【摘要】：咪唑啉1型受體(imidazoline-1 receptor,I1R)是近年發(fā)現(xiàn)的一種新型受體,能夠介導(dǎo)許多生理作用。2000年P(guān)iletz教授成功地克隆出了咪唑啉受體抗體選擇性蛋白(imidazoline receptor antisera selected protein,IRAS),為進(jìn)一步研究I1R的功能和信號(hào)轉(zhuǎn)導(dǎo)過程奠定了基礎(chǔ)。 本研究應(yīng)用本實(shí)驗(yàn)室已經(jīng)建立的穩(wěn)定轉(zhuǎn)染IRAS的細(xì)胞(CHO-IRAS),對(duì)IRAS偶聯(lián)的信號(hào)轉(zhuǎn)導(dǎo)途徑進(jìn)行了初步研究。經(jīng)典I1R激動(dòng)劑莫索尼定、利美尼定及其內(nèi)源性配體胍丁胺激活I(lǐng)RAS后并不能顯著提高CHO-IRAS細(xì)胞的[~(35)S]-GTPγS結(jié)合量,表明IRAS不與G蛋白偶聯(lián)。利美尼定、莫索尼定及胍丁胺激活I(lǐng)RAS后時(shí)間及濃度依賴性地引起磷脂酰膽堿特異性磷脂酶C(phosphatidylcholine selectivephospholipase C,PC-PLC)活性升高,且這一作用能被PC-PLC特異性拮抗劑D609及I1R選擇性拮抗劑依法克生所抑制。上述藥物在引起PC-PLC活性升高的同時(shí),也能顯著促進(jìn)甘油二酯(diacylglyceride,DAG)含量增加。利美尼定、莫索尼定及胍丁胺都能引起細(xì)胞外信號(hào)調(diào)節(jié)激酶(extracellular signal-regulated kinase,ERK)磷酸化水平發(fā)生改變。利美尼定和莫索尼定可以濃度依賴性地升高ERK磷酸化水平;但胍丁胺對(duì)ERK磷酸化有雙重作用,低濃度時(shí)(1 nM-10 nM)降低ERK磷酸化水平,高濃度時(shí)(10 μM-100μM)促進(jìn)ERK磷酸化。利美尼定、莫索尼定及胍丁胺對(duì)ERK磷酸化的作用能被PC-PLC特異性拮抗劑D609和I1R拮抗劑依法克生所抑制。 本研究首次直接證實(shí)IRAS不是G蛋白偶聯(lián)受體。IRAS的信號(hào)轉(zhuǎn)導(dǎo)途徑可能是通過活化PC-PLC繼而產(chǎn)生DAG,其后引起了絲裂原激活蛋白激酶(mitogen-activated protein kinase,MAPK)酶促級(jí)聯(lián)反應(yīng)過程。本研究為進(jìn)一步闡明IRAS作為I1R候選蛋白而產(chǎn)生相應(yīng)功能的分子機(jī)制提供了重要線索。
[Abstract]:Imidazoline type 1 receptor (I1R) is a new type of receptor found in recent years. In 2000, Professor Piletz successfully cloned Imidazoline receptor antibody selective protein (imidazoline receptor antisera selected protein (IRAs), which laid a foundation for the further study of I1R function and signal transduction process. In this study, we studied the signal transduction pathway of IRAS coupling using stable transfected IRAS cells (CHO-IRAS) established in our laboratory. Moxonidine, riminidine and its endogenous ligand agmatine did not significantly increase the binding capacity of [35S] -GTP 緯 S in CHO-IRAS cells, suggesting that IRAS was not coupled with G protein. The activity of phosphatidylcholine-specific phospholipase (C (phosphatidylcholine selectivephospholipase) was increased in a dose-and time-dependent manner after the activation of IRAS by laminidine, moxonidine and agmatine. This effect was inhibited by PC-PLC specific antagonist D609 and I1R selective antagonist Fokeson. The above drugs can increase the activity of PC-PLC and increase the content of diacylglyceride. Laminidine, moxonidine and agmatine can all cause changes in extracellular signal regulated kinase (extracellular signal-regulated kinase) phosphorylation. The level of ERK phosphorylation was increased in a concentration-dependent manner by nimenidine and moxonidine, but agmatine had a dual effect on ERK phosphorylation. At low concentration (1 nM-10 nm), ERK phosphorylation level was decreased, and ERK phosphorylation was promoted at high concentration (10 渭 M-100 渭 M). The phosphorylation of ERK by liminidine, moxonidine and agmatine was inhibited by PC-PLC specific antagonists D609 and I1R antagonists Fokeson. It is the first direct evidence that IRAS is not a G protein-coupled receptor. IRAs signal transduction pathway may be by activating PC-PLC to produce PC-PLC, and then induce the process of mitogen-activated protein kinase-activated protein kinase (mitogen-activated protein kinase) enzymatic cascade reaction. This study provides important clues for further elucidating the molecular mechanism of IRAS as a candidate protein for I1R.
【學(xué)位授予單位】：中國人民解放軍軍事醫(yī)學(xué)科學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2005
【分類號(hào)】：R33

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1 李斐;咪唑啉1型受體候選蛋白信號(hào)轉(zhuǎn)導(dǎo)的研究[D];中國人民解放軍軍事醫(yī)學(xué)科學(xué)院;2005年

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