Bcl11a和Bcl11b基因抑制病毒增殖功能研究
發(fā)布時(shí)間：2018-07-20 11:46
【摘要】：Bcl11a和Bcl11b是鋅指蛋白樣轉(zhuǎn)錄因子,分別對(duì)B淋巴和T淋巴細(xì)胞的發(fā)育、分化和增殖起到重要調(diào)節(jié)作用。已有的研究也表明Bcl11a和Bcl11b作為核轉(zhuǎn)錄因子調(diào)控了一系列下游基因的表達(dá),對(duì)細(xì)胞的增殖、周期調(diào)控和免疫的調(diào)節(jié)發(fā)揮了重要作用。同時(shí),這兩個(gè)基因還被證實(shí)不僅與癌癥有密切關(guān)系,也可以抑制HIV-1病毒在細(xì)胞內(nèi)的轉(zhuǎn)錄和復(fù)制,發(fā)揮一定的抗病性。鑒于Bcl11a和Bcl11b基因在淋巴細(xì)胞中的特異表達(dá)和對(duì)淋巴細(xì)胞生長(zhǎng)發(fā)育具有重要作用,然而,在免疫反應(yīng)及抗豬圓環(huán)病毒PCV2的功能不清楚,本研究以小鼠和PK15細(xì)胞為試驗(yàn)對(duì)象,豬圓環(huán)病毒2型PCV2為病原,主要采用基因敲除、基因超表達(dá)、熒光定量、Western blot和免疫熒光等方法,對(duì)Bcl11a和Bcl11b抑制PCV2增殖的作用和機(jī)理進(jìn)行了初步研究。同時(shí),還以昆明小鼠為攻毒對(duì)象,初步探討了PCV2對(duì)淋巴細(xì)胞發(fā)育的影響。主要研究成果如下:(1)半敲除Bcl11a后(Bcl11a+/-,Bcl11a表達(dá)量降低一半)小鼠免疫能力顯著降低,血清中PCV2病毒含量顯著上升,PCV2對(duì)脾臟細(xì)胞的感染能力也增強(qiáng),對(duì)脾臟等免疫器官的損傷更加嚴(yán)重。(2)超表達(dá)Bcl11a和Bcl11b能顯著抑制PCV2在PK15細(xì)胞中的增殖,病毒m RNA、DNA和蛋白水平都顯著降低。(3)Bcl11a、Bcl11b和PCV2病毒沒(méi)有明顯的共定位。(4)Bcl11a可以通過(guò)抑制Caspase3的活性抑制由PCV2引起的細(xì)胞凋亡,而B(niǎo)cl11b通過(guò)抑制NF-κB的表達(dá)并促進(jìn)抗凋亡蛋白bcl2、抗病毒蛋白IRF3和MX1的表達(dá),抑制病毒增殖,減少細(xì)胞凋亡,說(shuō)明Bcl11a、Bcl11b抑制病毒增殖的機(jī)制可能是通過(guò)不同途徑實(shí)現(xiàn)的。(5)PCV2能導(dǎo)致發(fā)育中的淋巴細(xì)胞大量損耗,尤其是對(duì)B淋巴細(xì)胞的影響非常明顯。PCV2也可以抑制體外培養(yǎng)淋巴細(xì)胞的增殖,并對(duì)淋巴細(xì)胞發(fā)育相關(guān)重要基因有顯著的抑制作用,說(shuō)明PCV2可以抑制早期淋巴細(xì)胞的發(fā)育和增殖。
[Abstract]:Bcl11a and Bcl11b are zinc finger protein like transcription factors, which play an important role in regulating the development, differentiation and proliferation of B lymphatic and T lymphocytes. The previous studies have also shown that Bcl11a and Bcl11b have regulated the expression of a series of downstream genes as nuclear transcription factors, which play an important role in cell proliferation, cycle regulation and immune regulation. At the same time, these two genes have also been proved not only closely related to cancer, but also inhibit the transcription and replication of HIV-1 virus in cells and play a certain resistance to disease. In view of the specific expression of Bcl11a and Bcl11b genes in lymphocytes and the important role in lymphocyte growth and development, however, in the immune response and anti porcine circovirus. The function of PCV2 is not clear. In this study, mice and PK15 cells were used as the test object. The porcine circovirus type 2 PCV2 was the pathogen. The effect and mechanism of Bcl11a and Bcl11b on the inhibition of PCV2 proliferation were primarily studied by gene knockout, gene overexpression, fluorescence quantitative, Western blot and immunofluorescence. Meanwhile, the mice were attacked by Kunming mice. The effect of PCV2 on the development of lymphocyte was preliminarily discussed. The main research results were as follows: (1) after half knockout Bcl11a (Bcl11a+/-, Bcl11a expression was reduced by half), the immune ability of mice decreased significantly, the content of PCV2 virus in serum increased significantly, the infection ability of PCV2 to spleen cells increased, and the spleen and other immune organs were more damaged. (2) overexpression of Bcl11a and Bcl11b could significantly inhibit the proliferation of PCV2 in PK15 cells, and the level of M RNA, DNA and protein decreased significantly. (3) Bcl11a, Bcl11b and PCV2 viruses have no obvious co location. (4) Bcl11a can inhibit the apoptosis induced by the inhibition of Caspase3 activity. Promoting the expression of anti apoptotic protein BCL2, antiviral protein IRF3 and MX1, inhibiting the proliferation of virus and