發(fā)布時(shí)間：2018-06-17 08:29

  本文選題：長(zhǎng)牡蠣 + 免疫球蛋白超家族�。� 參考：《中國(guó)科學(xué)院研究生院(海洋研究所)》2016年博士論文

【摘要】：免疫球蛋白超家族(IgSF)是所有含有免球蛋白樣三維拓?fù)浣Y(jié)構(gòu)的蛋白分子的群體集合。脊椎動(dòng)物中含有基于免疫球蛋白的適應(yīng)性免疫,無(wú)脊椎動(dòng)物雖然缺乏高度可變的免疫球蛋白,但已有研究表明無(wú)脊椎動(dòng)物中具有類似適應(yīng)性免疫的“選擇適應(yīng)性免疫”(alternative adaptive immunity),無(wú)脊椎動(dòng)物中的免疫球蛋白超家族(IgSF)成員分子可能在無(wú)脊椎動(dòng)物的“選擇適應(yīng)性免疫”應(yīng)答中發(fā)揮重要作用。目前對(duì)于軟體動(dòng)物IgSF成員結(jié)構(gòu)和免疫功能的了解非常有限。本文采用生物信息學(xué)、分子生物學(xué)、免疫學(xué)等手段,鑒定并分析了長(zhǎng)牡蠣I(yíng)g SF分子的結(jié)構(gòu)和進(jìn)化特征,探討了IgSF分子在長(zhǎng)牡蠣個(gè)體發(fā)育和免疫微生物(河口弧菌、溶藻弧菌、塔氏弧菌、燦爛弧菌和鰻弧菌)和PAMPs(LPS、PGN、GLU和poly(I:C))刺激的表達(dá)模式,查明了幾種IgSF成員分子(CgJAM-A-L、CgSiglec-1和CgNCAM)的結(jié)構(gòu)與免疫學(xué)功能。通過結(jié)構(gòu)域分析,從長(zhǎng)牡蠣中共鑒定得到了268個(gè)IgSF成員分子,分別含有1-36個(gè)Ig結(jié)構(gòu)域,其中162個(gè)基因由Ig結(jié)構(gòu)域串聯(lián)重復(fù)而成,而其余106個(gè)基因(39.6%)則由Ig結(jié)構(gòu)域與其它結(jié)構(gòu)域重組而成。注釋分析提示長(zhǎng)牡蠣I(yíng)gSF主要具有識(shí)別和粘附的功能,其中共有217個(gè)IgSF成員分子被預(yù)測(cè)具有識(shí)別和結(jié)合的功能。HMM-3D功能聚類的分子結(jié)果對(duì)GO注釋的結(jié)果做了進(jìn)一步的補(bǔ)充,將部分在Blast2GO軟件中未能成功注釋基因與其他明確注釋基因聚類到了一起。根據(jù)已報(bào)道海洋動(dòng)物基因組中IgSF的進(jìn)化分析發(fā)現(xiàn),IgSF家族體現(xiàn)出以海綿動(dòng)物、棘皮動(dòng)物和半索動(dòng)物為節(jié)點(diǎn)的階段性擴(kuò)張趨勢(shì)。轉(zhuǎn)錄組分析發(fā)現(xiàn)PAMPs和弧菌刺激后表達(dá)量顯著升高的IgSF分子數(shù)量分別達(dá)到116和166個(gè),提示長(zhǎng)牡蠣I(yíng)gSF可能在免疫應(yīng)答反應(yīng)中發(fā)揮重要功能。發(fā)現(xiàn)大量的IgSF在免疫響應(yīng)中具有高度的特異性,且含較少Ig結(jié)構(gòu)域的IgSF分子功能多樣性更為豐富,表明長(zhǎng)牡蠣I(yíng)gSF在進(jìn)化擴(kuò)張過程中存在結(jié)構(gòu)和功能的分化。在燦爛弧菌刺激的轉(zhuǎn)錄組分析中篩選獲得大量的IgSF單堿基突變和可變剪接,并且有大量突變位點(diǎn)具備對(duì)燦爛弧菌二次刺激的特異性,暗示長(zhǎng)牡蠣I(yíng)gSF可能通過點(diǎn)突變和可變剪接產(chǎn)生多樣性,且點(diǎn)突變形成的新個(gè)體變異型可能參與“選擇適應(yīng)性免疫”應(yīng)答。另外通過分析長(zhǎng)牡蠣不同時(shí)期的轉(zhuǎn)錄組數(shù)據(jù)發(fā)現(xiàn),部分長(zhǎng)牡蠣igsf能夠在特定發(fā)育時(shí)期上調(diào)表達(dá),可能參與母源免疫和卵裂期、胚胎期、幼蟲期和稚貝型的發(fā)育調(diào)控。非跨膜型的cgjam-a-l是在長(zhǎng)牡蠣中發(fā)現(xiàn)的一個(gè)jam家族分子同源基因,含有三個(gè)串聯(lián)的i-set型ig結(jié)構(gòu)域。它能夠識(shí)別并結(jié)合多種pamps(lps、pgn、man、lta、poly(i:c)和glu)和微生物(燦爛弧菌、鰻弧菌、大腸桿菌、金黃色葡萄球菌、藤黃微球菌、耶羅維亞酵母和畢赤酵母),并在長(zhǎng)牡蠣的固有免疫應(yīng)答中發(fā)揮調(diào)理作用。同時(shí)cgjam-a-l的結(jié)構(gòu)進(jìn)化信息表明了jam家族分子與抗體分子可能具有一個(gè)多功能的“祖先基因”,并在進(jìn)化中實(shí)現(xiàn)了功能的特異性分化,從而分別形成jam和抗體分子�？缒ば偷腸gsiglec-1基因包含兩個(gè)ig結(jié)構(gòu)域和兩個(gè)胞內(nèi)的受體酪氨酸抑制基序(itim),并在長(zhǎng)牡蠣的免疫識(shí)別和誘導(dǎo)下游級(jí)聯(lián)反應(yīng)中發(fā)揮連接作用。它在各組織中廣泛分布,并且血淋巴細(xì)胞中的轉(zhuǎn)錄本能夠在燦爛弧菌刺激后顯著地上調(diào)。cgsiglec-1能夠結(jié)合多種pamps(psias、lps和pgn),并且通過抑制下游信號(hào)通路,在血淋巴細(xì)胞的凋亡、吞噬和細(xì)胞因子的釋放中發(fā)揮調(diào)節(jié)作用。跨膜型的cgncam包含五個(gè)ig結(jié)構(gòu)域、兩個(gè)fniii結(jié)構(gòu)域和一個(gè)跨膜區(qū),并可能在長(zhǎng)牡蠣的固有免疫中發(fā)揮模式識(shí)別作用。它與一種植物的凝集素具有較高的相似性,并且在微生物刺激后,血淋巴細(xì)胞中的mrna能夠顯著上調(diào)。重組的rcgncam蛋白具有對(duì)多種pamps(lps、glu和man)和微生物(燦爛弧菌、鰻弧菌、大腸桿菌、藤黃微球菌、耶羅維亞酵母和畢赤酵母)的廣譜識(shí)別能力。同時(shí)cgncam還可能在長(zhǎng)牡蠣extracellulartraps的形成中發(fā)揮重要的作用。三中不同類型的igsf共同說(shuō)明長(zhǎng)牡蠣igsf的主要功能是識(shí)別和粘附,游離型的igsf可能發(fā)揮調(diào)理分子的作用,而跨膜型igsf能夠作為抗原分子的表面受體并調(diào)節(jié)下游的免疫通路。綜上所述,長(zhǎng)牡蠣中的igsf數(shù)目眾多,種類豐富,并能夠在免疫應(yīng)答中發(fā)揮重要作用。長(zhǎng)牡蠣igsf可能通過點(diǎn)突變和可變剪接產(chǎn)生基因的多樣性,且新形成的個(gè)體變異型可能參與“選擇適應(yīng)性免疫”應(yīng)答。igsf成員分子cgjam-a-l、cgsiglec-1和cgncam能識(shí)別結(jié)合多種外源微生物,并分別發(fā)揮調(diào)理分子、免疫調(diào)節(jié)分子和表面受體的作用。該結(jié)果探索了無(wú)脊椎動(dòng)物中igsf分子與高等動(dòng)物的異質(zhì)性,初步揭示無(wú)脊椎動(dòng)物IgSF分子在“選擇適應(yīng)性免疫”中的作用,為進(jìn)一步研究IgSF分子的進(jìn)化提供了分子生物學(xué)證據(jù),豐富和發(fā)展了海洋無(wú)脊椎動(dòng)物免疫學(xué)內(nèi)容。
