【摘要】：干細胞是一類具有極強自我更新及多向分化潛能的細胞,在體外特定條件下可被誘導(dǎo)分化為成熟的神經(jīng)元、胰島細胞,從而為細胞移植治療糖尿病、帕金森氏綜合癥、中風(fēng)等多種疾病提供優(yōu)良的種子細胞,具有重要的臨床應(yīng)用價值。然而在干細胞向神經(jīng)元、胰島細胞分化的過程中存在著一系列問題,包括分化效率低、分化的細胞功能不成熟等,歸根結(jié)底是因為分化的機制目前尚不清楚。文獻調(diào)研及我們實驗室前期的工作都表明神經(jīng)元限制性沉默因子(neuron-restrictive silencer factor,NRSF;又稱RE-1 silencing transcription factor,REST)在干細胞向神經(jīng)元和胰島細胞分化的過程中發(fā)揮重要的作用,此外,本實驗室的研究還表明NRSF調(diào)控神經(jīng)及胰島相關(guān)的靶基因如胰島素、可卡因-苯丙胺調(diào)節(jié)轉(zhuǎn)錄肽(Cocaine and amphetamine-regulated transcript peptide,CART)等的表達。基于這樣的工作基礎(chǔ),進一步研究NRSF對其靶基因的轉(zhuǎn)錄調(diào)控機制,有助于我們更好的了解NRSF在干細胞向神經(jīng)元或胰島細胞分化及成熟細胞功能維持方面的作用,從而為今后指導(dǎo)干細胞向成熟的神經(jīng)元或胰島細胞分化提供新的思路。 NRSF是一種含有9個Cys2/His2型鋅指結(jié)構(gòu)的蛋白,具有一個DNA結(jié)合結(jié)構(gòu)域、一個賴氨酸富含區(qū)、一個脯氨酸富含區(qū)和兩個轉(zhuǎn)錄抑制結(jié)構(gòu)域——N端抑制結(jié)構(gòu)域和C端抑制結(jié)構(gòu)域,廣泛表達于胚胎干細胞(embryonic stem cells,ES細胞)、神經(jīng)干細胞(neural stem cells,NSCs)和非神經(jīng)細胞中,在大部分分化的神經(jīng)元中沒有表達,通過與特異的作用元件NRSE(neuron-restrictive silencer element,NRSE;又稱Repressor Element,RE1)相結(jié)合,招募不同的抑制復(fù)合物,發(fā)揮對靶基因如II型鈉離子通道、SCG10、連接蛋白36(Connexin36)等轉(zhuǎn)錄水平的調(diào)控,從而在維持ES細胞全能性與自我更新、胚胎早期發(fā)育、干細胞向神經(jīng)元、胰島細胞的分化等多個過程中扮演重要角色。 CART是一種新型的神經(jīng)肽類物質(zhì),分布于中樞和周圍神經(jīng)系統(tǒng)及腎上腺、胰島等內(nèi)分泌組織,眾多研究表明其參與到進食與肥胖、應(yīng)激、能量代謝、神經(jīng)保護等多個生理過程。近年來,有研究表明CART參與調(diào)控胰島細胞的糖反應(yīng)性。目前對CART的研究主要集中在CART肽的定位、加工及其功能發(fā)揮,關(guān)于CART基因轉(zhuǎn)錄調(diào)控方面包括轉(zhuǎn)錄因子、調(diào)控元件及CART肽發(fā)揮神經(jīng)保護作用的機制的研究尚為數(shù)不多,而這些科學(xué)問題的探討,可以使我們弄清如何使CART發(fā)揮生物學(xué)功能的分子機制,為基礎(chǔ)研究和糖尿病、神經(jīng)系統(tǒng)疾病等的臨床治療提供有力的理論指導(dǎo)。 我們之前的研究工作表明CART基因是NRSF作用的靶基因之一,其核心啟動子區(qū)存在的NRSE樣基序與NRSF的序列特異性結(jié)合可以發(fā)揮對CART基因的轉(zhuǎn)錄抑制調(diào)控。本文通過進一步的生物信息學(xué)分析發(fā)現(xiàn),CART基因第一個內(nèi)含子區(qū)也存在與公認的NRSE共有序列極為相似的NRSE樣基序,并且在物種間具有較好的保守性,那么這段序列是否也參與了CART基因的轉(zhuǎn)錄調(diào)控?它又是如何與CART基因啟動子區(qū)的NRSE相互協(xié)調(diào)發(fā)揮對CART的轉(zhuǎn)錄調(diào)節(jié)作用?