Mechanisms regulating peptidoglycan fragment production

调节肽聚糖片段产生的机制

• 批准号: 9896150
• 负责人: Joseph P Dillard
• 金额: $ 24.28万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-18 至 2022-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9896150
• 关键词: Affect; Agonist; Antisense RNA; Bacteria; Binding; Binding Sites; Biochemical Genetics; Cause of Death; Cell Death; Cell Wall; Cells; Cervical; Cervix Uteri; Cicatrix; Colony-forming units; Data; Defect; Disease; Ectopic Pregnancy; Endotoxins; Enzymes; Family; Female; Genes; Genetic Transcription; Growth; Helix-Turn-Helix Motifs; Heptoses; Host Defense; Human; In Vitro; Infection; Infertility; Inflammation; Inflammatory; Inflammatory Response; Knock-out; Lytic; Male urethral structure; Mammalian Oviducts; Measures; Messenger RNA; Metabolism; Methods; Molecular; Monitor; Muramidase; Mutation; Neisseria gonorrhoeae; Pain; Pattern recognition receptor; Pelvic Inflammatory Disease; Peptidoglycan; Phenotype; Physiologic pulse; Point Mutation; Process; Production; Promoter Regions; Proteins; Proteomics; Publishing; RNA; Recycling; Regulation; Regulon; Reporter; Rest; Risk; Symptoms; Testing; Tissues; Transcript; Transcriptional Activation; Transcriptional Regulation; Tube; VDAC1 gene; Vas deferens structure; chronic pelvic pain; dimer; human tissue; mutant; pathogen; pathogenic bacteria; peptidoglycan monomer; receptor; reproductive tract; response; small molecule libraries; survival outcome; transcriptome; transcriptome sequencing; transcriptomics

In symptomatic Neisseria gonorrhoeae infections, the inflammatory response to bacterial products results in tissue damage and pain in the human host. Peptidoglycan fragments released by the bacteria during growth represent a major pro-inflammatory factor in these infections. In particular, the peptidoglycan monomers are known to cause the death and sloughing of ciliated cells in human Fallopian tubes, and one of the peptidoglycan monomers is an agonist for the intracellular pattern-recognition receptor NOD1. Peptidoglycan dimers, also released by the bacteria, are processed by host lysozyme to create peptidoglycan fragments that are agonists for another peptidoglycan sensing receptor, NOD2. Peptidoglycan monomers and dimers are created by enzymes called lytic transglycosylases that function to degrade strands of peptidoglycan in the cell wall so that additional strands can be inserted to facilitate growth. The lytic transglycosylase LtgA produces nearly half of the peptidoglycan monomers released by gonococci and a vast majority of the peptidoglycan fragments that are recycled. LtgD produces the rest of the peptidoglycan monomers released by N. gonorrhoeae. Published transcriptomic studies indicate that ltgA is subject to transcriptional regulation, and this regulation may therefore affect gonococcal cell wall metabolism and release of pro-inflammatory peptidoglycan fragments. To understand how N. gonorrhoeae might control peptidoglycan fragment metabolism or release, we previously performed a proteomics analysis of five different mutants that have distinct defects in peptidoglycan fragment recycling. This analysis identified a putative transcriptional regulator, NGO1982, as significantly increased in three of the mutants. Deletion of ngo1982 resulted in 8-9 fold increased transcript levels for ltgA, suggesting that NGO1982 is a repressor of ltgA. Since published studies had identified MtrR as an activator of ltgA and the antisense RNA NgncR_246 as an additional regulator of ltgA expression, ltgA appears to be regulated by three distinct factors. To understand the consequences of this regulation for infection, we will: 1) Determine how growth in human tissues affect ltgA and ltgD regulation, and how this regulation affects cell wall-related phenotypes in infection, and 2) Use biochemical and genetic methods to determine the molecular mechanisms used by these regulators. Preliminary results demonstrate that ltgA is strongly regulated, and that regulatory factors can greatly increase or decrease peptidoglycan fragment release. In cervical infection, LtgA and LtgD levels decreased even as colony forming units increased. Significantly reducing or greatly increasing peptidoglycan fragment release may allow gonococci to cause asymptomatic infection in some tissues and highly inflammatory infections other tissues.

在有症状的淋病奈瑟菌感染中，对细菌产物的炎症反应导致 人体内的组织损伤和疼痛。细菌在生长过程中释放的肽聚糖片段 在这些感染中是一个主要的促炎因子。特别是，肽多聚糖的单体是 已知会导致人体输卵管中纤毛细胞的死亡和脱落，其中一种 肽聚糖单体是细胞内模式识别受体NOD1的激动剂。 同样由细菌释放的肽聚糖二聚体被宿主溶菌酶加工成 肽聚糖片段是另一种肽聚糖感受器NOD2的激动剂。 肽聚糖单体和二聚体是由称为裂解转糖基酶的酶产生的，这种酶的功能是 降解细胞壁中的肽链，以便可以插入额外的链以促进 成长。裂解的转糖基酶Ltd.A产生了近一半由 淋球菌和绝大多数被回收的肽聚糖碎片。Ltd.D生产其余的 淋病奈瑟氏菌释放的肽聚糖单体。已发表的转录学研究表明 LtgA受转录调控，因此这种调控可能会影响淋球菌细胞壁。 促炎症肽多聚糖片段的代谢和释放。 为了了解淋病奈瑟菌如何控制肽聚糖片段的新陈代谢或释放，我们 之前对五个不同的突变体进行了蛋白质组学分析，这些突变体在 肽多聚糖片段回收。这一分析确定了一个假定的转录调控因子NGO1982，即 在其中三个突变体中显著增加。Ngo1982的缺失导致转录本增加8-9倍 LtgA水平，提示NGO1982是LtgA的抑制子。自那以后发表的研究发现了mtrr 作为ltgA的激活剂和作为ltgA表达附加调节的反义RNA NgncR_246，ltgA 似乎受到三个不同因素的影响。 为了了解这一规定对感染的影响，我们将：1)确定 人类组织影响ltgA和ltgD调节，以及这种调节如何影响细胞壁相关表型。 感染，以及2)使用生化和遗传方法来确定 这些监管者。初步结果表明，ltgA受到很强的调控，并且调控因子 可以极大地增加或减少肽聚糖片段的释放。在宫颈感染中，LtgA和LtgD水平 即使在菌落形成单位增加的情况下也会减少。显著减少或大大增加 肽聚糖片段的释放可能会使淋球菌在某些组织和组织中引起无症状感染 高度炎症感染其他组织。