Quantitative proteomic screen identifies Annexin A2 as a host target for Salmonella pathogenicity island-2 effectors SopD2 and PipB2

In this work we use stable isotope labeling of amino acids in cell culture (SILAC) and a S. Typhimurium mutant that secretes increased amounts of effectors to identify cognate effector binding partners during infection. Using this method, we identified the host protein annexin A2 as a binding partner for both SopD2 and PipB2 and were able to confirm its binding to SopD2 by reciprocal pull down. This indicates that SopD2 and PipB2 likely both interact with annexin A2 through protein-protein interactions to establish the S. Typhimurium intracellular replicativeniche. This demonstrates the value of studying effector interactions using proteomic techniques and natural effector delivery during infection rather than transfection.

本研究采用细胞培养氨基酸稳定同位素标记（stable isotope labeling of amino acids in cell culture, SILAC）技术，结合一株可过量分泌效应蛋白的鼠伤寒沙门氏菌（Salmonella Typhimurium, S. Typhimurium）突变株，以鉴定感染过程中效应蛋白的同源结合伴侣。借助该方法，我们鉴定到宿主膜联蛋白A2（annexin A2）同时作为SopD2与PipB2的结合伴侣，并通过反向pull-down实验验证了其与SopD2的相互作用。结果表明，SopD2与PipB2大概率通过蛋白质-蛋白质相互作用与膜联蛋白A2结合，进而构建鼠伤寒沙门氏菌的细胞内复制微环境。本研究证实，相较于转染手段，采用蛋白质组学技术并结合感染过程中天然的效应蛋白递送途径来探究效应蛋白相互作用，具备更高的研究价值。