Identifying Chlamydia trachomatis factors that mediate PD-L1 upregulation

鉴定介导 PD-L1 上调的沙眼衣原体因子

• 批准号: 10724569
• 负责人: MICHAEL N STARNBACH
• 金额: $ 24.78万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-06 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10724569
• 关键词: Antibiotics; Bacteria; Bacterial Genes; CD8-Positive T-Lymphocytes; CD8B1 gene; Candidate Disease Gene; Cell Separation; Cell physiology; Cells; Chimera organism; Chlamydia Infections; Chlamydia muridarum; Chlamydia trachomatis; Chromosomes; Chronic; Collection; Communities; Ectopic Pregnancy; Epidemic; Epithelial Cells; Foundations; Future; Genes; Genetic; Genomic Library; Goals; Hela Cells; Homing; Immune; Immune Evasion; Immune response; Immunity; Immunologic Memory; Impairment; In Vitro; Individual; Infection; Infection prevention; Infertility; Left; Lentivirus Vector; Libraries; Ligands; Malignant Neoplasms; Mediating; Memory; Mus; Pathway interactions; Pelvic Inflammatory Disease; Phenotype; Proliferating; Proteins; Public Health; Regulation; Reporting; Sampling; Secondary to; Sexually Transmitted Diseases; Signal Transduction; T cell response; T memory cell; T-Lymphocyte; Testing; Transcript; United States; Up-Regulation; Uterus; Vaccines; Virulence Factors; Virus; Work; adaptive immunity; chronic depression; chronic infection; experimental study; gain of function; immunoregulation; in vivo; mRNA Expression; mutant; null mutation; pathogen; pathogenic bacteria; prevent; programmed cell death ligand 1; programmed cell death protein 1; reproductive; response; screening; tool; urogenital tract; vaccine development; virulence gene

Chlamydia trachomatis is an obligate intracellular bacterium and the most prevalent sexually transmitted infection in the United States. If untreated, infection can lead to pelvic inflammatory disease, ectopic pregnancy, and infertility. Although C. trachomatis infection is treatable with antibiotics, many cases are asymptomatic, and repeat infections are common. Therefore, a vaccine is the best public health solution for managing and preventing infection. To develop a truly effective vaccine, it is critical to understand how C. trachomatis evades adaptive immunity to establish persistent infection during natural infection. CD8 T cells are normally integral for controlling intracellular pathogen infections, but during C. trachomatis infection, the CD8 T cell response is significantly impaired. We have shown that upregulation of the immunoinhibitory ligand PD-L1 during infection contributes to the diminution of the CD8 T cell response. Here we propose to identify C. trachomatis virulence genes that are responsible for upregulating PD-L1, as we hypothesize that these virulence genes are necessary to manipulate CD8 T cell immunity. In our first aim, we propose two complimentary approaches to identify C. trachomatis genes responsible for PD-L1 upregulation during infection. The first is to create and screen a lentiviral library containing individual C. trachomatis genes, testing the ability of individual bacterial genes to upregulate PD-L1. The second approach is based on a fortuitous phenotype in HeLa cells where only C. muridarum and not C. trachomatis is capable of upregulating PD-L1. By using a C. trachomatis/C. muridarum chimera collection developed by collaborators, we will identify portions of the C. muridarum chromosome that are sufficient to upregulate PD-L1. After generating a list of gene candidates from these two approaches, we will create C. trachomatis strains with null mutations in genes responsible for upregulating PD-L1 (CtNP), to validate their necessity for PD-L1 upregulation during infection in vitro and in vivo. In aim 2, we will use the CtNP strains to test if bacterial genes responsible for PD-L1 upregulation are required to manipulate the CD8 T cell response and hinder CD8 T cell-mediated protection. First, we will compare the proliferation and homing of C. trachomatis specific CD8 T cells during the course of infection with CtNP strains or WT C. trachomatis. Secondly, we propose to evaluate the protective capacity of memory CD8 T cells isolated from mice infected with CtNP strains or WT C. trachomatis by performing CD8 T cell transfer experiments into naïve mice. This work will provide the foundation for future studies to interrogate the mechanism by which C. trachomatis upregulates PD-L1. By understanding how C. trachomatis impairs the T cell response, we can develop vaccines that stimulate superior immunity compared to natural infection.

沙眼衣原体是一种专性细胞内细菌，也是最流行的性传播方式。 在美国的感染。如果不治疗，感染可导致盆腔炎、异位 怀孕和不孕不育。虽然沙眼衣原体感染可以用抗生素治疗，但许多病例是 无症状和重复感染很常见。因此，疫苗是治疗以下疾病的最佳公共卫生解决方案 管理和预防感染。要开发一种真正有效的疫苗，关键是要了解C. 在自然感染期间，沙眼衣原体逃避适应性免疫以建立持续感染。CD8 T细胞 通常对控制细胞内病原体感染是不可或缺的，但在沙眼衣原体感染期间，CD8T 细胞反应明显受损。我们已经证明，免疫抑制配体PD-L1的上调 在感染过程中，CD8 T细胞反应减弱。在这里，我们建议将C. 沙眼衣原体的毒力基因负责上调PD-L1，因为我们假设这些 毒力基因是调控CD8T细胞免疫所必需的。在我们的第一个目标中，我们提出两个 鉴定沙眼衣原体PD-L1上调基因的补充方法 感染。第一个是创建和筛选包含沙眼衣原体单个基因的慢病毒文库，测试 单个细菌基因上调PD-L1的能力。第二种方法是基于一次偶然的 HeLa细胞的表型，其中只有沙眼衣原体而不是沙眼衣原体能够上调PD-L1。通过 使用合作者开发的沙眼衣原体/小鼠衣原体嵌合体集合，我们将识别 足以上调PD-L1基因的小鼠染色体。在生成一份基因列表之后 这两种方法的候选者，我们将创造基因零突变的沙眼衣原体菌株。 负责上调PD-L1(CtNP)，以验证他们在感染期间上调PD-L1的必要性 体外和体内。在目标2中，我们将使用CtNP菌株来测试是否有导致PD-L1的细菌基因 需要上调来操纵CD8T细胞的反应，并阻碍CD8T细胞介导的保护。 首先，我们将比较沙眼衣原体特异性CD8T细胞在感染过程中的增殖和归巢情况。 感染沙眼衣原体或沙眼衣原体。其次，我们建议评估其防护能力。 从感染CtNP株和沙眼衣原体的小鼠体内分离CD8记忆T细胞 幼稚小鼠的细胞移植实验。这项工作将为以后的研究奠定基础 沙眼衣原体上调PD-L1的机制。通过了解沙眼衣原体如何损害 通过T细胞反应，我们可以开发出比自然感染更能激发免疫力的疫苗。