VISA相互作用蛋白的篩選與鑒定
發(fā)布時間：2018-11-09 21:33
【摘要】：病原微生物感染對人類健康構(gòu)成巨大威脅,先天免疫系統(tǒng)是生物體在長期進化過程中建立起來的天然保護系統(tǒng),是機體對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與下游信號轉(zhuǎn)導通路的一個接頭蛋白,病毒入侵機體時,VISA在病毒激發(fā)的IFN-β信號通路起關(guān)鍵作用,VISA通過招募RIG-I樣受體(包括RIG-I和MDA5)以及下游信號蛋白到線粒體上的VISA信號平臺上,激活轉(zhuǎn)錄因子IRF3和NF-κB,從而誘導IFNs和其他炎癥因子表達。研究結(jié)果表明VISA無論在TLR3非依賴的或者TLR3介導的抗病毒IFN信號途徑中都具有關(guān)鍵作用,但目前對VISA信號轉(zhuǎn)導的詳細機制還不十分清楚。更多VISA相互作用蛋白的發(fā)現(xiàn)以及它們之間的作用機制的研究能完善我們對VISA參與的信號轉(zhuǎn)導機制的認識。本研究利用酵母雙雜交系統(tǒng),用VISA蛋白的全長做誘餌(Bait)篩選293T細胞cDNA表達文庫,并結(jié)合免疫共沉淀實驗,雙熒光素酶報告基因系統(tǒng)驗證篩選到的陽性克隆和VISA作用的真實性。本研究成功篩選到Sec13L1候選基因并驗證了其與VISA的全長蛋白相互作用,在293T細胞中過表達這個基因,我們發(fā)現(xiàn)Sec13L1過表達能促進VISA對IFNβ的誘導,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.
【學位授予單位】：江西師范大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R392
本文編號：2321614
[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.
【學位授予單位】：江西師范大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R392
【參考文獻】
相關(guān)期刊論文 前4條
1 楊文婷;唐麗;;模式識別受體及其功能[J];細胞與分子免疫學雜志;2013年08期
2 葉志云;張婷婷;賀穎;陳巧玉;劉琪;;酵母雙雜交技術(shù)篩選與Axin相互作用的蛋白質(zhì)[J];廈門大學學報(自然科學版);2009年03期
3 閆鴻斌;高閃電;賈萬忠;才學鵬;;細胞內(nèi)模式識別受體研究進展[J];免疫學雜志;2008年03期
4 麥瑞琴,許麗艷,方王楷,韓溟,牛永東,王子良,李恩民;酵母雙雜交篩選NGAL相互作用蛋白[J];腫瘤防治雜志;2005年04期
，本文編號：2321614
本文鏈接：http://sikaile.net/yixuelunwen/jichuyixue/2321614.html
最近更新
教材專著