reducing cell apoptosis, the mechanism of Bcl11a, Bcl11b to inhibit the proliferation of the virus may be realized through different ways. (5) PCV2 can lead to a large amount of lymphocyte depletion in developing cells, especially the effect on B lymphocytes very obviously.PCV2 can also be suppressed. The proliferation of lymphocytes was cultured in vitro, and it has a significant inhibitory effect on the key genes related to lymphocyte development. It shows that PCV2 can inhibit the development and proliferation of early lymphocyte.
【學(xué)位授予單位】：華中農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：S852.65
本文編號(hào)：2133414
[Abstract]:Bcl11a and Bcl11b are zinc finger protein like transcription factors, which play an important role in regulating the development, differentiation and proliferation of B lymphatic and T lymphocytes. The previous studies have also shown that Bcl11a and Bcl11b have regulated the expression of a series of downstream genes as nuclear transcription factors, which play an important role in cell proliferation, cycle regulation and immune regulation. At the same time, these two genes have also been proved not only closely related to cancer, but also inhibit the transcription and replication of HIV-1 virus in cells and play a certain resistance to disease. In view of the specific expression of Bcl11a and Bcl11b genes in lymphocytes and the important role in lymphocyte growth and development, however, in the immune response and anti porcine circovirus. The function of PCV2 is not clear. In this study, mice and PK15 cells were used as the test object. The porcine circovirus type 2 PCV2 was the pathogen. The effect and mechanism of Bcl11a and Bcl11b on the inhibition of PCV2 proliferation were primarily studied by gene knockout, gene overexpression, fluorescence quantitative, Western blot and immunofluorescence. Meanwhile, the mice were attacked by Kunming mice. The effect of PCV2 on the development of lymphocyte was preliminarily discussed. The main research results were as follows: (1) after half knockout Bcl11a (Bcl11a+/-, Bcl11a expression was reduced by half), the immune ability of mice decreased significantly, the content of PCV2 virus in serum increased significantly, the infection ability of PCV2 to spleen cells increased, and the spleen and other immune organs were more damaged. (2) overexpression of Bcl11a and Bcl11b could significantly inhibit the proliferation of PCV2 in PK15 cells, and the level of M RNA, DNA and protein decreased significantly. (3) Bcl11a, Bcl11b and PCV2 viruses have no obvious co location. (4) Bcl11a can inhibit the apoptosis induced by the inhibition of Caspase3 activity. Promoting the expression of anti apoptotic protein BCL2, antiviral protein IRF3 and MX1, inhibiting the proliferation of virus and reducing cell apoptosis, the mechanism of Bcl11a, Bcl11b to inhibit the proliferation of the virus may be realized through different ways. (5) PCV2 can lead to a large amount of lymphocyte depletion in developing cells, especially the effect on B lymphocytes very obviously.PCV2 can also be suppressed. The proliferation of lymphocytes was cultured in vitro, and it has a significant inhibitory effect on the key genes related to lymphocyte development. It shows that PCV2 can inhibit the development and proliferation of early lymphocyte.
【學(xué)位授予單位】：華中農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：S852.65
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