Salmonella YrbD protein mediates invasion into the host by interacting with β2 integrin

doi:10.1016/j.jia.2023.12.035

Journal of Integrative Agriculture

2025, Vol. 24

Issue (3): 1181-1197 DOI: 10.1016/j.jia.2023.12.035

Animal Science · Veterinary Medicine

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Salmonella YrbD protein mediates invasion into the host by interacting with β2 integrin

Jun Cao, Jinliang Wang, Guanghui Dang, Shihao Ding, Yao Lu, Qiu Xu, Siguo Liu^#, Shenye Yu^#

State Key Laboratory for Animal Disease Control and Prevention, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150069, China

Highlights

● β2 integrin is an important host factor that promotes the invasion of Salmonella into host cells.

● The soluble protein in the extracellular domain of β2 integrin can neutralize Salmonella infection.

● The direct interaction between β2 integrin and Salmonella protein YrbD promotes the adhesion and internalization

of Salmonella to host cells.

Abstract
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摘要

鼠伤寒沙门菌是胃肠炎的病原体，是最成功的细胞内病原体之一。尽管沙门菌感染的某些宿主因素已经被揭示，但介导沙门菌进入的因素，特别是入侵过程，仍然不清楚。本研究首先在质谱筛选出的28种潜在沙门菌宿主因子中选择β2整合素为研究目标，在HeLa细胞中过表达β2整合素或用siRNA转染，再用沙门菌感染。结果显示β2整合素过表达的HeLa细胞中沙门菌感染显著高于对照组，siRNA转染组的β2整合素mRNA转录下降了90%，表明β2整合素是促进沙门菌入侵的重要宿主因子。其次，用沙门菌分别感染β2整合素过表达与沉默的HeLa细胞，对细胞表面的沙门菌进行共聚焦显微观察，结果显示沙门菌与β2整合素沉默的HeLa细胞的结合减少，而与β2整合素过表达的HeLa细胞的结合增加。将识别β2整合素胞外结构域的单克隆抗体或多克隆抗体分别与HeLa、Caco-2和RAW264.7细胞预孵育，对膜表面的β2整合素进行抗体阻断，测定感染率，结果显示阻断细胞表面的β2整合素显著减少了沙门菌感染，而且呈剂量依赖性，表明β2整合素与沙门菌感染存在特异的相关性。再次，分别进行细胞感染性中和试验和小鼠感染性中和试验，结果显示β2整合素胞外结构域的可溶性重组蛋白以剂量依赖的方式有效地中和了HeLa、Caco-2和RAW264.7细胞中的沙门菌感染；重组蛋白组的存活期比对照组延长了48小时，表明β2整合素胞外结构域可溶性蛋白在细胞（体外）和小鼠（体内）中和了沙门菌感染。然后，通过β2整合素与沙门菌外膜蛋白的pull-down试验以及β2整合素和YrbD的免疫共沉淀和pull-down试验，证明β2整合素与沙门菌YrbD直接相互作用，并且I-like结构域是相互作用所必需的。庆大霉素保护试验显示，沙门菌的yrbD缺失显著降低了对HeLa细胞的侵袭，表明YrbD有助于沙门菌入侵细胞。用纯化的YrbD蛋白包被的乳胶珠孵育HeLa细胞，透射电镜显示乳胶珠被非吞噬性HeLa细胞特异性吸收并内化。本研究首次表明，YrbD蛋白通过与β2整合素相互作用介导沙门菌粘附和内化到宿主细胞中。这些发现不仅拓宽了我们对沙门菌进入机制的理解，而且确定了治疗控制的潜在靶点。

Abstract

Salmonella enterica serovar Typhimurium, the causative agent of gastroenteritis, is one of the most successful intracellular pathogens. Although certain host factors for Salmonella infection have been unveiled, the factors mediating Salmonella entry, particularly the invasion process, remain obscure. Here, we have unearthed β2 integrin, a crucial member of the integrin family, as an important host factor facilitating Salmonella invasion. It is demonstrated that overexpression of β2 integrin promotes Salmonella invasion, while the knockdown of β2 integrin significantly diminishes the extent of invasion. Moreover, Salmonella exhibits specific binding affinity towards β2 integrin, and the block of β2 integrin on cell surface substantially reduces the infection of cells in vitro. The ectodomain soluble protein of β2 integrin neutralized Salmonella infection both in cells (in vitro) and in mice (in vivo). Additionally, Salmonella protein YrbD directly interacts with β2 integrin to facilitate its invasion. To our knowledge, this study showed for the first time that the protein YrbD mediates Salmonella adhesion and internalization into host cells by interacting with β2 integrin. These findings not only broaden our understanding of the mechanisms underlying Salmonella entry, but also identify a prospective target for therapeutic control.

Keywords: β2 integrin adhesion internalization Salmonella protein YrbD

Received: 25 August 2023 Accepted: 31 October 2023

Fund:

This work was supported by the National Key R&D Program of China (2022YFF0710500), the the National Natural Science Foundation, China (31802192, 32172853 and 32373013), the Natural Science Foundation of Heilongjiang Province of China (C2018070), China Postdoctoral Science Foundation (2017M620076), and the Central Public-interest Scientific Institution Basal Research Fund, China (1610302022001).

About author: Jun Cao, E-mail: yufei@126.com; #Correspondence Shenye Yu, Tel: +86-451-51051733, E-mail: yushenye@caas.cn

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