目前關(guān)于CART基因轉(zhuǎn)錄調(diào)控機制方面的研究比較清楚的只有cAMP/PKA/CREB正性調(diào)控通路,那么,我們研究發(fā)現(xiàn)的NRSF-NRSE負性調(diào)控系統(tǒng)與cAMP/PKA/CREB正性調(diào)控系統(tǒng)之間是如何競爭性調(diào)節(jié)CART基因,共同實現(xiàn)對CART基因轉(zhuǎn)錄水平的調(diào)控呢?以上關(guān)鍵問題的解決也就成為了本研究的目的。 本研究主要包括三部分內(nèi)容: 一、內(nèi)含子序列對CART基因轉(zhuǎn)錄調(diào)控作用的研究 通過應(yīng)用rVista軟件,我們對從美國國家生物醫(yī)學(xué)信息中心NCBI數(shù)據(jù)庫檢索得到的人、大鼠和小鼠CART基因的內(nèi)含子序列進行分析,預(yù)測到了一系列可能調(diào)控CART基因表達的元件,包括ZF5、E47、NRSE、AREB6等,結(jié)合之前的工作基礎(chǔ),我們確定以人、大鼠、小鼠CART基因第一個內(nèi)含子區(qū)保守性較好的NRSE樣基序為研究對象,進一步開展CART基因轉(zhuǎn)錄調(diào)控機制方面的研究。為了探討CART基因第一個內(nèi)含子及其中的NRSE元件對CART基因的轉(zhuǎn)錄調(diào)控作用,我們基于CART啟動子-熒光素酶報告載體pGL3-Basic-CART32(P-Luc),構(gòu)建了熒光素酶基因3’端連入內(nèi)含子序列(去或不去除NRSE基序)的熒光素酶報告載體pGL3-Basic-CART32-Intron(P-Luc-I)、pGL3-Basic-CART32-NRSEnon Intron(P-Luc-tI),將上述載體與含海腎熒光素酶的內(nèi)參質(zhì)粒pRL-CMV共轉(zhuǎn)染入NTera2/CloneD1細胞,通過檢測熒光素酶活性探討CART基因第一個內(nèi)含子區(qū)域及NRSE樣基序?qū)ζ滢D(zhuǎn)錄活性的影響,結(jié)果顯示P-Luc-I轉(zhuǎn)錄活性相對于對照組P-Luc下調(diào)57.9%(P0.05),表明CART基因第一個內(nèi)含子序列參與CART基因的轉(zhuǎn)錄負調(diào)控;P-Luc-tI相對于P-Luc-I熒光素酶活性上調(diào)53.8%(P0.05),提示CART基因內(nèi)含子通過其區(qū)域內(nèi)的NRSE發(fā)揮對CART基因的轉(zhuǎn)錄負調(diào)控作用。之后我們通過應(yīng)用anti-NRSF抗體進行染色質(zhì)免疫共沉淀( chromatin immunoprecipitation,ChIP)實驗,分析CART基因的內(nèi)含子區(qū)域與NRSF的結(jié)合狀況,結(jié)果顯示二者之間存在直接的結(jié)合,證實NRSF與內(nèi)含子區(qū)NRSE的結(jié)合真實的存在于天然染色質(zhì)區(qū)。 二、NRSF對CART基因轉(zhuǎn)錄的調(diào)控作用及其機制研究 結(jié)合之前我們實驗室對CART基因啟動子區(qū)NRSE元件研究的工作基礎(chǔ)及本文第一部分對CART基因第一個內(nèi)含子區(qū)NRSE樣基序的初步研究,為進一步探討NRSF是如何通過CART基因啟動子及內(nèi)含子序列發(fā)揮對該基因的轉(zhuǎn)錄調(diào)控作用,我們開展了一系列的工作。通過電泳遷移率變動分析實驗(Eelectrophoretic Mobility Shift Assay,EMSA)證實CART基因啟動子區(qū)及第一個內(nèi)含子區(qū)NRSE基序在體外均可以與NRSF發(fā)生序列特異性結(jié)合;染色質(zhì)免疫共沉淀實驗表明在NRSF表達量不同的細胞系,NRSF與NRSE結(jié)合后,通過招募組蛋白以不同的強度結(jié)合,形成不同的抑制復(fù)合物,進而發(fā)揮對CART基因的轉(zhuǎn)錄調(diào)控作用;為弄清NRSF是如何通過啟動子及內(nèi)含子序列發(fā)揮對CART基因的轉(zhuǎn)錄調(diào)控作用,我們使用了CART啟動子-熒光素酶報告載體P-Luc、P-Luc-tI、P-Luc-I,同時構(gòu)建了去除NRSE基序的CART啟動子-熒光素酶報告載體pGL3-Basic-NRSEnon promoter (tP-Luc),及在其熒光素酶基因3’端連入內(nèi)含子序列(去或不去除NRSE基序)的熒光素酶報告載體pGL3-Basic-NRSEnon promoter-Intron ( tP-Luc-I )、pGL3-Basic-NRSEnon promoter-NRSEnon Intron(tP-Luc-tI),將這些報告載體分別轉(zhuǎn)染到HeLa細胞中,通過檢測熒光素酶活性的分析結(jié)果表明CART基因內(nèi)含子與啟動子協(xié)同的參與CART基因的轉(zhuǎn)錄負調(diào)控,啟動子區(qū)NRSF-NRSE對CART基因發(fā)揮更強的轉(zhuǎn)錄負調(diào)控作用。