Department of Microbiology and Immunology, Virginia Commonwealth University Medical Center, School of Medicine, Richmond, VA, United States.
Department of Clinical Sciences and the Intracellular Pathogens Research Laboratory, Comparative Medicine Institute, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, United States.
Front Cell Infect Microbiol. 2020 Sep 23;10:500. doi: 10.3389/fcimb.2020.00500. eCollection 2020.
is an obligate intracellular bacterium that invades monocytes to cause the emerging and potentially severe disease, monocytic ehrlichiosis. Ehrlichial invasion of host cells, a process that is essential for the bacterium's survival and pathogenesis, is incompletely understood. In this study, we identified ECH_0377, henceforth designated as EplA ( PDI ligand A) as an adhesin that interacts with host cell protein disulfide isomerase (PDI) to mediate bacterial entry into host cells. EplA is an outer membrane protein that expresses during growth in THP-1 monocytic cells. Canine sera confirmed to be positive for exposure to spp. recognized recombinant EplA, indicating that it is expressed during infection . EplA antiserum inhibited the bacterium's ability to infect monocytic cells. The EplA-PDI interaction was confirmed via co-immunoprecipitation. Treating host cell surfaces with antibodies that inhibit PDI and/or thioredoxin-1 thiol reductase activity impaired infection. Chemical reduction of host cell surfaces, but not bacterial surfaces with tris(2-carboxyethyl)phosphine (TCEP) restored ehrlichial infectivity in the presence of the PDI-neutralizing antibody. Antisera specific for EplA C-terminal residues 95-104 (EplA) or outer membrane protein A amino acids 53-68 (OmpA) reduced infection of THP-1 cells. Notably, TCEP rescued ehrlichial infectivity of bacteria that had been treated with anti-EplA, but not anti-EcOmpA. These results demonstrate that EplA contributes to infection of monocytic cells by engaging PDI and exploiting the enzyme's reduction of host cell surface disulfide bonds in an EplA C-terminus-dependent manner and identify EplA and EcOmpA as novel ehrlichial receptor binding domains.
是一种专性细胞内细菌，它侵入单核细胞引起新兴的潜在严重疾病，单核细胞埃立克体病。埃立克体对宿主细胞的入侵，这是细菌生存和发病机制的必要过程，目前还不完全了解。在这项研究中，我们鉴定了 ECH_0377，此后称为 EplA（PDI 配体 A），它是一种黏附素，与宿主细胞蛋白二硫键异构酶（PDI）相互作用，介导细菌进入宿主细胞。EplA 是一种外膜蛋白，在 THP-1 单核细胞中生长时表达。经证实对 spp. 暴露呈阳性的犬血清识别重组 EplA，表明它在感染期间表达。EplA 抗血清抑制了细菌感染单核细胞的能力。EplA-PDI 相互作用通过共免疫沉淀得到证实。用抑制 PDI 和/或硫氧还蛋白-1 巯基还原酶活性的抗体处理宿主细胞表面，会损害 的感染。用三（2-羧乙基）膦（TCEP）化学还原宿主细胞表面，但不还原细菌表面，可在中和 PDI 的抗体存在下恢复埃立克体的感染力。针对 EplA C 端残基 95-104（EplA）或外膜蛋白 A 氨基酸 53-68（OmpA）的抗血清降低了 THP-1 细胞中 的感染。值得注意的是，TCEP 挽救了用抗 EplA 处理但不抗 EcOmpA 处理的细菌的埃立克体感染力。这些结果表明，EplA 通过与 PDI 结合并利用该酶还原宿主细胞表面二硫键，以 EplA C 端依赖性方式促进单核细胞的感染，并将 EplA 和 EcOmpA 鉴定为新型埃立克体受体结合结构域。
