Functional assessment of TprC/D and TprK proteins of syphilis causing spirochete, Treponema pallidum

科学抽象 梅毒的全球流行率估计为3600万例，新感染人数超过1100万 每年在世界各地。仅在美国，传染性梅毒病例的数量就稳步上升， 自2000年以来有所增加，表明这种疾病仍然是一个公共卫生问题，特别是考虑到它 会导致严重的神经和心血管后遗症了解外膜的生物学功能 膜蛋白（OMP）的梅毒剂，梅毒螺旋体亚种。苍白球（T. pallidum），是 梅毒发病机制研究的最大挑战。功能表征的进展 由于缺乏一个独立的、纯培养的T.苍白球的遗传难治性 病原体，需要使用家兔进行细菌繁殖，T.外苍白球 膜，以及这种细胞区室的脆弱性。尽管有如此多的限制， T.苍白球外膜蛋白可以大大提高我们对这些毒力因子在梅毒中的作用的理解 发病机制，甚至提供了新的方法，成功地控制梅毒的重要线索，如 开发保护性疫苗12个推测的OMPs/毒力因子属于T.苍白球重复序列 (Tpr)家族，其是一组预测与密螺旋体同源的高免疫原性蛋白质 denticola（T.主要鞘蛋白（Msp）是一种具有孔蛋白和粘附蛋白性质的表面毒力因子。 到目前为止，关于各种激素的功能，实验证据有限或没有。基于 与Msp的同源性，Tcb的结构模型和我们的初步数据，我们假设几个Tcb 也具有作为粘附素和孔蛋白转运蛋白的双重作用。T.梅毒螺旋体和伯氏疏螺旋体， 莱姆病，是生理和结构相关的螺旋体。与T.苍白球、B. B. Burgdorferi可以是 基因操作和转化以表达所选的TbR，就好像它们是其OM的成分一样。的 我们的初步数据表明，当TprD 2和TprD 2的水平高于TprD 1时， TprK在非感染性和粘附性差的B的表面上表达。burgdorferi菌株，这些蛋白质 增强对胶质瘤和上皮细胞的粘附，并促进氨基酸和肽的摄取。在这里，我们计划 为了进一步验证我们的假设，并建立这些重要的毒力因子的功能， 尝试了B。burgdorferi和T.齿垢MSP突变体替代系统。我们会进行以下研究 为了验证我们的假设：（1）确定T.苍白球TprC、D2和K蛋白介导与宿主的附着 通过识别每种细胞类型上的特异性宿主受体/ECM组分，以差异化方式识别细胞，以及 (2)确定T的作用。苍白球TprC、D2和K蛋白作为参与营养转运的通道。 意义：本研究将有助于更好地了解T.苍白球粘连 宿主成分，这是先天性传播和感染期间神经侵入的关键步骤， 提供了重要的信息，以进一步支持这些抗原在未来作为候选疫苗的使用。

SCIENTIFIC ABSTRACT Syphilis has an estimated global prevalence of 36M cases, with more than 11M new infections occurring per year around the world. In the United States alone, the number of cases of infectious syphilis has steadily increased since 2000, indicating that this disease is still a public health concern, particularly considering that it can lead to serious neurological and cardiovascular sequelae. Understanding the biological function of the outer membrane proteins (OMPs) of the syphilis agent, Treponema pallidum subsp. pallidum (T. pallidum), is one of the greatest challenges in the study of syphilis pathogenesis. Progress towards functional characterization of OMPs is hindered by the lack of an independent, pure culture system for T. pallidum, genetic intractability of the pathogen, the need to use of rabbits for bacterial propagation, the paucity of OMPs in T. pallidum outer membrane, and the fragility of this cellular compartment. Despite so many limitations, undertaking the study of T. pallidum OMPs can greatly enhance our understanding of the role of these virulence factors in syphilis pathogenesis and even provide important clues on new approaches to successfully control syphilis, such as by developing a protective vaccine. Twelve of putative OMPs/virulence factors belong to the T. pallidum repeat (Tpr) family, which are a group of highly immunogenic proteins predicted to be homologous to the Treponema denticola (T. denticola) major sheath protein (Msp), a surface virulence factor with porin and adhesin properties. Limited or no experimental evidence is available until now regarding functions of various Tprs. Based on homology to Msp, structural models for the Tprs and our preliminary data, we hypothesize that several Tprs also have dual roles as adhesins and porin transporters. T. pallidum and Borrelia burgdorferi, which causes Lyme disease, are physiologically and structurally related spirochetes. Unlike T. pallidum, B. burgdorferi can be genetically manipulated and transformed to express selected Tprs as if they were constituents of its OM. The validity of the surrogate system to study Tprs is supported by our preliminary data showing that when TprD2 and TprK are expressed on the surface of a non-infectious and poorly adherent B. burgdorferi strain, these proteins enhance adhesion to Glioma and epithelial cells and facilitate amino acids and peptides uptake. Here, we plan to further test our hypothesis and establish function of these important virulence factors by using both our well- tried B. burgdorferi, as well as the T. denticola msp mutant surrogate systems. We will conduct following studies to test our hypothesis: (1) Determine whether T. pallidum TprC, D2, and K proteins mediate attachment to host cells in differential manner by recognizing the specific host receptors/ECM components on each cell type, and (2) Determine the role of T. pallidum TprC, D2, and K proteins as channels involved in nutrient transport. Significance: Our studies will lead to a better understanding of the molecular basis of T. pallidum adhesion to host components, which is a pivotal step in congenital transmission and neuroinvasion during infection and will provide important information to further support the use of these antigens as vaccine candidates in the future.