共轉(zhuǎn)染熒光素酶報告載體、NRSF表達質(zhì)粒pcDNA3.1-NRSF到NRSF低表達的SK-N-SH細胞中,也得到相似的結(jié)論;為了驗證CART基因啟動子與第一個內(nèi)含子發(fā)揮的轉(zhuǎn)錄負調(diào)控作用是否依賴于其各自區(qū)域內(nèi)的NRSE元件,我們在本室原有的含有雙拷貝CART啟動子NRSE基序(正常或突變序列)的報告載體的基礎(chǔ)上,在其熒光素酶基因的3’端連入雙拷貝CART內(nèi)含子NRSE基序(正�；蛲蛔冃蛄�),將這些熒光素酶報告載體分別與含海腎熒光素酶的內(nèi)參質(zhì)粒pRL-CMV共轉(zhuǎn)染入HeLa細胞和SK-N-SH細胞,檢測熒光素酶活性的分析結(jié)果證實CART基因啟動子及第一個內(nèi)含子區(qū)對CART基因轉(zhuǎn)錄發(fā)揮的負調(diào)控作用依賴于其各自區(qū)域內(nèi)NRSE元件的協(xié)同作用。 三、CART基因表達的正性與負性調(diào)控系統(tǒng)及其作用機制的研究 為了進一步考察NRSF-NRSE負性調(diào)控系統(tǒng)與cAMP/PKA/CREB正性調(diào)控系統(tǒng)之間是如何競爭性調(diào)節(jié)CART基因的表達,首先我們共轉(zhuǎn)染P-Luc-I、對照質(zhì)粒pcDNA3.1或呈濃度梯度的NRSF表達質(zhì)粒pcDNA3.1-NRSF(100ng、200ng、400ng/2×105個細胞),檢測熒光素酶活性的分析結(jié)果表明轉(zhuǎn)染pcDNA3.1-NRSF 400ng/2×105個細胞可以抑制Forskolin/cAMP/PKA/CREB對CART的正性調(diào)控,提示高水平的NRSF表達引起的NRSF-NRSE負性調(diào)控作用可以抑制cAMP/PKA/CREB正性調(diào)控系統(tǒng)功能的發(fā)揮。 在此基礎(chǔ)上,我們建立了神經(jīng)細胞的氧糖剝奪(oxygen-glucose deprivation,OGD)模型,結(jié)合CART肽可以減輕缺血/缺氧導(dǎo)致的細胞死亡的功能開展了相關(guān)實驗。近年來有研究表明局部腦缺血的大鼠或OGD處理的皮質(zhì)神經(jīng)元NRSF的表達出現(xiàn)上調(diào),利用此模型我們可以進一步研究NRSF表達的變化是否會引起CART基因表達的變化,并影響cAMP/PKA/CREB正性調(diào)控通路的作用。 我們將接種在12孔板內(nèi)的SK-N-SH細胞放置在37°C,0.3% O2,95% N2的低氧手套箱中處理3h,體外模擬缺血/缺氧的環(huán)境。通過RT-PCR和Western Blot檢測處理的SK-N-SH細胞復(fù)氧24 h、48 h后NRSF、CART表達的變化,并應(yīng)用流式細胞儀檢測處理前后細胞凋亡率的變化,結(jié)果顯示經(jīng)OGD處理的SK-N-SH細胞復(fù)氧后NRSF表達量上調(diào),CART表達量下調(diào),以24 h最為顯著,細胞凋亡率則隨著復(fù)氧時間的延長而升高;應(yīng)用此模型進行cAMP/PKA/CREB和NRSF-NRSE調(diào)控系統(tǒng)相互作用關(guān)系的研究中,我們在OGD處理SK-N-SH細胞3 h后,進行報告載體P-Luc-I、對照質(zhì)粒pcDNA3.1或NRSF表達質(zhì)粒pcDNA3.1-NRSF的共轉(zhuǎn)染,熒光素酶活性檢測的結(jié)果提示OGD處理可以引起NRSF表達的上調(diào),但不足以抑制Forskolin/cAMP/PKA/CREB對CART的正性調(diào)控,當(dāng)外源性轉(zhuǎn)染NRSF表達質(zhì)粒200ng、400ng/2×105個細胞時,則可以抑制Forskolin正性作用的發(fā)揮;細胞凋亡率的流式檢測結(jié)果同樣提示高水平的NRSF表達引起的NRSF-NRSE負性調(diào)控作用,可以抑制cAMP/PKA/CREB正性調(diào)控作用的發(fā)揮。 