Virulence Factors of Chlamydiae

描述(申请人提供)：沙眼衣原体D-K血清型在美国和全世界是细菌获得性传播疾病及其后遗症的最常见原因，包括前列腺炎、附睾炎、盆腔炎、宫外孕和不孕症。衣原体疾病是潜伏的，它们构成了重大的初级、二级和三级健康问题，其中妇女承担着特别的负担，因为不良生殖后果的风险增加。25年来，我们实验室的目标一直是试图了解衣原体在宿主上皮细胞中生长的基本生物学，以了解感染过程，并允许解剖持续感染的细胞/分子后果，因为大多数衣原体输卵管疾病似乎是由亚临床、持续感染引起的。这一建议是对关键的依恋步骤的持续努力的继续，这些步骤在体外受到女性激素的调节。在特定的目标1中，我们将确定雌激素受体(ER)是否参与E血清型与人子宫内膜上皮细胞的结合，或者，如果雌激素对宿主细胞的生理作用导致其他受体的显示。除了Biacore、FACS和显微镜分析外，还将使用ER细胞系的特定标记EB结合分析，并结合siRNA技术来敲除表面ER量，以查看是否发生EB结合减少。在具体目标2中，我们将通过显微镜和梯度囊泡分离来监测E血清型外膜泡膜抗原通过包涵膜以及从包涵膜外翻小泡逃逸的机制，并确定转运，包括MHC I类负载的内质网膜融合潜力。最后，为了具体目标3，我们分离了衣原体突起(小规模)，并在大量收集和产生抗体后，计划表征这些表面结构；明显的问题是，这些突起是否是衣原体III型分泌物注射剂的等价物。

DESCRIPTION (provided by applicant): Chlamydia trachomatis serovars D-K are the most common cause, in the USA and worldwide, of bacterially acquired sexually transmitted diseases and their sequelae, including prostatitis, epididymitis, pelvic inflammatory disease, ectopic pregnancy and sterility. Chlamydial diseases are insidious and they constitute significant primary, secondary and tertiary health concerns in which women bear a special burden because of increased risk of adverse reproductive consequences. The goal of our laboratory for 25 years has been to try to understand the basic biology of Chlamydial growth in its host epithelial cell in order to understand the infectious process and to permit dissection of the cellular/molecular consequences of persistent infection, since the majority of Chlamydial tubal disease appears to result from subclinical, persistent infection. This proposal is a continuation of on-going efforts to understand the crucial attachment steps, which are modulated by female hormones in vitro. In Specific Aim 1, we shall determine whether or not estrogen receptors (ER) are involved in serovar E attachment to human endometrial epithelial cells or, alternatively, if the estrogen-induced physiological effects on the host cell result in display of other receptors. Specific labeled-EB binding assays to ER+ cell lines will be used in addition to BiaCore, FACS, and microscopy analyses and coupled with siRNA technology to knockdown surface ER amounts to see if reduced EB binding does occur. In Specific Aim 2, we shall monitor, by microscopy and gradient vesicle isolation, mechanisms of serovar E outer membrane bleb antigen escape through the inclusion membrane as well as from inclusion membrane everted vesicles and determine trafficking, including endoplasmic reticulum membrane fusion potential for MHC Class I loading. Finally, for Specific Aim 3, we have isolated Chlamydial projections (small scale) and, after bulk collection and generation of antibodies, plan to characterize these surface structures; the obvious question is whether or not the projections are the Chlamydial Type III secretion injectisome-equivalent.