CRASP-mediated Serum Resistance by Borrelia burgdorferi

 描述（由申请方提供）：莱姆病由伯氏疏螺旋体螺旋体引起，是美国B最常见的病媒传播疾病。伯氏菌可以感染蜱叮咬部位的皮肤，并通过血液传播到不同的组织，这表明该细菌 可以抵抗血液的防御补体系统是一种血流防御系统，通过三种不同的途径激活。该系统活化的关键步骤是C3转化酶C4 b2 a或C3 bBb的形成，其催化导致炎症、调理作用和病原体裂解的事件。补体激活后潜在的宿主损伤需要通过结合补体组分并促进其降解的血清补体调节剂进行严格调节。例如，补体调节因子H（FH）和C4 BP分别调节C3 bBb和C4 b2 a的形成。我们最近证明了B。Burgdorferi产生DbpA，DbpA是一种通过结合硫酸皮肤素和核心蛋白聚糖促进组织定殖的表面蛋白，也通过结合C4 BP促进菌血症。A B。产生DbpA-I156 A（一种C4 BP结合缺陷的点突变体）的伯氏菌株显示出菌血症和联合定殖的延迟。此外，B. burgdorferi还产生与补体调节因子结合的CRASP（补体调节因子获得表面蛋白）。旁系同源物CspA、CspZ与FH和FH样蛋白（FHL）结合，旁系同源物ErpP、ErpC和ErpA与FH和补体因子H相关蛋白（CFHR）结合。尽管已经证明几种CRASP有助于体外血清抗性，但没有一种明确显示有助于哺乳动物感染。最近，我们发现一个B。携带erpA：：Tn插入的伯氏菌株在感染的早期阶段表现出组织定殖的缺陷。这些结果表明DbpA和ErpA分别调节C4 b2 a和C3 bBb，以促进菌血症和组织定殖。DbpA和ErpA抑制两类C3转化酶形成的能力可能意味着B对宿主血流防御的协同攻击。burgdorferi。为了检验这一假设，将追求以下目标。(1)测试FH结合在菌血症和定植中的作用。我们将测试靶向的非极性erpA缺失突变体是否在FH/CFHR结合、体外补体激活以及鼠模型中的菌血症和组织定植中显示缺陷。如果突变体显示缺陷，我们将测试FH与ErpA的结合是否对这些功能至关重要，以及其他CRASP是否可以在功能上弥补这些缺陷。（二）、通过B检测C4 BP和FH是否结合。burgdorferi提供促进菌血症和定植的非冗余作用。为了确定显著的播散缺陷是否需要C4 BP和FH结合活性的丧失，我们将在B的菌株背景中突变erpA。BurgdorferiDbpA-I156 A突变体。将在鼠模型中检测这些双突变菌株的重度菌血症和定植缺陷。

 DESCRIPTION (provided by applicant): Lyme disease, caused by the spirochete Borrelia burgdorferi, is the most common vector-borne disease in U.S. B. burgdorferi can infect the skin at the site of the tick bite and spread via blood to diverse tissues, indicating that the bacterium can withstand bloodstream defenses. The complement system is a bloodstream defense that is activated via three different pathways. A critical step of the activation of this system is the formation of C3 convertases, C4b2a or C3bBb, which catalyze events resulting in inflammation, opsonization and pathogen lysis. Potential host damage following complement activation necessitates tight regulation by serum complement regulators that bind to complement components and promote their degradation. For example, the complement regulator factor H (FH) and C4BP modulates formation of C3bBb and C4b2a, respectively. We recently showed that B. burgdorferi produces DbpA, a surface protein that promotes tissue colonization by binding to dermatan sulfate and decorin, also fosters bacteremia by binding to C4BP. A B. burgdorferi strain producing DbpA-I156A, a point mutant deficient in C4BP binding, showed a delay in bacteremia and joint colonization. In addition, B. burgdorferi also produces CRASPs (Complement Regulator Acquiring Surface Proteins) that bind to complement regulators. The paralogs CspA, CspZ bind to FH and FH-like protein (FHL), and the paralogs ErpP, ErpC, and ErpA bind to FH and complement factor H-related protein (CFHR). Although several CRASPs have been demonstrated to contribute to serum resistance in vitro, none have been definitely shown to contribute to mammalian infection. Recently, we found that a B. burgdorferi strain harboring an erpA::Tn insertion displayed a defect in tissue colonization at early stages of infection. These results suggest that DbpA and ErpA modulate C4b2a and C3bBb, respectively, to promote bacteremia and tissue colonization. The ability of DbpA and ErpA to inhibit the formation of both classes of C3 convertases may signify a coordinated attack on host bloodstream defenses by B. burgdorferi. To test this hypothesis, the following aims will be pursued. (1) Test the role of FH-binding in bacteremia and colonization. We will test if a targeted, non-polar erpA deletion mutant display defects in FH/CFHR binding, complement activation in vitro, and bacteremia and tissue colonization in the murine model. If the mutant displays defects, we will test if FH binding by ErpA is essential for these functions, and whether other CRASPs can functionally complement the defects. (2). Test if C4BP- and FH-binding by B. burgdorferi provide non-redundant roles to promote bacteremia and colonization. To determine if a dramatic dissemination defect requires loss of both C4BP- and FH-binding activities, we will mutate erpA in the strain background of the B. burgdorferi DbpA-I156A mutant. These double mutant strains will be tested for the severe bacteremia and colonization defect in the murine model.