Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, Indiana, USA.
Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska, USA.
mSphere. 2023 Jun 22;8(3):e0010423. doi: 10.1128/msphere.00104-23. Epub 2023 Apr 5.
Coxiella burnetii is an intracellular bacterium that causes the human disease Q fever. C. burnetii forms a large, acidic -containing vacuole (CCV) and uses a type 4B secretion system to secrete effector proteins into the host cell cytoplasm. While the CCV membrane is rich in sterols, cholesterol accumulation in the CCV is bacteriolytic, suggesting that C. burnetii regulation of lipid transport and metabolism is critical for successful infection. The mammalian lipid transport protein ORP1L (oxysterol binding protein-like protein 1 Long) localizes to the CCV membrane and mediates CCV-endoplasmic reticulum (ER) membrane contact sites. ORP1L functions in lipid sensing and transport, including cholesterol efflux from late endosomes and lysosomes (LELs), and the ER. Its sister isoform, ORP1S (oxysterol binding protein-like protein 1 Short) also binds cholesterol but has cytoplasmic and nuclear localization. In ORP1-null cells, we found that CCVs were smaller than in wild-type cells, highlighting the importance of ORP1 in CCV development. This effect was consistent between HeLa cells and murine alveolar macrophages (MH-S cells). CCVs in ORP1-null cells had higher cholesterol content than CCVs in wild-type cells at 4 days of infection, suggesting ORP1 functions in cholesterol efflux from the CCV. While the absence of ORP1 led to a C. burnetii growth defect in MH-S cells, there was no growth defect in HeLa cells. Together, our data demonstrated that C. burnetii uses the host sterol transport protein ORP1 to promote CCV development, potentially by using ORP1 to facilitate cholesterol efflux from the CCV to diminish the bacteriolytic effects of cholesterol. Coxiella burnetii is an emerging zoonotic pathogen and bioterrorism threat. No licensed vaccine exists in the United States, and the chronic form of the disease is difficult to treat and potentially lethal. Postinfectious sequelae of C. burnetii infection, including debilitating fatigue, place a significant burden on individuals and communities recovering from an outbreak. C. burnetii must manipulate host cell processes in order to promote infection. Our results establish a link between host cell lipid transport processes and C. burnetii's avoidance of cholesterol toxicity during infection of alveolar macrophages. Elucidating the mechanisms behind bacterial manipulation of the host will yield insight for new strategies to combat this intracellular pathogen.
贝氏考克斯氏体是一种引起人类 Q 热的细胞内细菌。贝氏考克斯氏体形成一个大型的、含有酸性物质的空泡（CCV），并利用一种 4B 型分泌系统将效应蛋白分泌到宿主细胞质中。虽然 CCV 膜富含固醇，但 CCV 中的胆固醇积累会导致细菌裂解，这表明贝氏考克斯氏体对脂质运输和代谢的调节对于成功感染至关重要。哺乳动物脂质转运蛋白 ORP1L（氧化固醇结合蛋白样蛋白 1 长）定位于 CCV 膜，并介导 CCV-内质网（ER）膜接触位点。ORP1L 参与脂质感应和运输，包括晚期内体和溶酶体（LEL）以及 ER 中的胆固醇外排。其姐妹同工型 ORP1S（氧化固醇结合蛋白样蛋白 1 短）也结合胆固醇，但具有细胞质和核定位。在 ORP1 缺失细胞中，我们发现 CCV 比野生型细胞小，这突出了 ORP1 在 CCV 发育中的重要性。这种效应在 HeLa 细胞和鼠肺泡巨噬细胞（MH-S 细胞）之间是一致的。在感染 4 天时，ORP1 缺失细胞中的 CCV 胆固醇含量高于野生型细胞中的 CCV，表明 ORP1 参与了 CCV 中的胆固醇外排。虽然 ORP1 的缺失导致 MH-S 细胞中的贝氏考克斯氏体生长缺陷，但在 HeLa 细胞中没有生长缺陷。总之，我们的数据表明，贝氏考克斯氏体利用宿主固醇转运蛋白 ORP1 来促进 CCV 的发育，这可能是通过利用 ORP1 促进 CCV 中的胆固醇外排，从而减轻胆固醇的杀菌作用。
贝氏考克斯氏体是一种新兴的人畜共患病原体和生物恐怖威胁。在美国，没有许可的疫苗，疾病的慢性形式难以治疗，且可能致命。感染贝氏考克斯氏体后的后遗症，包括使人虚弱的疲劳，给个人和社区从疫情中恢复带来了巨大的负担。贝氏考克斯氏体必须操纵宿主细胞过程才能促进感染。我们的研究结果建立了宿主细胞脂质运输过程与贝氏考克斯氏体在感染肺泡巨噬细胞时避免胆固醇毒性之间的联系。阐明细菌操纵宿主的机制将为对抗这种细胞内病原体提供新的策略提供洞察力。