[Abstract]:The immunoglobulin superfamily (IgSF) is a collection of all protein molecules containing a globulin like three-dimensional topology. Vertebrates contain adaptive immunity based on immunoglobulin, and invertebrates, although lack of highly variable immunoglobulin, have shown that invertebrates have similar adaptive immunity. "Alternative adaptive immunity", the immunoglobulin superfamily (IgSF) members of invertebrates may play an important role in the "selective adaptive immune response" response to invertebrates. At present, the understanding of the IgSF member structure and immune function of mollusks is very limited. Informatics, molecular biology and immunology have been used to identify and analyze the structure and evolution characteristics of Ig SF in oysters, and to explore the expression patterns of IgSF molecules in the oysters' ontogeny and immune microorganism (Vibrio estuarine, Vibrio alginolyticus, Vibrio TA, Vibrio resplenden and Vibrio anguinae) and PAMPs (LPS, PGN, GLU and poly (I:C)). Structural and immunological functions of several IgSF members (CgJAM-A-L, CgSiglec-1 and CgNCAM). Through domain analysis, 268 IgSF members were identified from long oysters, containing 1-36 Ig domains, of which 162 genes were repeated in series with the Ig domain, while the rest of the 106 genes (39.6%) were from Ig domain and other junctions. Annotated analysis suggests that oysters IgSF mainly have the function of identification and adhesion, in which a total of 217 IgSF members are predicted to have a functional cluster of functional.HMM-3D, which is identified and combined. The results of the GO annotation are further supplemented by the failure to annotate the genes in the Blast2GO software. According to the reported evolutionary analysis of IgSF in the genome of marine animals, it was reported that the IgSF family showed a phased expansion of the node with spongy, eanthoderma and semi cord animals. The number of IgSF molecules with a significant increase in the expression of PAMPs and Vibrio stimulated by the transcriptional analysis was 11, respectively. 6 and 166, suggesting that the long oyster IgSF may play an important role in the immune response. It is found that a large number of IgSF are highly specific in the immune response, and the functional diversity of the IgSF molecules with less Ig domains is more abundant, indicating the structural and functional differentiation of the long oyster IgSF in the process of evolution and expansion. A large number of IgSF single base and variable splicing were screened in the stimulated transcriptional analysis, and a large number of mutation sites were specific to the two stimulation of Vibrio radivibrio, suggesting that the long oyster IgSF may produce diversity through point mutation and variable splicing, and the new individual variant formed by point mutation may be involved in "selecting adaptive immunity." In addition to the analysis of transcriptional data from different periods of oysters, some long oysters IgSF can be up-regulated at a specific developmental stage and may be involved in developmental regulation of mother source and cleavage, embryo, larval and juvenile