Table 6_The T3SS structural and effector genes of Chlamydia trachomatis are expressed in distinct phenotypic cell forms.xlsx

Bacteria in the Chlamydiales order are obligate intracellular parasites of eukaryotic cells. Within this order, the genus Chlamydia contains the causative agents of a number of clinically important infections in humans. Biovars of Chlamydia trachomatis are the causative agents of trachoma and the leading cause of preventable blindness worldwide, as well as sexually transmitted infections with the potential to cause pelvic inflammatory disease and infertility. Irrespective of the resulting disease, all chlamydial species share the same obligate intracellular life cycle and developmental cell forms. They are reliant on an infectious cycle consisting of at least three phenotypically distinct cell forms termed the reticulate body (RB), the intermediate body (IB), and the elementary body (EB). The EB is infectious but does not replicate. The RB replicates in the host cell but is non-infectious, while the IB is an intermediate form that transitions to the EB form. In this study, we ectopically expressed the transcriptional repressor Euo, the two nucleoid-associated proteins HctA and HctB, and the two-component sensor kinase CtcB in the RB. Transcriptional analysis using RNA-seq, differential expression clustering, and fluorescence in situ hybridization analysis shows that the chlamydial developmental cycle is driven by three distinct regulons corresponding to the RB, IB, or EB cell forms. Moreover, we show that the genes for the type III secretion system (T3SS) were cell type restricted, suggesting defined functional roles for the T3SS in specific cell forms.
ImportanceChlamydia trachomatis, a sexually transmitted bacterial infection, poses a significant global health threat, causing over 100 million infections annually and leading to complications like ectopic pregnancy and infertility. This study investigates the gene expression patterns of C. trachomatis during its unique life cycle within human cells. As an obligate intracellular parasite, C. trachomatis transitions through distinct developmental stages—one for infection and dissemination, another for replication, and a third for transitioning back to the infectious form. By analyzing gene expression profiles at each stage, we identified key genes involved in these processes. Interestingly, our research also reveals the presence of two separate type III secretion system (T3SS) translocons expressed in distinct stages, suggesting their crucial roles in specific functions during the infection cycle.

衣原体目（Chlamydiales）的细菌是真核细胞的专性胞内寄生虫。在该目中，衣原体属（Chlamydia）包含多种引发人类临床重要感染的病原体。沙眼衣原体（Chlamydia trachomatis）的生物型是沙眼以及全球范围内可预防性失明的首要病因，同时也是可引发盆腔炎与不孕的性传播感染的病原体。无论引发何种疾病，所有衣原体物种均共享一致的专性胞内生命周期与发育细胞形态。它们依赖一套至少包含三种表型迥异细胞形态的感染周期，分别称为网状体（reticulate body, RB）、中间体（intermediate body, IB）与原体（elementary body, EB）。原体具备感染性但无法增殖；网状体可在宿主细胞内复制却不具备感染性；中间体则是一类过渡形态，可转化为原体。本研究在网状体中异位表达了转录阻遏蛋白Euo、两种类核结合蛋白HctA与HctB，以及双组分传感激酶CtcB。通过RNA测序（RNA-seq）、差异表达聚类与荧光原位杂交分析开展转录组研究，结果显示衣原体的发育周期由对应网状体、中间体或原体细胞形态的三类不同调控子（regulon）所驱动。此外，本研究还发现Ⅲ型分泌系统（type III secretion system, T3SS）的基因呈现细胞类型限制性表达，表明Ⅲ型分泌系统在特定细胞形态中承担明确的功能性角色。

重要性
沙眼衣原体引发的细菌性性传播感染对全球健康构成重大威胁，每年感染病例超1亿例，并可导致异位妊娠与不孕等并发症。本研究探究了沙眼衣原体在人类细胞内独特生命周期中的基因表达模式。作为专性胞内寄生虫，沙眼衣原体会经历不同的发育阶段：其一负责感染与播散，其二负责细胞内增殖，其三负责过渡回感染性形态。通过分析各阶段的基因表达谱，本研究鉴定出了参与这些过程的关键基因。值得关注的是，本研究还发现了两套分别在不同阶段表达的Ⅲ型分泌系统转位复合体（translocon），表明它们在感染周期的特定功能中发挥关键作用。