本文關鍵詞：重組法國梧桐花粉過敏原（rPla a2）對小鼠ORMDL3基因轉錄調控機制初步研究　出處：《南京醫(yī)科大學》2017年碩士論文　論文類型：學位論文

  更多相關文章： 植物吸入性過敏原 重組法國梧桐花粉過敏原2 小鼠血清類黏蛋白1樣蛋白3 GATA結合蛋白3 轉錄調控

【摘要】：目的:分析轉錄因子GATA3對小鼠ORMDL3基因轉錄的影響機制,探索重組法國梧桐花粉過敏原rPla a2對小鼠ORMDL3基因轉錄表達的影響及相關機制,為植物吸入性過敏原所致過敏性哮喘的防治尋找新的靶點提供思路。方法:(1)染色質免疫沉淀、RNA干擾、轉錄因子過表達、雙熒光素酶活性分析等實驗,鑒定GATA3與小鼠ORMD3啟動子結合和啟動子活性的調控作用;(2)通過實時定量PCR和Western Blot實驗,觀察GATA3對小鼠ORMDL3表達水平的影響;(3)利用法國梧桐花粉重組過敏原rPla a2探索對小鼠ORMDL3啟動子活性、mRNA及蛋白表達的影響,用ChIP-qPCR觀察rPla a2刺激后GATA3與小鼠ORMDL3啟動子區(qū)結合的影響。結果:(1)ChIP實驗表明GATA3與小鼠ORMDL3核心啟動子-74bp至+141 bp結合,但不能與小鼠ORMD3基因轉錄起始位點上游-921至-835 bp序列結合。(2)RNA干擾、轉錄因子過表達、實時熒光定量PCR、Western Blot實驗證明GATA3能正向調控ORMDL3的表達。(3)rPla a2上調小鼠ORMDL3啟動子活性,突變GATA3和STAT6結合序列后該上調作用受抑制,rPla a2促進ORMDL3和GATA3表達,干擾GATA3表達可抑制rPla a2對ORMDL3的上調作用。ChIP-qPCR實驗發(fā)現(xiàn)rPlaa2增強GATA3與ORMDL3啟動子區(qū)的結合。結論:(1)GATA3通過直接結合小鼠ORMDL3核心啟動子區(qū)域正向調控ORMDL3基因表達,為以GATA3為靶點的哮喘治療提供理論依據(jù)。(2)rPla a2通過上調內源性GATA3表達和增強GATA3與核心啟動子區(qū)域的結合上調ORMDL3表達,為植物過敏原誘發(fā)的過敏性哮喘治療提供新的思路。
[Abstract]:Objective: to analyze the influence mechanism of transcription factor GATA3 on mouse ORMDL3 gene transcription, to explore the effect of recombinant Platanus pollen allergen rPla A2 expression of mouse ORMDL3 gene transcription and related mechanisms to control the plant inhaled allergen induced allergic asthma to find new targets to provide ideas. Methods: (1) chromatinimmune precipitation, RNA interference and overexpression of the transcription factor, dual luciferase activity analysis and experimental identification, GATA3 and mouse ORMD3 promoter binding and promoter activity regulation; (2) by real-time quantitative PCR and Western Blot experiment, the level of the effect of GATA3 on the expression of ORMDL3 in mice; (3) using recombinant Platanus pollen allergen rPla A2 to explore the promoter activity of ORMDL3 in mice, the expressions of mRNA and protein, GATA3 and mouse ORMDL3 promoter region with rPla after A2 stimulation was observed by ChIP-qPCR. Results: (1) ChIP. The results show that GATA3 and mouse ORMDL3 core promoter -74bp to +141 with BP, but not with the mouse ORMD3 gene upstream of the transcription initiation site of -921 to -835. (2) BP sequence combined with RNA interference and overexpression of the transcription factor Western, real-time PCR, Blot experiments show that GATA3 can positively regulate the expression of ORMDL3 (3) rPla. Upregulation of A2 mouse ORMDL3 promoter activity, GATA3 mutations and STAT6 binding sequence after the upregulation of rPla was inhibited, A2 promotes the expression of ORMDL3 and GATA3, GATA3 interference can inhibit the expression of A2 on rPla ORMDL3.ChIP-qPCR induced test showed that rPlaa2 enhanced GATA3 and ORMDL3 binds promoter region. Conclusion: (1) GATA3 by directly binding mouse ORMDL3 core promoter region of the positive regulation of ORMDL3 gene expression, and provide a theoretical basis for the treatment of asthma by targeting GATA3. (2) rPla A2 by upregulating the expression of endogenous GATA3 and enhanced GATA3 and core promoter The combination of subregions up-regulated the expression of ORMDL3 provides a new way of thinking for the treatment of allergic asthma induced by plant allergen.

【學位授予單位】：南京醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R593.1

【參考文獻】

相關期刊論文 前10條

1 岳鳳敏，，常乃鳳;法國梧桐花粉所致的呼吸系統(tǒng)過敏分析[J];中華結核和呼吸雜志;1995年03期

2 楊波,褚以德;西寧地區(qū)花粉癥患者致敏花粉和季節(jié)分析[J];青海醫(yī)藥雜志;2001年02期

3 謝水祥,李良東,劉建新;南昌市中心氣傳花粉調查[J];贛南醫(yī)學院學報;2003年01期

4 齊名;魏花;朱琴;王艾麗;熊華;;二球懸鈴木花粉變應原特應性體質與HLA-DRB1等位基因相關性分析[J];臨床檢驗雜志;2006年06期

5 邢銳;齊名;姚凱;黃宇烽;季曉輝;;二球懸鈴木花粉主要變應原蛋白編碼基因克隆的制備[J];南京醫(yī)科大學學報(自然科學版);2006年12期

6 姚麗娜;張宏譽;;北京市氣傳柏樹花粉濃度監(jiān)測分析[J];中國公共衛(wèi)生;2009年06期

7 董尚雄;劉淮;肖正軍;廖明;付穎;;湖北省荊門地區(qū)氣傳花粉與過敏性哮喘的相關性[J];華中醫(yī)學雜志;2009年05期

8 鄭銘;王妍;周兵;杜芝燕;徐元基;于曉Y

本文編號：1379025