生命科學院: 生化科學研究所指導教授: 蔡明道魏同佑Wei, Tong-You WadeTong-You WadeWei2017-03-022018-07-062017-03-022018-07-062016http://ntur.lib.ntu.edu.tw//handle/246246/272315NF-κB控制著各方面的免疫反應，並調控著细胞的存活，增殖，和分化。由於其功能的多樣性，NF-κB的失調已經被發現與許多疾病有關連。我們先前的研究指出，TIFA蛋白質的的多聚化現象需要第九號蘇胺酸殘基的磷酸化， 進而活化NF-κB訊息調控路徑。此外，我們我們發現有與PI3K-Akt信號傳遞途徑相關的激酶參與磷酸化第九號蘇胺酸殘基。 在這項研究中，我們找到了上游能夠對TIFA蛋白質第九號蘇胺酸磷酸化的磷酸化激酶Aurora A， 在急性骨髓性白血病中，我們也證實前人所提出的Aurora A激酶活化NF-κB訊息傳遞路徑是由於TIFA蛋白質所調控的。 我們發現TIFA蛋白質表現量在急性骨髓性白血病中與Aurora A及NF-κB所調控的腫瘤存活因子的表現量有高度正相關性，且與病人的預後表現呈現負相關。我們進一步發現，抑制活體急性骨髓性白血病細胞中的TIFA表現量能夠擾亂白血病細胞激素的分泌，顯著地提昇化學治療的敏感度，並且提高人類急性骨髓性白血病細胞在裸鼠模式中的清除率。這些結果共同顯示，TIFA可以支持的AML的發展，並且標靶TIFA可以提高治療AML的效果。 我們也發現TIFA蛋白在肝癌細胞株與肝癌病人細胞中過度表達。而且越高的TIFA表現量，患者的無疾病存活期越短。這可能與TIFA調控上皮間質轉化的訊息傳遞作為肝癌腫瘤侵襲和轉移的分子機制有關。我們也發現了抑制TIFA可以特別地降低肝癌細胞株的細胞活性，提高化學治療毒性，並抑制其移動和侵襲的能力。這些結果顯示TIFA在未來將是一個嶄新的肝癌化學治療標把。 另一方面，在血管內皮細胞面臨促氧化和炎性刺激時，先天免疫反應主要主要是受到類鐸受體調控的NF-κB的訊息調控路徑所活化。我們發現ＴIFA在內皮細胞中是一個新穎的NLRP3炎性體啟動（訊息一）與活化（訊息二）的調控者。氧化和發炎的壓力例如剪切應力與氧化修飾的低密度脂蛋白皆會誘導並活化TIFA。 在訊息一中的炎性細胞激素和炎性體組成成分的轉錄活性中，TIFA的誘導被認為是必需的。 此外，Akt增加TIFA蛋白質第九號蘇胺酸的磷酸化並促使NLRP3炎性體組裝，可視為訊息二的炎性體活化。這些結果顯示TIFA透過訊息一以及訊息二來啟動與活化NLRP3炎性體，使其在內皮先天免疫反應扮演一個關鍵的調節者。 此研究所發現的機制提供了重要的轉譯意義，尤其是在哺乳動物細胞中的炎性反應。我們所得到的整體的結果不僅對腫瘤壞死因子到NF-κB的訊息傳遞有了詳細的解釋，並且在免疫疾病和癌症的治療中提供了潛在的治療靶標。Nuclear factor-κB (NF-κB) controls various aspects of immune responses and regulates cell survival, proliferation, and differentiation. In light of its functional diversity, dysregulation of NF-κB has been linked to a variety of diseases. Our previous study demonstrated that Thr9 phosphorylation-dependent oligomerization of TRAF-interacting protein with a FHA domain (TIFA) triggers the activation of NF-κB. In addition, we investigated the involvement of the kinase in PI3K-Akt signaling pathway is response for the Thr9 phosphorylation. In this study, we identified that Aurora A is an essential kinase for the Thr9 phosphorylation of TIFA, and that TIFA functionally mediates the Aurora A-driven NF-κB survival pathway in acute myeloid leukemia (AML). We found that TIFA protein is overexpressed concurrently with Aurora A and NF-κB signaling factors in de novo AML patients relative to healthy individuals, which is also correlated with the poor prognosis of patients. In addition, TIFA inhibition perturbs leukemic cytokine secretion, significantly enhances chemotoxicity, and potentiates the clearance of leukemic myeloblasts in a xenograft model. These results collectively demonstrate that TIFA may support AML progression, and that targeting TIFA can enhance therapeutic efficacies in the treatment of AML. In accord, we showed that TIFA protein is also overexpressed in hepatocellular carcinoma (HCC) lines and patient cells relative to normal ones. Higher TIFA expression showed significantly shorter disease free survival (DFS) than those with lower TIFA expression in HCC patients. This may correlate with TIFA directed epithelial-mesenchymal transition (EMT) signaling as the molecular mechanism underlying tumor invasion and metastasis in HCC. We also found that silencing of TIFA specifically reduces viability, enhances the chemotoxicity, and retards the migration and invasion abilities of HCC lines. These results may propose TIFA as a novel therapeutic target in the treatment of HCC. On the other hand, toll-like receptor-mediated NF-κB activation is a major innate immune response in vascular endothelial cells (ECs) in response to pro-oxidative and inflammatory stimuli. We identified TIFA as a novel regulator of both priming (Signal 1) and activating (Signal 2) signals of NOD-like receptor pyrin domain-containing protein 3 (NLRP3) inflammasome in ECs. Oxidative and inflammatory stresses such as atheroprone flow and oxidized-LDL induce and activate TIFA. Induction of TIFA is required for the transcriptional activation of inflammatory cytokines and inflammasome components which is considered Signal 1. Additionally, Akt-enhanced phosphorylation of TIFA Thr9 promotes the assembly of NLRP3 inflammasome which is considered Signal 2 of inflammasome activation. The results suggest that TIFA is a crucial mediator in the endothelial innate immune response by potentiating and amplifying NLRP3 inflammasome via augmenting Signal 1 and 2. This newly defined mechanism has important translational implications, particularly toward the inflammatory responses in mammalian cells. The overall results obtained in this study are expected to not only decipher the detailed molecular mechanism within TNF and NF-κB axis, but also provide a potential therapeutic target in the treatment of immune disorders and cancers.25879208 bytesapplication/pdf論文公開時間: 2021/8/3論文使用權限: 同意有償授權(權利金給回饋本人)NF-κBTIFAAurora A急性骨髓性白血病肝癌藥物抗性上皮-間質轉化AktNLRP3炎性體AMLHCCchemoresistanceEMTNLRP3 inflammasome[SDGs]SDG3thesis10.6342/NTU201601470http://ntur.lib.ntu.edu.tw/bitstream/246246/272315/1/ntu-105-D99b46010-1.pdf