綜上所述,我們通過生物信息學(xué)分析、電泳遷移率變動分析實驗、染色質(zhì)免疫共沉淀實驗、熒光素酶報告系統(tǒng)活性檢測證實了CART基因第一個內(nèi)含子中的NRSE樣基序通過與NRSF蛋白發(fā)生序列特異性結(jié)合從而發(fā)揮對CART基因的轉(zhuǎn)錄負調(diào)控作用,CART基因啟動子、第一個內(nèi)含子依賴于其各自區(qū)域內(nèi)的NRSE元件之間的相互作用協(xié)同的發(fā)揮對CART基因的轉(zhuǎn)錄阻遏功能;高水平的NRSF表達引起的NRSF-NRSE負性調(diào)控作用可以有效的抑制cAMP/PKA/CREB正性調(diào)控系統(tǒng)作用的發(fā)揮。這一研究工作使我們對NRSF的轉(zhuǎn)錄調(diào)控作用、其對靶基因CART的轉(zhuǎn)錄調(diào)控機制研究又有了新的認識,為今后探討該調(diào)控機制在干細胞向成熟神經(jīng)元、胰島細胞的發(fā)育分化及在病理損傷狀態(tài)下,如何通過干預(yù)NRSF、CART基因的表達發(fā)揮治療作用等方面提供了新的思路。
[Abstract]:Stem cells are a kind of cells with strong self-renewal and multi-directional differentiation potential. They can be induced to differentiate into mature neurons and islet cells under specific conditions in vitro, thus providing excellent seed cells for cell transplantation in the treatment of diabetes, Parkinson's syndrome, stroke and other diseases. However, they have important clinical application value. In the process of stem cells differentiating into neurons and islet cells, there are a series of problems, including inefficient differentiation, immature function of differentiated cells, etc. In the final analysis, the mechanism of differentiation is still unclear. NCER factor, NRSF; also known as RE-1 silencing transcription factor (REST) plays an important role in the differentiation of stem cells into neurons and islet cells. In addition, our laboratory studies have shown that NRSF regulates neuronal and islet-related target genes such as insulin, cocaine and amphetamine-regu. Based on this work, further study on the transcriptional regulation mechanism of NRSF on its target genes will help us better understand the role of NRSF in the differentiation of stem cells into neurons or islet cells and the maintenance of function of mature cells, so as to guide stem cells to mature nerves in the future. Cell or islet cell differentiation provides new ideas.
