【摘要】：對于自己和非己的識別是機體免疫反應的基礎,而完成這一功能的分子被稱作模式識別受體(pattern recognition receptor, PRRs)。C型凝集素(C-type lectins)作為一類新型的模式識別分子,近年來逐漸顯示出在病原體識別中的重要作用。同時,某些C型凝集素與配體結(jié)合后可以產(chǎn)生免疫抑制信號,因而許多病原微生物包括艾滋病毒、丙肝病毒等借此以逃避機體的免疫監(jiān)視,但迄今對其胞內(nèi)的信號轉(zhuǎn)導途徑仍不清楚,此方面的研究引起了人們的高度關注。LSECtin(liver and lymph node sinusoidal endothelial cells C-type lectin)是本室鑒定的一個新的C型凝集素,特異表達在肝臟和淋巴結(jié)竇內(nèi)皮細胞上,具有糖基結(jié)合能力,但是其胞內(nèi)的信號轉(zhuǎn)導途徑尚未揭示,其生理功能和分子機制了解甚少,我們對此開展了研究。 本文首先通過酵母雙雜交方法篩選LSECtin的胞內(nèi)區(qū)結(jié)合蛋白,并選擇了四對可能存在的相互作用在酵母中進行了初步驗證,證明均真實存在。繼而在綜合比較結(jié)合力、功能相關性、亞細胞定位的基礎上,選取了其中兩個蛋白IFP35(interferon-induced protein 35)和TRIP-1(TGF-β type Ⅱ receptor interacting protein-1)在哺乳動物細胞中進行進一步驗證。熒光定位實驗表明,當單獨表達時,LSECtin為質(zhì)膜上的不均勻分布和細胞質(zhì)中的少量點狀分布,IFP35為細胞質(zhì)中的點狀或彌散狀分布,TRIP-1在質(zhì)膜和細胞質(zhì)中均有分布。而當IFP35或TRIP-1分別與LSECtin共表達時,二者均被募集到LSECtin高表達的區(qū)域,并且存在明顯的共定位。同時,免疫共沉淀實驗也證明二者確與LSECtin存在相互作用。此外,我們對LSECtin與TRIP-1的相互作用的生物學意義進行了探索。報告基因?qū)嶒烇@示,LSECtin對TGF-β誘導的靶基因PAI-1表達具有抑制作用,而同家族的DC-SIGNR(DC-SIGN related gene)沒有此功能,可能與其特有的生理功能相關。這些結(jié)果提示,LSECtin的胞內(nèi)信號轉(zhuǎn)導可能與TGF-β和干擾素途徑有關。同時,為闡明LSECtin的生理功能,我們還開展了小鼠LSECtin的基因打靶工作,完成了mLSECtin基因組序列的篩選。
[Abstract]:The recognition of self and non-self is the basis of the body's immune response, and the molecule that completes this function is called the pattern recognition receptor (pattern recognition receptor, PRRs). C lectin (C-type lectins) as a new type of pattern recognition molecule. In recent years, it has gradually shown the important role in pathogen recognition. At the same time, some type C lectin binding with ligands can produce immunosuppressive signals, so many pathogenic microorganisms, including HIV, hepatitis C virus and so on, can escape the body's immune surveillance. However, the intracellular signal transduction pathway is still unclear, and this study has aroused great concern that. LSECtin (liver and lymph node sinusoidal endothelial cells C-type lectin is a new type C lectin identified in our laboratory. Specifically expressed in liver and lymph node sinusoidal endothelial cells, it has glycosyl binding ability, but its intracellular signal transduction pathway has not been revealed, and its physiological function and molecular mechanism are not well understood. In this paper, the intracellular binding proteins of LSECtin were screened by yeast two-hybrid method, and four pairs of possible interactions were selected to verify the existence of these proteins in yeast. On the basis of comprehensive comparison of binding ability, functional correlation and subcellular localization, two proteins IFP35 (interferon-induced protein 35) and TRIP-1 (TGF- 尾 type 鈪，

本文編號：2150153