IMMUNODOMINANT STRESS PROTEINS OF P. GINGIVALIS

牙龈卟啉单胞菌的免疫显性应激蛋白

• 批准号: 6900249
• 负责人: DENNIS E LOPATIN
• 金额: $ 31.74万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-04-01 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6900249
• 关键词: Bacteroides gingivalis; bacterial antigens; bacterial genetics; bacterial proteins; bactericidal immunity; disease /disorder model; gel electrophoresis; gene expression; genetic strain; immune tolerance /unresponsiveness; immunoelectron microscopy; immunologic assay /test; immunoprecipitation; laboratory mouse; laboratory rabbit; monoclonal antibody; mutant; nucleic acid sequence; periodontium disorder; polymerase chain reaction; protease inhibitor; protein sequence; protein structure function; stress proteins; virulence

Studies performed in our laboratory implicate the Porphyromonas gingivalis HtpG stress protein, the prokaryotic homologue of Hsp90, in the etiology of periodontal disease. We have reported that elevated levels of anti-Hsp90 antibodies, concomitant with P. gingivalis colonization, are associated with periodontal health. Transcription of HtpG message was also found to be upregulated 7-10-fold in P. gingivalis obtained from diseased subgingival plaque. There is a precedence for Hsp90 homologues contributing to pathogenicity of other microorganisms. Immunity to a single Hsp90 epitope of Candida albicans has been demonstrated to confer protection against systemic candidiasis. Studies performed by our laboratory have revealed that P. gingivalis HtpG has a significant degree of homology with human Hsp90, but remains clearly distinct from other HtpG proteins due to its unique C-terminal region. We have found that HtpG is localized to P. gingivalis membranes and extracellular vesicles, and that it cross-reacts with other prokaryotic and eukaryotic Hsp90 homologues. Our findings suggest that HtpG is readily accessible to participate in host cellular invasion processes, as well as to interfere with normal host cell functions one P. gingivalis enters the host cytoplasmic compartment. Transfection of KB cells with the P. gingivalis htpG gene stimulates IL-8 production by these cells. This application proposes to extend our investigations into the role that molecular mimicry by HtpG plays in the pathogenicity of P. gingivalis. Previous studies of other pathogenic microorganisms which appear to use the Hsp90 homologue as a virulence factor have been purely descriptive. Our application is unique in that while will propose to evaluate the role of HtpG in adherence and invasion mechanisms, we also propose to elucidate novel pathogenic mechanism(s) by which microorganisms such as P. gingivalis utilize molecular mimicry to disrupt normal eukaryotic cell function(s). Since the most clearly defined eukaryotic Hsp90-mediated mechanisms involved signal transduction pathways, these will be the primary foci of our investigations. The hypothesis to be tested in this study is: 1) HtpG plays a role in adherence and invasion of host cells; and 2) once internalized, signal transduction mechanisms mediated by Hsp90/TRAP1 within eukaryotic cells are disrupted by the HtpG of P. gingivalis through molecular mimicry. This leads to disruption of normal inflammatory cytokine responses to microbial invasion by P. gingivalis and other oral microorganisms.

我们实验室进行的研究表明，牙龈卟啉单胞菌HtpG应激蛋白，Hsp90的原核同源物，与牙周病的病因学有关。我们已经报道，高水平的抗Hsp90抗体，伴随着牙龈假单胞菌的定植，与牙周健康有关。从患病的牙龈下菌斑中获得的牙龈假单胞菌中，HtpG信息的转录也被上调了7-10倍。Hsp90同系物对其他微生物的致病作用有先例。对白色念珠菌的单一Hsp90表位的免疫已被证明对系统性念珠菌病具有保护作用。我们实验室的研究表明，牙龈假单胞菌HtpG与人类Hsp90有很大程度的同源性，但由于其独特的C-末端区域，仍明显有别于其他HtpG蛋白。我们发现HtpG定位于牙龈假单胞菌的细胞膜和胞外小泡，并与其他原核和真核Hsp90同源物发生交叉反应。我们的发现表明，HtpG很容易参与宿主细胞的入侵过程，并在牙龈假单胞菌进入宿主细胞质时干扰正常的宿主细胞功能。将牙龈假单胞菌htpG基因导入KB细胞，可刺激这些细胞产生IL-8。这项应用建议扩大我们对HtpG分子模拟在牙龈假单胞菌致病性中所起作用的研究。以前对其他似乎使用Hsp90同源物作为毒力因子的病原微生物的研究都是纯粹的描述性的。我们的应用是独特的，虽然我们将建议评估HtpG在黏附和侵袭机制中的作用，但我们也建议阐明新的致病机制(S)，通过该机制，牙龈假单胞菌等微生物利用分子模拟来破坏正常的真核细胞功能(S)。由于最明确定义的真核细胞Hsp90介导的机制涉及信号转导途径，这些将是我们研究的主要焦点。本研究要验证的假设是：1)HtpG在宿主细胞的黏附和侵袭中发挥作用；2)一旦内化，真核细胞内Hsp90/TRAP1介导的信号转导机制就会被牙龈假单胞菌的HtpG通过分子模拟来破坏。这会导致正常的炎性细胞因子对牙龈假单胞菌和其他口腔微生物入侵的反应中断。