Majeed M, Krause K H, Clark R A, Kihlström E, Stendahl O
Divisions of Medical Microbiology and Clinical Microbiology, Faculty of Health Sciences, Linköping University, S-581 85 Linköping, Sweden.
J Cell Sci. 1999 Jan;112 ( Pt 1):35-44. doi: 10.1242/jcs.112.1.35.
Chlamydia trachomatis elementary bodies (EBs) enter epithelial cells within membrane-bound endosomes that aggregate with each other in a calcium-regulated process, but avoid fusion with lysosomes. Annexin III but not I translocates to chlamydial aggregates and inclusions. In this study, we localize the intracellular Ca2+ stores during the course of infection by analyzing the distribution of three intracellular Ca2+ store proteins: calreticulin, type-1 inositol-1,4, 5-trisphosphate receptor (IP3-R), and Sarcoplasmic/Endoplasmic Reticulum Ca2+ ATPase type 2 (SERCA2) in HeLa cells infected with C. trachomatis serovar L2. In uninfected cells, immunofluorescence staining of the proteins showed a fine granular distributed pattern for all three proteins. After infection with C. trachomatis, calreticulin was found at the periphery of chlamydial aggregates and inclusions from 3 to 48 hours post-infection. In infected cells, SERCA2 was intimately associated with chlamydial inclusions after 3 and 24 hours, but not after 48 hours. Moreover, IP3-R was translocated to and colocalized with EB aggregates and chlamydial inclusions and had a distribution very similar to that of SERCA 2. After 24 hours incubation with chlamydiae, there was a local accumulation of [Ca2+]i (105+/-17 nM) in the proximity of chlamydial inclusions, compared to 50+/-13 nM in other parts of the cell cytoplasm. In the absence of extracellular Ca2+, this local accumulation of Ca2+ increased to 295+/-50 nM after adding 50 microM ATP, and to a similar extent after adding 100 nM thapsigargin (Tg). These data indicate that during infection of HeLa cells with chlamydiae, intracellular Ca2+ stores are redistributed, causing local accumulation of Ca2+ in the vicinity of chlamydial inclusions. These changes may trigger the association of certain proteins such as annexins with chlamydia-containing vesicles, and thereby regulation of membrane-membrane interaction during endosome aggregation and inclusion formation.
沙眼衣原体原体（EBs）进入膜结合的内体中的上皮细胞，这些内体在钙调节过程中相互聚集，但避免与溶酶体融合。膜联蛋白III而非膜联蛋白I易位至衣原体聚集体和包涵体。在本研究中，我们通过分析三种细胞内钙储存蛋白的分布来定位感染过程中的细胞内钙储存：钙网蛋白、1型肌醇-1,4,5-三磷酸受体（IP3-R）和2型肌浆网/内质网钙ATP酶（SERCA2），这些蛋白存在于感染沙眼衣原体血清型L2的HeLa细胞中。在未感染的细胞中，这些蛋白的免疫荧光染色显示所有三种蛋白均呈细颗粒状分布模式。感染沙眼衣原体后，在感染后3至48小时，钙网蛋白出现在衣原体聚集体和包涵体的周边。在感染的细胞中，SERCA2在3小时和24小时后与衣原体包涵体密切相关，但在48小时后则不然。此外，IP3-R易位至EB聚集体和衣原体包涵体并与其共定位，其分布与SERCA2非常相似。与衣原体孵育24小时后，衣原体包涵体附近的细胞内钙离子浓度局部积累至（105±17 nM），而细胞质其他部位为50±13 nM。在无细胞外钙的情况下，添加50 μM ATP后，钙离子的这种局部积累增加至295±50 nM，添加100 nM毒胡萝卜素（Tg）后也增加至类似程度。这些数据表明，在沙眼衣原体感染HeLa细胞的过程中，细胞内钙储存重新分布，导致衣原体包涵体附近的钙离子局部积累。这些变化可能触发某些蛋白（如膜联蛋白）与含衣原体囊泡的结合，从而在内体聚集和包涵体形成过程中调节膜-膜相互作用。
