Costa Tiago R D, Francis Monika K, Farag Salah I, Edgren Tomas, Francis Matthew S
Department of Molecular Biology, Umeå University, Umeå, Sweden.
Umeå Centre for Microbial Research, Umeå University, Umeå, Sweden.
Methods Mol Biol. 2019;2010:211-229. doi: 10.1007/978-1-4939-9541-7_15.
Many Gram-negative pathogens produce a type III secretion system capable of intoxicating eukaryotic cells with immune-modulating effector proteins. Fundamental to this injection process is the prior secretion of two translocator proteins destined for injectisome translocon pore assembly within the host cell plasma membrane. It is through this pore that effectors are believed to travel to gain access to the host cell interior. Yersinia species especially pathogenic to humans and animals assemble this translocon pore utilizing two hydrophobic translocator proteins-YopB and YopD. Although a full molecular understanding of the biogenesis, function and regulation of this translocon pore and subsequent effector delivery into host cells remains elusive, some of what we know about these processes can be attributed to studies of bacterial infections of erythrocytes. Herein we describe the methodology of erythrocyte infections by Yersinia, and how analysis of the resultant contact-dependent hemolysis can serve as a relative measurement of YopB- and YopD-dependent translocon pore formation.
许多革兰氏阴性病原体产生一种III型分泌系统，该系统能够用免疫调节效应蛋白使真核细胞中毒。这一注射过程的基础是预先分泌两种转运蛋白，它们用于在宿主细胞质膜内组装注射体转运孔。效应蛋白据信是通过这个孔进入宿主细胞内部的。对人和动物尤其致病的耶尔森氏菌属利用两种疏水转运蛋白YopB和YopD组装这个转运孔。尽管对这个转运孔的生物发生、功能和调节以及随后效应蛋白递送至宿主细胞的过程仍缺乏全面的分子层面理解，但我们对这些过程的部分了解可归因于对红细胞细菌感染的研究。在此，我们描述耶尔森氏菌感染红细胞的方法，以及如何通过分析由此产生的接触依赖性溶血来相对测量依赖YopB和YopD的转运孔形成。
