CALRETICULIN--A SECRETED PROTEIN IN IXODID TICK SALIVA

扁虱传播更多类型的微生物，感染更多的人 莱姆病是美国最常见的节肢动物 对于扁虱的担忧死灰复燃，尽管落基山发现 发热、科罗拉多州壁虱热和复发热仍然是一个持续的威胁。 在区域基础上对人类人口的影响。我们提案的重点是 在扁虱的唾液腺上，促进了疾病的传播和 产生止血、抗炎和免疫抑制作用 控制叮咬部位，使寄生虫可以觅食的制剂。我们的 总体目标是确定主要唾液成分的特征 调节喂食点的环境。理解继承人的角色应该 提供对疾病传播过程的基本见解，并 寄主对寄生虫的反应是如何中和的，甚至是如何用来 扁虱的优势。直到最近，在定性方面取得了进展 有机唾液成分的含量只是适中的，根据反应 唾液或腺体提取物的体外检测。基本利率- 限制措施一直是组织(唾液腺)丰度。 分子遗传学技术的进步现在为 克服这一限制。我们已经确定了一种形式的钙网硬蛋白 两个重要的医学物种(摄食美洲钝线虫和 雌性变色尘螨)，基于 与兔骨骼肌推导的氨基酸序列同源性为69% 钙网织蛋白)和腺体提取物和唾液的阳性免疫印迹 用山羊抗兔钙网织蛋白。钙网硬蛋白是一种高度保守的 非肌细胞内质网内钙结合蛋白和 平滑肌肌浆网。非肌肉组织中的蛋白质 细胞内质网和平滑肌肌浆网。 蛋白质很可能被合成用于分泌，这一点得到了支持 通过序列数据，因为ER保持信号(KDEL)显然不 存在；而且，它存在于唾液中。光和超微结构水平 该蛋白的免疫定位及原位杂交研究 形态发生过程中的腺体有助于将蛋白质分泌与 与宿主相关的反应，并确定蛋白质在哪里合成 并位于细胞内。止血(血小板聚集)和 血管(平滑肌收缩)检测将提供有价值的 关于腺体蛋白作为分泌产物的作用的信息。 我们将研究钙离子依赖和独立的过程。 使用这些方法表征钙网硬蛋白将呈现一种 在描述其何时何地分泌方面取得了重大进展， 在开始定义其生理意义的过程中 在进食过程中。这一理解将对 了解寄生虫-宿主关系和可能的疾病 硬蜱传播。

Ticks transmit more types of microorganisms and infect more people with disease than any other arthropod in the U.S. Lyme disease is responsible for a resurgence of concern about tick, although Rocky Mountain spotted fever, Colorado tick fever and relapsing fever remain a continuous threat to the human population on a regional basis. The focus of our proposal is on the tick's salivary glands that facilitate disease transmission and produce t he antihemostasis, antiinflammatory and immunosuppressive agents, which control the bite site so the parasite can feed. Our overall objective is to characterize the primary salivary components that mediate the feeding-site environment. Understanding heir roles should provide fundamental insights into the disease transmission process and how the host's responses to the parasite are neutralized or even used to the tick's advantage. Until recently, progress towards characterization of organic salivary components was only modest, and based upon responses of in vitro assays to saliva or gland extracts. The primary rate- limiting steps has been that of tissue (salivary gland) abundance. Advances in molecular genetics technology now provide the opportunity to overcome this limitation. We have identified a form of calreticulin in two medically important tick species (feeding Amblyomma americanum & Dermacentor variabilis females), based upon complete sequencing of a cDNA, (69% deduced amino acid homology with rabbit skeletal muscle calreticulin) and positive immunoblotting of gland extracts and saliva with goat anti-rabbit calreticulin. Calreticulin is a highly conserved Ca2+ binding protein within non-muscle cell endoplasmic reticulum and smooth muscle sarcoplasmic reticulum. That the protein within non-muscle cell endoplasmic reticulum and smooth muscle sarcoplasmic reticulum. That the protein is likely to be synthesized for secretion is supported by sequence data as the ER retention signal (KDEL) is apparently not present; and, it is found in the saliva. Light and ultrastructural level immunolocalization of the protein and in situ hybridization studies of glands during morphogenesis will help correlate protein secretion with host-related responses, and determine where the protein is synthesized and located within the cell. Hemostasis (platelet aggregation) and vascular (smooth muscle contraction) assays will provide valuable information about the role of the gland protein as a secreted product. Ca2+-dependent and independent processes will be investigated. Characterization of calreticulin using these methods will present a significant advancement in describing when and where its is secreted, while beginning the process of defining its physiological significance in the feeding process. This understanding will be critical in understanding the parasite-host relationship and possibly disease transmission by ixodid ticks.