2009年8月3日，北京生命科学研究所邵峰博士实验室在Proc Natl Acad Sci USA杂志在线发表题为题为“A Legionella type IV effector activates the NF-κB pathway by phosphorylating the IκB family of inhibitors”的文章。该文章首次报道了肺炎军团菌通过其四型分泌系统向宿主细胞注入一个新的能激活宿主中具有抗凋亡作用的NF-κB免疫炎症调节信号通路及其具体分子作用机制。
肺炎军团菌（Legionella pneumophila）感染肺泡巨噬细胞，并在宿主细胞内大量繁殖和裂解细胞，随后扩散侵袭其它的巨噬细胞产生新一轮的感染最终引发严重的肺炎。肺炎军团菌通过其四型分泌系统分泌毒力效应蛋白分子进入宿主细胞内，进而阻断或调节宿主免疫防御相关的信号通路。目前关于肺炎军团菌四型分泌系统效应蛋白直接调节宿主免疫信号通路的研究还鲜有报道。
邵峰小组在这篇文章中筛选了100多个可能的军团菌四型分泌系统效应蛋白，发现其中只有一个叫做LegK1的蛋白，在导入真核细胞后显示出非常强的激活NF-κB信号通路的活性。同时，LegK1对其它包括MAPK激酶和干扰素(IFNb)在内的相关免疫信号通路没有激活作用。LegK1编码了一个类真核的丝氨酸/苏氨酸蛋白激酶。作者也证明了LegK1的激酶活性对激活NF-κB信号通路是必不可少的，并且LegK1确实能通过肺炎军团菌四型分泌系统被注入到宿主巨噬细胞内。IKK激酶复合物介导的IκB家族蛋白的磷酸化是NF-κB信号通路激活的关键步骤。邵峰小组通过RNA干扰的方法发现knockdown 诸多已知的IKK上游的信号分子对LegK1激活NF-κB信号通路没有影响。作者进一步发现LegK1在IKK遗传缺失的细胞中仍然能够激活NF-κB信号通路。这些结果在后续的基于细胞提取物的无细胞体系重组实验中也得到了验证。深入的生物化学研究表明LegK1能够和宿主中的IKK激酶一样直接磷酸化IκBa蛋白的32位和36位的丝氨酸，从而导致IκBa的泛素化和降解，进而释放NF-κB进入细胞核内并激活转录。有趣并值得注意的是，NFκB2（也叫p100）蛋白在生化上也属于IκB家族，同样受IKK磷酸化调节，但NFκB2介导的是与天然免疫不相关的非经典NF-κB信号通路。作者发现LegK1也能磷酸化NFκB2并直接诱导其成熟为活性形式的p52蛋白。根据上述实验结果，作者认为肺炎军团菌四型分泌系统效应蛋白LegK1是模拟了宿主的IKK激酶来激活宿主的NF-κB信号通路，并进一步推测这种激活作用可能起到了抗凋亡作用使得宿主细胞在感染后不会立刻启动具有保护性的凋亡程序。
这篇文章不仅揭示了肺炎军团菌如何通过其四型分泌系统来调节宿主NF-κB免疫信号通路的具体分子作用机制，同时LegK１也是目前为止第一个报道的能直接激活宿主NF-κB信号通路的病原菌效应蛋白分子。（生物谷Bioon.com）
生物谷推荐原始出处：
PNAS August 3, 2009, doi: 10.1073/pnas.0907200106
A Legionella type IV effector activates the NF-κB pathway by phosphorylating the IκB family of inhibitors
Jianning Gea,b,1, Hao Xua,b,1, Ting Lib, Yan Zhoua,b, Zhibin Zhanga,b, Shan Lib, Liping Liub and Feng Shaob,2
aCollege of Life Sciences, Beijing Normal University, Beijing 100875, China; and
bNational Institute of Biological Sciences, Beijing 102206, China
NF-κB is critical in innate immune defense responses against invading microbial pathogens. Legionella pneumophila infection of lung macrophages causes Legionnaire's disease with pneumonia symptoms. A set of NF-κB-controlled genes involved in inflammation and anti-apoptosis are up-regulated in macrophages upon L. pneumophila infection in a Legionella Dot/Icm type IV secretion system-dependent manner. Among ≈100 Dot/Icm substrates screened, we identified LegK1 as the sole Legionella protein that harbors a highly potent NF-κB-stimulating activity. LegK1 does not affect MAPK and IFN pathways. Activation of the NF-κB pathway by LegK1 requires its eukaryotic-like Ser/Thr kinase activity and is independent of upstream components in the NF-κB pathway, including TRAFs, NIK, MEKK3, and TAK1. Cell-free reconstitution revealed that LegK1 stimulated NF-κB activation in the absence of IKKα and IKKβ, and LegK1 efficiently phosphorylated IκBα on Ser-32 and Ser-36 both in vitro and in cells. LegK1 seems to mimic the host IKK as LegK1 also directly phosphorylated other IκB family of inhibitors including p100 in the noncanonical NF-κB pathway. Phosphorylation of p100 by LegK1 led to its maturation into p52. Thus, LegK1 is a bacterial effector that directly activates the host NF-κB signaling and likely plays important roles in modulating macrophage defense or inflammatory responses during L. pneumophila infection.