type. Non transmembrane cgjam-a-l is a jam family homologous group found in long oysters. It contains three series of i-set type Ig domains. It can identify and combine a variety of PAMPS (LPS, PGN, man, LTA, poly (i:c) and Glu) and microbes (Vibrio, Vibrio eel, Escherichia coli, Staphylococcus aureus, Micrococcus luticus, Pichia Hierro and Pichia pastoris), and play a regulating role in the inherent immune response of long oysters. Cgjam-a-l's structural evolution information indicates that jam family and antibody molecules may have a multi-functional "ancestral gene", and the specific differentiation of functions has been achieved in evolution, thus forming jam and antibody molecules respectively. The transmembrane cgsiglec-1 gene contains two Ig domains and two intracellular receptor tyrosine inhibitors The sequence (ITIM), which plays a connection in the immuno identification and induced cascade reaction of long oysters, is widely distributed in various tissues, and the transcriptional transcript in the blood lymphocytes can be significantly up-regulated after Vibrio splendeni can be combined with a variety of PAMPS (psias, LPS and PGN), and by inhibiting downstream signaling pathways in the blood. Apoptosis, phagocytosis, and cytokine release play an regulatory role. Transmembrane cgncam contains five Ig domains, two FNIII domains and a transmembrane region, and may play a pattern recognition role in the inherent immunity of long oysters. It has a higher similarity with a plant lectin and is stimulated by microbes, MRNA in blood lymphocytes can be significantly up-regulated. The recombinant rcgncam protein has broad spectrum recognition ability for a variety of PAMPS (LPS, Glu and man) and microbes (Vibrio, Vibrio eel, Escherichia coli, Pichia Garcinia, Pichia Hierro and Pichia pastoris). Meanwhile, cgncam may also play an important role in the formation of long oyster extracellulartraps. The main functions of the three different types of IgSF show that the main function of the oyster IgSF is to recognize and adhere, the free type of IgSF may play the role of the modulators, and the transmembrane IgSF can be used as the surface receptor of the antigen molecule and regulate the downstream immune pathway. In summary, the number of IgSF in the oyster is numerous, rich and capable. IgSF may play an important role in the immune response. The variety of gene may be produced by point mutation and alterable splicing, and the newly formed individual variant may participate in the "selective adaptive immune" response to the.Igsf member cgjam-a-l, cgsiglec-1 and cgncam can identify the combination of multiple exogenous microbes and play a regulating molecule, respectively. The effect of immunomodulatory molecules and surface receptors. The results explored the heterogeneity of IgSF and higher animals in invertebrates, preliminarily revealed the role of IgSF in the selection of adaptive immunity, providing molecular biological evidence for further study of the evolution of IgSF molecules and enriching and developing the oceanic spine. The content of animal immunology.
【學(xué)位授予單位】：中國(guó)科學(xué)院研究生院(海洋研究所)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：S917.4
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本文編號(hào)：2030396