Homologues of the Chlamydia trachomatis and Chlamydia muridarum Inclusion Membrane Protein IncS Are Interchangeable for Early Development but Not for Inclusion Stability in the Late Developmental Cycle.
Homologues of the Chlamydia trachomatis and Chlamydia muridarum Inclusion Membrane Protein IncS Are Interchangeable for Early Development but Not for Inclusion Stability in the Late Developmental Cycle.
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沙眼衣原体是一种专性胞内细菌，在称为包涵体的空泡内经历双相发育周期。包埋在包涵体膜中的衣原体特异性效应蛋白，即Inc蛋白，促进包涵体与细胞器的相互作用。Inc蛋白的一个子集在内质网（ER）-包涵体膜接触位点（MCS）与特定宿主因子接合，MCS是包涵体膜和内质网（ER）之间的离散接触点。在这里，我们报告说，C。沙眼衣原体蛋白CTL 0402/IncSCt是ER-包含MCS的一种新组分，其特异性地与STIM 1相互作用并募集STIM 1，STIM 1是先前鉴定的ER-包含MCS的宿主组分，在包含膜处具有未分配的相互作用伴侣。相比之下，鼠衣原体IncS同源物（TC 0424/IncSCm）不与STIM 1相互作用或将STIM 1募集到内含物中，表明种特异性。为了进一步研究IncS的功能，并克服最近报道的早期发育缺陷的IncS突变体，我们实现了时间互补的表达IncS专门在早期阶段的发展周期。此外，我们使用等位基因交换将C中的incSCt开放阅读框架替换为incSCm。沙眼染色体包含这些菌株的进展，通过发展周期，但STIM 1阴性，并显示增加包涵体裂解感染后48小时。incSCt的反式表达补充了这些表型。总之，我们的研究结果表明，IncS是必要的和足够的招募STIM 1到C。沙眼衣原体包涵体中的IncS和C. trachomatis和C.以及在包合物稳定性中的晚期作用。沙眼专性细胞内病原体严格依赖宿主进行复制。专门的病原体编码的效应蛋白在复杂的宿主细胞操纵机制中发挥着核心作用。在衣原体中，这些效应蛋白的一个子集，即包涵体膜蛋白，嵌入细菌复制的液泡膜中。衣原体编码50至100个推定的Inc蛋白。许多是物种之间的保守，包括人类和小鼠病原体沙眼衣原体和鼠衣原体，分别。然而，Inc蛋白的功能是否在物种之间确实是保守的知之甚少。在此，我们对C. trachomatis和C. muridarum。我们的工作表明，一个单一的效应蛋白可以在发育周期的不同阶段发挥多种功能。然而，这些功能不一定是跨物种的保守，这表明衣原体物种之间的复杂进化路径。
Chlamydia trachomatis is an obligate intracellular bacterium, which undergoes a biphasic developmental cycle inside a vacuole termed the inclusion. Chlamydia-specific effector proteins embedded into the inclusion membrane, the Inc proteins, facilitate inclusion interaction with cellular organelles. A subset of Inc proteins engages with specific host factors at the endoplasmic reticulum (ER)-inclusion membrane contact site (MCS), which is a discrete point of contact between the inclusion membrane and the endoplasmic reticulum (ER). Here, we report that the C. trachomatis Inc protein CTL0402/IncSCt is a novel component of the ER-inclusion MCS that specifically interacts with and recruits STIM1, a previously identified host component of the ER-inclusion MCS with an unassigned interacting partner at the inclusion membrane. In comparison, the Chlamydia muridarum IncS homologue (TC0424/IncSCm) does not interact with or recruit STIM1 to the inclusion, indicating species specificity. To further investigate IncS function and overcome the recently reported early developmental defect of the incS mutant, we achieved temporal complementation by expressing IncS exclusively during the early stages of the developmental cycle. Additionally, we used allelic exchange to replace the incSCt open reading frame with incSCm in the C. trachomatis chromosome. Inclusions harboring either of these strains progressed through the developmental cycle but were STIM1 negative and displayed increased inclusion lysis 48 h postinfection. Expression of incSCt in trans complemented these phenotypes. Altogether, our results indicate that IncS is necessary and sufficient to recruit STIM1 to C. trachomatis inclusion and that IncS plays an early developmental role conserved in C. trachomatis and C. muridarum and a late role in inclusion stability specific to C. trachomatis. IMPORTANCE Obligate intracellular pathogens strictly rely on the host for replication. Specialized pathogen-encoded effector proteins play a central role in sophisticated mechanisms of host cell manipulation. In Chlamydia, a subset of these effector proteins, the inclusion membrane proteins, are embedded in the membrane of the vacuole in which the bacteria replicate. Chlamydia encodes 50 to 100 putative Inc proteins. Many are conserved among species, including the human and mouse pathogens Chlamydia trachomatis and Chlamydia muridarum, respectively. However, whether the function(s) of Inc proteins is indeed conserved among species is poorly understood. Here, we characterized the function of the Inc protein IncS conserved in C. trachomatis and C. muridarum. Our work reveals that a single effector protein can play multiple functions at various stages of the developmental cycle. However, these functions are not necessarily conserved across species, suggesting a complex evolutionary path among Chlamydia species.
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DOI：
10.3791/60848
发表时间：
2020-01-30
期刊：
Journal of visualized experiments : JoVE
影响因子：
--
作者：
Keb G;Fields KA
通讯作者：
Fields KA
影响因子：
6.7
作者：
Derré I;Swiss R;Agaisse H
通讯作者：
Agaisse H
DOI：
10.1111/tra.12002
发表时间：
2012-12
期刊：
Traffic (Copenhagen, Denmark)
影响因子：
--
作者：
Dumoux M;Clare DK;Saibil HR;Hayward RD
通讯作者：
Hayward RD
影响因子：
3.2
作者：
Keb, G.;Hayman, R.;Fields, K. A.
通讯作者：
Fields, K. A.
影响因子：
6.7
作者：
Elwell CA;Jiang S;Kim JH;Lee A;Wittmann T;Hanada K;Melancon P;Engel JN
通讯作者：
Engel JN