Molecular Assembly on the Cell Surface of Actinomyces

放线菌细胞表面的分子组装

• 批准号: 8911399
• 负责人: Hung Ton-That
• 金额: $ 1.18万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-02-19 至 2017-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8911399
• 关键词: Abnormal Cell; Actinomyces; Adherence; Adhesives; Anti-Infective Agents; Arginine; Bacterial Adhesins; Binding; Cell Communication; Cell Wall; Cell Wall Alteration; Cell surface; Cells; Complex; Defect; Dental; Dental Enamel; Dental Plaque; Development; Disease; Economic Burden; Erythrocytes; Gene Expression Profile; Genes; Genetic; Gingivitis; Goals; Gram-Positive Bacteria; Grant; Growth; Health; Homeostasis; Housekeeping; Human; Immunoglobulin G; Lead; Maps; Mediating; Membrane; Membrane Proteins; Microbial Biofilms; Molecular Analysis; Morphology; Mutagenesis; Pathogenesis; Pathway interactions; Peptidyltransferase; Periodontitis; Pharmaceutical Preparations; Phenotype; Physiological; Play; Polymers; Polysaccharides; Population; Post-Translational Protein Processing; Proteins; Regulation; Role; Signal Transduction; Sorting - Cell Movement; Stress; Structure; Surface; Technology; Testing; Tooth Diseases; Trans-Activators; Twin Multiple Birth; Vaccines; base; cell envelope; cell growth; cellular targeting; disulfide bond; effective therapy; fimbria; fimbrillin; fitness; inhibitor/antagonist; man; microbial community; molecular assembly/self assembly; novel; oral bacteria; oral biofilm; oral streptococci; polymerization; prevent; protein B; receptor; salivary proline-rich proteins; sortase; tooth surface; type 1 fimbriae

DESCRIPTION (provided by applicant): Dental plaque represents one of the most complex microbial communities or biofilms known to afflict man. Oral biofilm-related diseases - dental carries, gingivitis and periodontitis - devastate a large human population and continue to pose a huge economic burden due to the lack of effective therapies. The long-term goal of this project is to elucidate the basic mechanisms of oral biofilm development and identify key players that may be attractive targets for developing drugs and vaccines. The development of dental plaque begins with the attachment of early bacterial colonizers to the tooth enamel, generating an adhesive matrix that then attracts the intermediate and late colonizers. Actinomyces is a key early colonizer that plays a prominent role in biofilm development by virtue of its ability to directly interact not only with the tooth surface but also with a number of both early and intermediate colonizers. Therefore, our studies have focused on dissecting the adhesive principles, i.e. fimbriae and other surface proteins, specifically involved in these interactions ad the mechanism of their assembly on the bacterial surface. During the past grant period, we succeeded in developing a facile new gene disruption technology for Actinomyces oris, and through it, identified the key components of two distinct fimbriae that are pivotal in the aforementioned cell-cell interactions. We showed that the tip fimbrillin FimQ serves dual functions, facilitating the assembly of type 1 fimbriae and directly mediating bacterial adherence to salivary proline-rich proteins known to coat the tooth surface. In contrast, the shaft fimbrilli FimA of the type 2 fimbriae mediates the receptor polysaccharide-dependent coaggregation with oral streptococci, adherence to erythrocytes and biofilm development. Structural studies revealed two adhesive IgG-like modules of FimA essential for its multivalent functions. Significantly, we showed that polymerization of these fimbrillins into fimbrial polymers requires their cognate fimbriae-specific sortase, a conserved transpeptidase in Gram-positive bacteria. The resulting polymers are anchored to the cell wall by the housekeeping sortase SrtA, which is also essential for the cell wall anchoring of many surface proteins with a cell wall sorting signal One of these, AcaF, is found to play a significant role in bacterial coaggregation. Most importantly, we discovered that inactivation of SrtA greatly perturbs bacterial morphology accompanied with abnormal cell wall and septa. Thus, the sortase machinery is a key player of bacterial pathogenesis and fitness in A. oris. Driven by these major advancements and several new hypotheses, the continuation proposal has three major aims: (1) Uncover the physiological function and regulation of sortase SrtA in cell surface homeostasis, (2) Dissect the molecular interactions of fimbrillins and non-fimbrial surface proteins with various cellular receptors, (3) Further elucidate the fundamental mechanisms of fimbrial assembly in Actinomyces, delineate the distinct mechanism of fimbrial assembly mediated by the tip fimbrillin FimQ, and identify trans- acting factors required for sortase-mediated fimbrial assembly.

描述（由申请人提供）：牙菌斑代表了已知折磨人类的最复杂的微生物群落或生物膜之一。口腔生物膜相关疾病-牙携带、牙龈炎和牙周炎-困扰着大量人口，并且由于缺乏有效的治疗方法而继续造成巨大的经济负担。该项目的长期目标是阐明口腔生物膜发展的基本机制，并确定可能成为开发药物和疫苗的有吸引力的目标的关键参与者。牙菌斑的形成始于早期细菌定植者附着在牙釉质上，产生一种粘附基质，然后吸引中期和晚期定植者。放线菌是一种关键的早期定殖菌，由于其不仅与牙齿表面而且与许多早期和中期定殖菌直接相互作用的能力，其在生物膜发育中起着突出的作用。因此，我们的研究集中在解剖粘附原则，即菌毛和其他表面蛋白，特别是参与这些相互作用及其在细菌表面组装的机制。在过去的资助期间，我们成功地开发了一种简便的口腔放线菌新基因破坏技术，并通过它确定了两种不同菌毛的关键成分，这两种菌毛在上述细胞间相互作用中起着关键作用。我们发现，尖端菌毛蛋白FimQ具有双重功能，促进1型菌毛的组装，并直接介导细菌粘附到已知覆盖牙齿表面的富含脯氨酸的唾液蛋白。相反，2型菌毛的轴菌毛FimA介导与口腔链球菌的受体多糖依赖性共聚集、粘附于红细胞和生物膜发育。结构研究揭示了FimA的两个粘附IgG样模块对其多价功能至关重要。值得注意的是，我们发现这些菌毛蛋白聚合成菌毛聚合物需要它们的同源菌毛特异性分选酶，一种革兰氏阳性菌中保守的转肽酶。由此产生的聚合物通过管家分选酶SrtA锚定到细胞壁上，SrtA也是许多具有细胞壁分选信号的表面蛋白质的细胞壁锚定所必需的。最重要的是，我们发现SrtA的失活极大地扰乱了细菌的形态，伴随着异常的细胞壁和隔膜。因此，分选酶机制是A.奥里斯。在这些重大进展和几个新假设的推动下，延续提案有三个主要目标：（1）揭示了分选酶SrtA在细胞表面稳态中的生理功能和调控;（2）剖析了菌毛蛋白和非菌毛表面蛋白与各种细胞受体的分子相互作用;（3）进一步阐明了放线菌菌毛组装的基本机制;描述由尖端菌毛蛋白FimQ介导的菌毛组装的独特机制，并鉴定分选酶介导的菌毛组装所需的反式作用因子。