Effect of Neisserial Porin in Immune Cell Apoptosis

奈瑟氏球菌孔蛋白对免疫细胞凋亡的影响

• 批准号: 6758513
• 负责人: LEE Mark WETZLER
• 金额: $ 33.42万
• 依托单位: BOSTON MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6758513
• 关键词: B lymphocyte; HeLa cells; Neisseriaceae; antigen antibody reaction; antigen presenting cell; apoptosis; bacteria infection mechanism; bacterial proteins; bacterial toxins; clone cells; confocal scanning microscopy; cytoprotection; electrophysiology; flow cytometry; gel electrophoresis; immunoprecipitation; intracellular transport; laboratory mouse; leukocyte activation /transformation; ligands; mitochondrial membrane; pore forming protein; protein protein interaction; protein transport; tissue /cell culture; western blottings

Neisserial porins (gonococcal Protein I [PIA or PIB] or meningococcal class 1 [PorA], 2 or 3 [PorB] proteins) are the major protein constituents of the Neisserial outer membrane. We have previously demonstrated that the Neisserial porins activate B lymphocytes, increasing the surface expression of the costimulatory ligand B7-2, which improves the B cell's ability to "costimulate" T lymphocytes. The induction of B7-2 expression is directly related to the porins' adjuvant ability. During the course of this work, we have discovered that the porins, when activating B cells, decreased the susceptibility of these B cells to activation induced cell death (AICD) or apoptosis. We found that the anti-apoptotic effect is due to a direct interaction of the porin with mitochondria (a central control point for apoptotic cell death). Immunoprecipitation experiments revealed that PorB interacts with the mitochondrial porin, VDAC. We hypothesize that this VDAC-Neisserial porin protein-protein interaction, which is similar to that of VDAC with the anti- apoptotic molecule Bcl-2, is responsible for the anti-apoptotic effect of the porins. This interaction results in an enhancement of cell survival and continued activation of B cells, which would potentiate the B cells' involvement in specific immune responses. Therefore, it is possible that this anti-apoptotic effect is a potential additional mechanism of the porins' immunopotentiating ability. This proposal will investigate the effect of Neisserial porins on the susceptibility of B cells, epithelial cells and other cell types to apoptosis. The basic aims include 1) examining the interaction of Neisserial porin with mitochondria and mitochondrial VDAC, 2) determining if alterations in cell culture conditions (especially serum withdrawal) can alter the effect of porins on apoptosis and whether this could be related to changes in cellular ATP levels and 3) determining the effect of intact Neisserial organisms on apoptosis and whether porin might play a role in this effect. These studies will yield significant findings that could both elucidate the immune stimulatory effect of Por (and potentially other bacterial components), allowing for the better use of Por as a vaccine adjuvant. In addition, these results can have significant implications in Neisserial pathogenesis, especially since we have new preliminary data that intact live meningococci can also protect cells from apoptosis.

奈瑟球菌孔蛋白（淋球菌蛋白 I [PIA 或 PIB] 或脑膜炎球菌 1 类 [PorA]、2 或 3 [PorB] 蛋白）是奈瑟球菌外膜的主要蛋白质成分。 我们之前已经证明奈瑟菌孔蛋白激活B淋巴细胞，增加共刺激配体B7-2的表面表达，从而提高B细胞“共刺激”T淋巴细胞的能力。 B7-2表达的诱导与孔蛋白的佐剂能力直接相关。 在这项工作的过程中，我们发现孔蛋白在激活 B 细胞时降低了这些 B 细胞对激活诱导细胞死亡 (AICD) 或细胞凋亡的敏感性。 我们发现抗凋亡作用是由于孔蛋白与线粒体（细胞凋亡的中心控制点）的直接相互作用所致。 免疫沉淀实验表明，PorB 与线粒体孔蛋白 VDAC 相互作用。 我们假设这种 VDAC-奈瑟氏球菌孔蛋白-蛋白质相互作用（类似于 VDAC 与抗凋亡分子 Bcl-2 的相互作用）负责孔蛋白的抗凋亡作用。 这种相互作用会增强细胞存活率并持续激活 B 细胞，从而增强 B 细胞参与特异性免疫反应。因此，这种抗凋亡作用可能是孔蛋白免疫增强能力的潜在附加机制。 该提案将研究奈瑟菌孔蛋白对 B 细胞、上皮细胞和其他细胞类型凋亡敏感性的影响。 基本目标包括 1) 检查奈瑟氏球菌孔蛋白与线粒体和线粒体 VDAC 的相互作用，2) 确定细胞培养条件的改变（尤其是血清撤出）是否可以改变孔蛋白对细胞凋亡的影响，以及这是否可能与细胞 ATP 水平的变化有关，3) 确定完整奈瑟球菌生物体对细胞凋亡的影响以及孔蛋白是否可能在这种影响中发挥作用。 这些研究将产生重要的发现，可以阐明 Por（以及可能的其他细菌成分）的免疫刺激作用，从而更好地使用 Por 作为疫苗佐剂。 此外，这些结果可能对奈瑟氏球菌发病机制产生重大影响，特别是因为我们有新的初步数据表明完整的活脑膜炎球菌也可以保护细胞免于凋亡。