Global mapping of Salmonella enterica-host protein-protein interactions during infection

Summary of the study: Intracellular bacterial pathogens inject effector proteins into host cells to hijack diverse cellular processes and promote their survival and proliferation. To systematically map effector-host protein-protein interactions (PPIs) during infection, we generated a library of 32 Salmonella enterica serovar Typhimurium (STm) strains expressing chromosomally encoded affinity-tagged effectors, and quantified PPIs in macrophages and epithelial cells by Affinity-Purification Quantitative Mass-Spectrometry. Thereby, we identified 25 previously described and 421 novel effector-host PPIs. While effectors converged on the same host cellular processes, most had multiple targets, which often differed between cell types. Using reciprocal co-immunoprecipitations, we validated 13 out of 22 new PPIs. We used this PPI resource to further demonstrate that SseJ, SseL and SifA modulate cholesterol accumulation at the Salmonella Containing Vacuole (SCV) partially via the cholesterol transporter Niemann-Pick C1 protein (NPC1), PipB recruits the organelle contact site protein PDZD8 to the SCV, and SteC promotes actin bundling by phosphorylating formin-like proteins. Annotation to this repository: This collection of supplementary and original data completes the associated manuscript of the same title. It comprises: (01-04) The enrichment plots of the large scale AP/QMS study (related to figures 2, 3 and S2) (05-06) The abundances of all proteins in the AP/QMS study, alongside the full limma results (Figures 2, 3, S2 and S4) (07) The data at the basis of all plots shown in the manuscript (Figures 4, 5, 6, 7, S6 and S7) (08) All original Western Blot scans (Figures 4, 6, 7, S1, S3, S5, S7 and S8) (09) The limma results of the validation in pBMDMs (Figure S5) (10) The limma results of the separate TMT-run to assess the interaction partners of SseJ (Figures 5 and S6) (11-12) Additional information about the antibodies and primers used in this study (KRT and Experimental Procedures)

研究概述：胞内细菌致病菌可向宿主细胞内注入效应蛋白，以劫持多样化的细胞进程，促进自身存活与增殖。为系统绘制感染过程中的效应蛋白-宿主蛋白-蛋白互作（protein-protein interactions, PPIs）图谱，本研究构建了32株表达染色体编码亲和标签标记效应蛋白的鼠伤寒沙门氏菌（Salmonella enterica serovar Typhimurium, STm）菌株文库，随后通过亲和纯化定量质谱法（Affinity-Purification Quantitative Mass-Spectrometry）在巨噬细胞与上皮细胞中对PPIs进行定量分析。本研究由此鉴定出25种已报道的效应蛋白-宿主PPIs与421种新型效应蛋白-宿主PPIs。尽管各类效应蛋白均汇聚于相同的宿主细胞进程，但多数效应蛋白拥有多个靶标，且这些靶标往往因细胞类型而异。本研究通过反向免疫共沉淀实验，验证了22种新型PPIs中的13种。利用本研究获得的PPIs资源，我们进一步证实：SseJ、SseL与SifA可部分通过胆固醇转运蛋白尼曼-匹克C1蛋白（Niemann-Pick C1 protein, NPC1）调控沙门氏菌内含泡（Salmonella Containing Vacuole, SCV）处的胆固醇蓄积；PipB可将细胞器接触位点蛋白PDZD8招募至沙门氏菌内含泡；SteC可通过磷酸化类formin蛋白促进肌动蛋白束形成。

本数据集注释：本套补充数据与原始数据完整配套于同标题的相关学术论文。数据集包含以下内容：
(01-04) 大规模亲和纯化定量质谱研究的富集分析图（对应正文图2、图3及补充图S2）
(05-06) 大规模亲和纯化定量质谱研究中所有蛋白的表达丰度数据，以及完整的limma分析结果（对应正文图2、图3、补充图S2及S4）
(07) 论文中所有展示图表的原始数据（对应正文图4、5、6、7及补充图S6、S7）
(08) 所有原始蛋白免疫印迹扫描图像（对应正文图4、6、7及补充图S1、S3、S5、S7、S8）
(09) 原代骨髓来源巨噬细胞（pBMDMs）验证实验的limma分析结果（对应补充图S5）
(10) 单独开展的TMT标记实验（用于鉴定SseJ的互作伴侣）的limma分析结果（对应正文图5及补充图S6）
(11-12) 本研究所用抗体与引物的详细信息（详见KRT与实验方法部分）