Table 3_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）下的细菌均为真核细胞专性胞内寄生菌。该目下的衣原体属（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）的不同生物型是沙眼的致病原，也是全球范围内可预防性失明的首要病因，同时还可引发性传播感染，进而可能导致盆腔炎与不孕不育。无论引发何种疾病，所有衣原体物种均共享统一的专性胞内生命周期与发育细胞形态。其生存依赖一套感染周期，该周期至少包含三种表型截然不同的细胞形态：网状体（reticulate body, RB）、中间体（intermediate body, IB）与原体（elementary body, EB）。其中原体（EB）具有感染性但无法增殖；网状体（RB）可在宿主细胞内增殖，却不具备感染性；而中间体（IB）则是一类可向原体转化的中间过渡形态。本研究在网状体（RB）中异位表达了转录阻遏物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开展的转录组分析显示，衣原体的发育周期由分别对应网状体、中间体或原体三种细胞形态的三类独立调控子所驱动。此外，本研究证实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，提示该系统在特定细胞形态中发挥明确的功能作用。

研究意义：沙眼衣原体（Chlamydia trachomatis）引发的细菌性性传播感染对全球公共卫生构成重大威胁，每年导致超过1亿例感染，并可引发异位妊娠、不孕不育等多种并发症。本研究针对沙眼衣原体（C. trachomatis）在人类细胞内的独特生命周期展开基因表达模式分析。作为专性胞内寄生菌，沙眼衣原体会经历不同的发育阶段：其一用于感染与播散，其二用于细胞内增殖，其三则用于向感染性形态逆转。通过解析各阶段的基因表达谱，本研究鉴定出参与上述发育与感染过程的关键基因。值得注意的是，本研究还发现两类独立的Ⅲ型分泌系统（T3SS）转位因子在不同发育阶段特异性表达，提示其在感染周期的特定功能环节中发挥关键作用。