Host-Parasite Interactions Section, Laboratory of Bacteriology, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Hamilton, Montana, USA.
mBio. 2024 Apr 10;15(4):e0345023. doi: 10.1128/mbio.03450-23. Epub 2024 Mar 6.
We compared the growth characteristics of a virulent strain (Sheila Smith) to an attenuated stain (Iowa) and a non-pathogenic species () in primary human dermal microvascular endothelial cells (HDMEC). All replicated in Vero cells, however, only the Sheila Smith strain productively replicated in HDMECs. The Iowa strain showed minimal replication over a 24-h period, while lost viability and induced lysis of the HDMECs via a rapid programmed cell death response. Both the virulent and attenuated strains, but not , induced an interferon-1 response, although the response was of lesser magnitude and delayed in the Sheila Smith strain. IFN-β secretion correlated with increased host cell lysis, and treatment with anti-IFNAR2 antibody decreased lysis from Iowa-infected but not Sheila Smith-infected cells. Both Sheila Smith- and Iowa-infected cells eventually lysed, although the response from Sheila Smith was delayed and showed characteristics of apoptosis. We, therefore, examined whether reconstitution of the Iowa strain with two recently described putative virulence determinants might enhance survival of Iowa within HDMECs. Reconstitution with RARP2, which is inhibitory to anterograde trafficking through the Golgi apparatus, reduced IFN-β secretion but had no effect on cell lysis. RapL, which proteolytically processes surface exposed autotransporters and enhances replication of Iowa in Guinea pigs, suppressed both IFN-β production and host cell lysis. These findings suggest distinct mechanisms by which virulent spotted fever group rickettsiae may enhance intracellular survival and replication.IMPORTANCEWe examined a naturally occurring non-pathogenic rickettsial species, , a laboratory-attenuated strain (Iowa), and a fully virulent strain (Sheila Smith) for growth in human dermal microvascular endothelial cells. The two avirulent strains replicated poorly or not at all. Only the virulent Sheila Smith strain replicated. IFN-β production correlated with the inhibition of Iowa. Reconstitution of Iowa with either of two recently described putative virulence determinants altered the IFN-β response. A rickettsial ankyrin repeat protein, RARP2, disrupts the Golgi network and inhibits IFN-β secretion. An autotransporter peptidase, RapL, restores proteolytic maturation of outer membrane autotransporters and diminishes the IFN-β response to enhance cell survival and permit replication of the recombinant strain. These studies point the way toward discovery of mechanisms for innate immune response avoidance by virulent rickettsia.
我们比较了一种强毒株（Sheila Smith）、一种弱毒株（Iowa）和一种非致病性种（）在原代人真皮微血管内皮细胞（HDMEC）中的生长特性。所有株系都能在 Vero 细胞中复制，但只有 Sheila Smith 株系能在 HDMEC 中有效复制。Iowa 株系在 24 小时内复制能力有限，而 株系则通过快速程序性细胞死亡反应失去活力并诱导 HDMEC 裂解。强毒株和弱毒株都能诱导干扰素-1 反应，尽管反应幅度较小且在 Sheila Smith 株系中延迟，但 株系不能诱导干扰素-1 反应。IFN-β 的分泌与宿主细胞裂解增加相关，用抗 IFNAR2 抗体治疗可减少 Iowa 感染细胞的裂解，但不能减少 Sheila Smith 感染细胞的裂解。Sheila Smith-和 Iowa 感染的细胞最终都发生了裂解，尽管 Sheila Smith 的反应延迟且具有凋亡特征。因此，我们研究了用最近描述的两种假定毒力决定因子重建 Iowa 株系是否能增强 Iowa 在 HDMEC 中的存活能力。用 RARP2 重建 Iowa，RARP2 可抑制高尔基体顺行运输，减少 IFN-β 的分泌，但对细胞裂解无影响。RapL 可蛋白水解处理表面暴露的自转运蛋白，并增强 Iowa 在豚鼠中的复制能力，抑制 IFN-β 的产生和宿主细胞的裂解。这些发现表明，强毒斑点热群立克次体可能通过不同机制增强细胞内生存和复制能力。
我们研究了一种天然非致病性立克次体种、一种实验室弱毒株（Iowa）和一种完全强毒株系（Sheila Smith）在人真皮微血管内皮细胞中的生长情况。两种非致病株系复制能力差或根本不复制。只有强毒 Sheila Smith 株系复制。IFN-β 的产生与 Iowa 的抑制相关。用最近描述的两种假定毒力决定因子中的任何一种重建 Iowa 都会改变 IFN-β 的反应。一种立克次体锚蛋白重复蛋白 RARP2 破坏了高尔基体网络并抑制了 IFN-β 的分泌。一种自转运体蛋白酶 RapL 恢复了外膜自转运体的蛋白水解成熟，并减弱了 IFN-β 对细胞存活的反应，从而促进了重组株系的复制。这些研究为发现强毒立克次体逃避先天免疫反应的机制指明了方向。
