Manipulation of pro-inflammatory cytokine production by the bacterial cell-penetrating effector protein YopM is independent of its interaction with host cell kinases RSK1 and PRK2
Manipulation of pro-inflammatory cytokine production by the bacterial cell-penetrating effector protein YopM is independent of its interaction with host cell kinases RSK1 and PRK2
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DOI：
10.4161/viru.29062
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发表时间：
2014-10-01
期刊：
影响因子：
5.2
通讯作者：
Rueter, Christian
中科院分区：
文献类型：
--
作者：
Hoefling, Sabrina;Scharnert, Julia;Rueter, Christian
小肠结肠炎耶尔森氏菌的效应蛋白耶尔森氏菌外部蛋白 M (YopM) 先前已被鉴定并表征为第一个细菌细胞穿透蛋白 (CPP)。我们发现重组YopM（rYopM）在自主易位后进入不同的真核细胞类型并下调几种促炎细胞因子（例如肿瘤坏死因子-α [TNF-α]）的表达。在感染小肠结肠炎耶尔森氏菌或转染宿主细胞后，YopM 与两种激酶核糖体 S6 蛋白激酶 (RSK) 和蛋白激酶 C 相关激酶 (PRK) 的亚型相互作用。由于干扰去磷酸化，这种相互作用导致 RSK 持续激活。在这里，我们通过免疫共沉淀证明 rYopM 在细胞渗透后与 RSK 和 PRK 相互作用。我们发现自主易位的 rYopM 与不同的 RSK 和 PRK 亚型形成三聚体复合物。此外，我们构建了一系列 rYopM 的截短版本，以映射形成复合物所需的结构域。 rYopM 的 C 末端被认为对于与 RSK1 的相互作用至关重要，而 rYopM 富含亮氨酸的重复结构域中的任何缺失都会消除 PRK2 的结合。此外，我们发现细胞穿透性 rYopM 与 RSK 的相互作用导致该激酶在丝氨酸 380 处的自磷酸化增强。最后，我们研究了三聚体 rYopM-RSK/PRK 复合物的下游信号传导是否调节促炎性 TNF-α 的表达。在这里，我们可以排除与 RSK1 和 PRK2 的相互作用对于 rYopM 的抗炎作用至关重要。
The effector protein Yersinia outer protein M (YopM) of Yersinia enterocolitica has previously been identified and characterized as the first bacterial cell-penetrating protein (CPP). We found that recombinant YopM (rYopM) enters different eukaryotic cell types and downregulates the expression of several pro-inflammatory cytokines (e. g., tumor necrosis factor-alpha [TNF-alpha]) after autonomous translocation. After infection with Y. enterocolitica or transfection of host cells, YopM interacts with isoforms of the two kinases ribosomal S6 protein kinase (RSK) and protein kinase C-related kinase (PRK). This interaction caused sustained RSK activation due to interference with dephosphorylation.Here we demonstrate by co-immunoprecipitation that rYopM interacts with RSK and PRK following cell-penetration. We show that autonomously translocated rYopM forms a trimeric complex with different RSK and PRK isoforms. Furthermore, we constructed a series of truncated versions of rYopM to map the domain required for the formation of the complex. The C-terminus of rYopM was identified to be essential for the interaction with RSK1, whereas any deletion in rYopM's leucin-rich repeat domains abrogated PRK2 binding. Moreover, we found that the interaction of cell-penetrating rYopM with RSK led to enhanced autophosphorylation of this kinase at serine 380. Finally, we investigated whether downstream signaling of the trimeric rYopM-RSK/PRK complex modulates the expression of pro-inflammatory TNF-alpha. Here, we could exclude that interaction with RSK1 and PRK2 is essential for the anti-inflammatory effects of rYopM.