Porphyromonas gingivalis traffics into endoplasmic reticulum-rich-autophagosomes for successful survival in human gingival epithelial cells.
Porphyromonas gingivalis traffics into endoplasmic reticulum-rich-autophagosomes for successful survival in human gingival epithelial cells.
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DOI：
10.1080/21505594.2018.1454171
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发表时间：
2018-12-31
期刊：
影响因子：
5.2
通讯作者：
Yilmaz Ö
中科院分区：
文献类型：
--
作者：
Lee K;Roberts JS;Choi CH;Atanasova KR;Yilmaz Ö
牙龈卟啉单胞菌是一种机会性病原体，它篡夺了牙龈上皮细胞（GECs）作为其在口腔粘膜定植的主要细胞内生态位。然而，牙龈假单胞菌在gec中的细胞内运输和命运的精确表征仍然不完整。因此，我们采用高分辨率三维透射电子显微镜来确定牙龈假单胞菌入侵时在人原代gec中的亚细胞位置。感染gec的连续切片及其断层扫描重建显示富含er的双膜自噬体空泡窝藏牙龈假单胞菌。Western-blotting和荧光共聚焦显微镜显示，P. gingivalis以时间依赖性的方式显著诱导LC3脂化，并与LC3、ER-lumen-protein Bip或ER-tracker共定位，后者是吞噬体膜的主要成分。此外，用fmn -绿色荧光转化菌株（PgFbFP）感染并被洋地黄苷选择性渗透的gec中，双膜-液泡- p的数量迅速增加。牙龈感染24小时以上，胞质游离菌比例低。此外，通过抗生素保护试验确定，使用3-甲基腺嘌呤或ATG5 siRNA抑制自噬可显著降低gec细胞内牙龈卟啉卟啉菌的活力。溶酶体标记LAMP-1显示与牙龈卟啉卟啉菌低程度共定位（约20%）。PgFbFP用于研究液泡- p与细胞质- p的命运。它们与泛素结合接头蛋白NDP52和p62相关。只有cytosolic-P。牙龈菌与这两种标记物都有显著的关联，这表明无细胞壁细菌可能注定要进入溶酶体降解途径，而液泡p。gingivalis幸存。因此，研究结果揭示了牙龈假单胞菌通过利用宿主自噬机制在口腔黏膜中建立成功的复制生态位和持久性而在gec中存活的新机制。
Porphyromonas gingivalis, an opportunistic pathogen usurps gingival epithelial cells (GECs) as primary intracellular niche for its colonization in the oral mucosa. However, the precise characterization of the intracellular trafficking and fate of P. gingivalis in GECs remains incomplete. Therefore, we employed high-resolution three-dimensional-transmission-electron-microscopy to determine the subcellular location of P. gingivalis in human primary GECs upon invasion. Serial sections of infected-GECs and their tomographic reconstruction depicted ER-rich-double-membrane autophagosomal-vacuoles harboring P. gingivalis. Western-blotting and fluorescence confocal microscopy showed that P. gingivalis significantly induces LC3-lipidation in a time-dependent-manner and co-localizes with LC3, ER-lumen-protein Bip, or ER-tracker, which are major components of the phagophore membrane. Furthermore, GECs that were infected with FMN-green-fluorescent transformant-strain (PgFbFP) and selectively permeabilized by digitonin showed rapidly increasing large numbers of double-membrane-vacuolar-P. gingivalis over 24 hours of infection with a low-ratio of cytosolically free-bacteria. Moreover, inhibition of autophagy using 3-methyladenine or ATG5 siRNA significantly reduced the viability of intracellular P. gingivalis in GECs as determined by an antibiotic-protection-assay. Lysosomal marker, LAMP-1, showed a low-degree colocalization with P. gingivalis (∼20%). PgFbFP was used to investigate the fate of vacuolar- versus cytosolic-P. gingivalis by their association with ubiquitin-binding-adaptor-proteins, NDP52 and p62. Only cytosolic-P. gingivalis had a significant association with both markers, which suggests cytosolically-free bacteria are likely destined to the lysosomal-degradation pathway whereas the vacuolar-P. gingivalis survives. Therefore, the results reveal a novel mechanism for P. gingivalis survival in GECs by harnessing host autophagy machinery to establish a successful replicative niche and persistence in the oral mucosa.
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影响因子：
3.4
作者：
Choi CH;Spooner R;DeGuzman J;Koutouzis T;Ojcius DM;Yilmaz Ö
通讯作者：
Yilmaz Ö
Keystone-Pathogen假设。
DOI：
10.1038/nrmicro2873
发表时间：
2012-10
期刊：
Nature reviews. Microbiology
影响因子：
--
作者：
通讯作者：
--
影响因子：
5.8
作者：
Atanasova KR;Yilmaz Ö
通讯作者：
Yilmaz Ö
影响因子：
13.3
作者：
Cemma, Marija;Kim, Peter Kijun;Brumell, John Hunter
通讯作者：
Brumell, John Hunter
影响因子：
5.8
作者：
Johnson L;Atanasova KR;Bui PQ;Lee J;Hung SC;Yilmaz Ö;Ojcius DM
通讯作者：
Ojcius DM