本文選題：鼻咽癌　切入點：14-3-3σ　出處：《中南大學(xué)》2010年博士論文　論文類型：學(xué)位論文

【摘要】： 鼻咽癌(nasopharyngeal carcinoma, NPC)是我國南方最常見的惡性腫瘤之一,嚴重威脅我國人民的生命和健康。絕大多數(shù)NPC分化差,惡性程度高,易早期發(fā)生頸淋巴結(jié)和遠處轉(zhuǎn)移。NPC轉(zhuǎn)移是一個多基因參與的過程,多個基因之間通過相互作用形成關(guān)鍵性的節(jié)點(node),這些節(jié)點在NPC轉(zhuǎn)移中具有重要作用。 我們前期實驗采用蛋白質(zhì)組學(xué)方法篩選NPC組織差異表達的蛋白質(zhì),鑒定了36個NPC組織的差異表達蛋白質(zhì),采用免疫組織化學(xué)染色檢測差異蛋白質(zhì)14-3-3σ(14-3-3 sigma)在正常鼻咽上皮組織、NPC組織以及頸淋巴結(jié)轉(zhuǎn)移NPC組織中的表達情況。結(jié)果發(fā)現(xiàn)：在NPC組織中存在高頻率的14-3-3σ蛋白表達缺失/下調(diào),而且其表達缺失/下調(diào)與NPC轉(zhuǎn)移相關(guān),這提示14-3-3σ表達缺失/下調(diào)在NPC轉(zhuǎn)移中可能具有重要作用,但14-3-3σ表達缺失/下調(diào)促進NPC轉(zhuǎn)移的分子機制尚未闡明。 為了研究14-3-3σ在NPC轉(zhuǎn)移中的作用機制,本研究采用靶向蛋白質(zhì)組學(xué)方法(免疫共沉淀結(jié)合質(zhì)譜分析)分離鑒定NPC細胞中14-3-3σ的相互作用蛋白,采用基因本體論(GO)、功能聚類、信號通路和蛋白—蛋白相互作用(protein-protein interaction, PPI)等生物信息學(xué)方法分析14-3-3σ的相互作用蛋白,揭示其生物學(xué)意義,并對關(guān)鍵相互作用蛋白(14-3-3σ/EGFR/Keratin 8)進行實驗驗證和功能研究。 主要結(jié)果如下： 1、采用靶向蛋白質(zhì)組學(xué)在NPC細胞中鑒定了111個14-3-30的相互作用蛋白,并對其中4個相互作用蛋白質(zhì)(Keratin 8、EGFR、RAB7和p53)進行了驗證。 2、GO和聚類分析顯示：111個14-3-3σ的相互作用蛋白可聚類成13功能相關(guān)的簇,這些蛋白的功能主要歸類為8個方面：細胞骨架、跨膜轉(zhuǎn)運、分子伴侶、核糖體蛋白、GTPase、蛋白轉(zhuǎn)運、ATPase和RNA結(jié)合。 3、KEGG和Biocarta信號通路分析顯示：14-3-3σ的相互作用蛋白涉及3條Biocarta信號通路和6條KEGG信號通路,其中16個相互作用蛋白涉及到細胞骨架信號通路,5個相互作用蛋白涉及到囊泡轉(zhuǎn)運信號通路。 4、蛋白相互作用分析顯示：14-3-3σ/p53/RAB7相互作用組(Interactome)調(diào)控囊泡轉(zhuǎn)運,14-3-3σ/EGFR/CK8相互作用組調(diào)控細胞骨架,它們可能與NPC轉(zhuǎn)移相關(guān)。 5、實驗證實在NPC細胞中存在14-3-3σ/EGFR/Keratin 7相互作用組。6、功能分析顯示：14-3-3σ下調(diào)能增加NPC細胞EGFR和Keratin 8表達,增強NPC細胞的體外侵襲能力。 研究結(jié)果提示,14-3-3σ表達下調(diào)可能通過14-3-3σ/EGFR/Keratin8相互作用組而促進NPC侵襲轉(zhuǎn)移。
[Abstract]:Nasopharyngeal carcinoma (NPCs) is one of the most common malignant tumors in southern China, which is a serious threat to the lives and health of our people. The early occurrence of cervical lymph node and distant metastasis. NPC metastasis is a process in which multiple genes interact to form a key node, which plays an important role in NPC metastasis. We used proteomics method to screen differentially expressed proteins in NPC tissues and identified the differentially expressed proteins in 36 NPC tissues. Immunohistochemical staining was used to detect the expression of differential protein 14-3-3 sigma in normal nasopharyngeal epithelial tissues and NPC tissues with cervical lymph node metastasis. The results showed that there were high frequency deletion / down-regulation of 14-3-3 蟽 protein expression in NPC tissues. The deletion / down-regulation of 14-3-3 蟽 expression may play an important role in NPC metastasis, but the molecular mechanism of 14-3-3 蟽 expression deletion / down-regulation promoting NPC metastasis has not been elucidated. In order to study the mechanism of 14-3-3 蟽 in NPC transfer, we used targeted proteomics (immunoprecipitation mass spectrometry) to isolate and identify 14-3-3 蟽 interacting proteins in NPC cells. Signal pathway and protein-protein interaction protein-protein interaction (PPI) were used to analyze 14-3-3 蟽 interaction proteins and reveal their biological significance. The key interacting proteins 14-3-3 蟽 / EGFR / Keratin 8 were tested and their functions were studied. The main results are as follows:. 1. 111 14-3-30 interacting proteins were identified by targeting proteomics in NPC cells, and four of them, Keratin 8G EGFRN RAB7 and p53, were identified. 2go and cluster analysis showed that 111 14-3-3 蟽 interacting proteins could be clustered into 13-functional clusters. The functions of these proteins were mainly classified into eight aspects: cytoskeleton, transmembrane transport, molecular chaperone. Ribosomal protein GTPase, protein transporter ATPase and RNA binding. 3Analysis of the KEGG and Biocarta signaling pathways showed that the interaction protein of 1: 14-3-3 蟽 involved three Biocarta signaling pathways and six KEGG signaling pathways. Sixteen interacting proteins were involved in cytoskeletal signaling pathways and five involved in vesicular transport signaling pathways. 4. The protein interaction analysis showed that the cytoskeleton was regulated by the interaction group of 14-3-3 蟽 -p53 / RAB7. The cytoskeleton was regulated by the 14-3-3 蟽 -EGFR / CK8 interaction group, which might be related to NPC metastasis. 5. The results showed that there were 14-3-3 蟽 -EGFR / Keratin 7 interactions in NPC cells. Functional analysis showed that the down-regulation of w _ (14-3-3) 蟽 could increase the expression of EGFR and Keratin _ 8 in NPC cells and enhance the invasiveness of NPC cells in vitro. The results suggest that the down-regulation of 14-3-3 蟽 expression may promote the invasion and metastasis of NPC through 14-3-3 蟽 / EGFR / Keratin8 interaction.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2010
【分類號】：R739.6

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本文編號：1559453