【摘要】：病原微生物感染對(duì)人類健康構(gòu)成巨大威脅,先天免疫系統(tǒng)是生物體在長(zhǎng)期進(jìn)化過(guò)程中建立起來(lái)的天然保護(hù)系統(tǒng),是機(jī)體對(duì)抗外界病原體的第一道防線。VISA(MAVS,CARDIF,IPS-1)是連接胞漿dsRNA受體RIG-I與下游信號(hào)轉(zhuǎn)導(dǎo)通路的一個(gè)接頭蛋白,病毒入侵機(jī)體時(shí),VISA在病毒激發(fā)的IFN-β信號(hào)通路起關(guān)鍵作用,VISA通過(guò)招募RIG-I樣受體(包括RIG-I和MDA5)以及下游信號(hào)蛋白到線粒體上的VISA信號(hào)平臺(tái)上,激活轉(zhuǎn)錄因子IRF3和NF-κB,從而誘導(dǎo)IFNs和其他炎癥因子表達(dá)。研究結(jié)果表明VISA無(wú)論在TLR3非依賴的或者TLR3介導(dǎo)的抗病毒IFN信號(hào)途徑中都具有關(guān)鍵作用,但目前對(duì)VISA信號(hào)轉(zhuǎn)導(dǎo)的詳細(xì)機(jī)制還不十分清楚。更多VISA相互作用蛋白的發(fā)現(xiàn)以及它們之間的作用機(jī)制的研究能完善我們對(duì)VISA參與的信號(hào)轉(zhuǎn)導(dǎo)機(jī)制的認(rèn)識(shí)。本研究利用酵母雙雜交系統(tǒng),用VISA蛋白的全長(zhǎng)做誘餌(Bait)篩選293T細(xì)胞cDNA表達(dá)文庫(kù),并結(jié)合免疫共沉淀實(shí)驗(yàn),雙熒光素酶報(bào)告基因系統(tǒng)驗(yàn)證篩選到的陽(yáng)性克隆和VISA作用的真實(shí)性。本研究成功篩選到Sec13L1候選基因并驗(yàn)證了其與VISA的全長(zhǎng)蛋白相互作用,在293T細(xì)胞中過(guò)表達(dá)這個(gè)基因,我們發(fā)現(xiàn)Sec13L1過(guò)表達(dá)能促進(jìn)VISA對(duì)IFNβ的誘導(dǎo),并存在劑量效應(yīng)。
[Abstract]:The infection of pathogenic microorganisms poses a great threat to human health. The innate immune system is a natural protective system established by organisms in the long evolution process and the first line of defense against external pathogens,. VISA (MAVS,CARDIF,. IPS-1) is a junction protein linking cytoplasmic dsRNA receptor RIG-I and downstream signal transduction pathway. VISA plays a key role in the IFN- 尾 signaling pathway stimulated by virus when the virus invades the organism. VISA induces the expression of IFNs and other inflammatory factors by recruiting RIG-I like receptors (including RIG-I and MDA5) and downstream signal proteins to the VISA signal platform on mitochondria to activate the transcription factors IRF3 and NF- 魏 B. The results show that VISA plays a key role in TLR3 independent or TLR3 mediated antiviral IFN signaling pathway, but the detailed mechanism of VISA signal transduction is not well understood. The discovery of more VISA interacting proteins and the study of their interaction mechanism can improve our understanding of the signal transduction mechanism in which VISA is involved. In this study, yeast two-hybrid system was used to screen 293T cell cDNA expression library using full-length (Bait) of VISA protein as bait, and combined with co-immunoprecipitation test. Double luciferase reporter gene system was used to verify the authenticity of the screened positive clones and the role of VISA. In this study, the candidate gene of Sec13L1 was successfully screened and its interaction with the full-length protein of VISA was verified. The gene was overexpressed in 293T cells. We found that overexpression of Sec13L1 promoted the induction of IFN 尾 by VISA, and there was a dose effect.
【學(xué)位授予單位】：江西師范大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R392

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