MISR，它与其同源传感器激酶失误合作，形成了两组分组信号转导系统（TCS）。我们的数据强烈表明MISRS感知到Luxs产品。我们的初步结果表明，LuxS操纵子通过MISRS充当经典的法定人数传感系统，并且该TCS控制了调节细胞分裂和N？赖斯胺乙酰化的基因。 qRT-PCR studies with Ng grown in broth cultures and continuous flow biofilms demonstrate that the product of luxS significantly influences the transcription of a number of genes important in biofilm formation, including the gonococcal nuclease that remodels biofilm matrix, cell division proteins (minD and ftsZ), peptidoglycan biosynthesis enzymes (murD-murF), regulatory基因（METR），SIR-2家族蛋白脱乙酰基酶和Misrs。指导该建议的假设是，法定人数感应和乙酰化协同调节生物膜形成以影响发病机理。为了检验这一假设，我们提出以下3个特定目的：1）确定luxS调节基因网络中关键基因的转录组以及与可逆乙酰化相关的基因。这些研究将对从生物膜和浮游态中分离出的mRNA进行，2）为了了解控制NG生物膜发育的因素，我们将进行全面的靶向分析，旨在确定旨在确定NG菌株的转录组，蛋白质组和乙酰菌株的变化。 E6/E7转化了人宫颈上皮细胞（HCEC），我们将对转录组，蛋白质组和乙酰基组进行类似的研究，以询问细菌/真核信号传导对Gonorrhoeae n. gonorrhoeae biofilm发育的可能作用。这些研究有可能为淋病治疗的抗生素开辟新的新靶标。