铜绿假单胞菌分泌型Ndk促进宿主非小细胞肺癌转移的分子机制研究

铜绿假单胞菌（PA）是肺部感染最常见病原菌之一，非小细胞肺癌（NSCLC）患者尤易感染。研究表明抑制MST3-T190位点自身磷酸化可促进肿瘤迁移。课题组前期发现，PA感染时其分泌型Ndk可进入细胞并与MST3互作，且伴有MST3-T190位点磷酸化水平降低，提示PA感染可能通过Ndk介导的MST3途径促进NSCLC患者肿瘤转移。本项目拟采用截断及位点突变等技术，考察Ndk与MST3互作关键区域及位点；考察该互作对MST3自身磷酸化、激酶活性、胞内分布、稳定性及对癌细胞侵袭、迁移的影响。构建肺癌皮下移植瘤动物模型，考察PA感染及Ndk蛋白对小鼠肿瘤生长、转移的影响。此外，考察人Ndk同源蛋白NM23-H1/H2与MST3互作方式及生物学效应。结果将对PA致病性的认识提供全新资料，并为临床NSCLC患者尽量避免PA感染，控制肿瘤转移，提供指导作用。

Pseudomonas aureginosa (P. aureginosa) is one of the most prevalent pathogens in lung infection, especially in patients suffered from non-small cell lung cancer (NSCLC). Researches have shown that the inhbition of the phosphorylation of mammalian ste20-like protein kinase 3 (MST3) on tyrosine 190 (T190) promotes tumor metastasis. In our previous study, we have found that the nucleoside diphosphate kinases (Ndk) secreted from P. aureginosa can be translocated into the host cells upon infection, and it interacts with the MST3, which results in the inhibition of the phosphorylation of MST3 on T190. This result indicates that P. aureginosa might promote tumor metastasis of NSCLC patients through the Ndk-mediated regulation of MST3. To verify this hypothesis, we firstly propose to identify the critical binding domains and sites between Ndk and MST3 by constructing the truncated and site-directed mutants. Secondly, we aim to clarify the effects on auto phosphorylation, kinase activity, sub-cellular location and protein stability of MST3, and on the tumor invasion and metastasis upon the interaction between Ndk and MST3. Thirdly, we set out to investigate the effect on tumor growth and metastasis upon P. aureginosa infection or Ndk treatment in the subcutaneous xenograft models of lung carcinoma. Lastly, we are intended to investigate the interaction pattern among Ndk, MST3 and NM23-H1 or NM23-H2 (the homologous protein of P. aureginosa Ndk), and the biological effects upon their interaction. This investigation might shed new light on the pathogenesis of P. aureginosa, and might also provide a new guidance for the prevention of tumor metastasis of NSCLC patients in clinic.