NRSF is a protein with nine Cys2/His2 zinc finger structures. It has one DNA binding domain, one lysine-rich domain, one proline-rich domain and two transcription inhibitory domains, N-terminal inhibitory domain and C-terminal inhibitory domain. It is widely expressed in embryonic stem cells (ES cells), neural stem cells (neural stem cells). Stem cells, NSCs, and non-neural cells, which are not expressed in most differentiated neurons, recruit different inhibitory complexes by binding to specific acting elements NRSE (neuron-restrictive silencer element, NRSE; also known as Repressor Element, RE1) to play a role in target genes such as type II sodium channel, SCG10, connectin 36 (Connex). In36) plays an important role in the maintenance of ES cell totipotency and self-renewal, early embryonic development, stem cell differentiation into neurons and islet cells.
CART is a new type of neuropeptide, which is distributed in the central and peripheral nervous system, adrenal glands, islets and other endocrine tissues. Many studies have shown that CART is involved in many physiological processes, such as food intake and obesity, stress, energy metabolism, neuroprotection and so on. The research mainly focuses on the localization, processing and function exertion of CART peptides. There are few studies on the transcriptional regulation of CART gene including transcription factors, regulatory elements and the neuroprotective mechanism of CART peptides. This system will provide powerful theoretical guidance for basic research and clinical treatment of diabetes, nervous system diseases and so on.
Our previous work has shown that CART gene is one of the target genes for NRSF. The NRSE-like motifs in the core promoter region of CART gene can specifically bind to NRSF sequence to regulate the transcriptional inhibition of CART gene. NRSE has a common NRSE-like motif that is very similar to each other and is conserved among species. Does this sequence also participate in the transcriptional regulation of CART gene? How does it coordinate with NRSE in the promoter region of CART gene to regulate the transcriptional regulation of CART gene? Only the positive regulatory pathway of cAMP/PKA/CREB is clear. So, how does the negative regulatory system of NRSF-NRSE and the positive regulatory system of cAMP/PKA/CREB compete to regulate the CART gene and jointly regulate the transcriptional level of CART gene?
This study mainly consists of three parts:
First, the transcriptional regulation of CART gene by intron sequence.
By using rVista software, we analyzed the intron sequences of human, rat and mouse CART genes retrieved from NCBI database of the National Center for Biomedical Information of the United States, and predicted a series of elements that might regulate the expression of CART genes, including ZF5, E47, NRSE, AREB6, and so on. Combining with previous work, we identified human, large and large. In order to investigate the transcriptional regulation of CART gene by the first intron of CART gene and its NRSE elements, we studied the NRSE-like motif of the first intron of CART gene in mice. Basic-CART32 (P-Luc), a luciferase reporter vector pGL3-Basic-CART32-Intron (P-Luc-I), pGL3-Basic-CART32-NRSEnon Intron (P-Luc-tI), was constructed and co-transfected into NTera2/CloneD1 cells with the luciferase-containing plasmid pRL-CMV. The results showed that the transcriptional activity of P-Luc-I was down-regulated by 57.9% (P 0.05) compared with the control group, indicating that the first intron sequence of CART gene was involved in the negative regulation of CART gene transcription. The activity of CART gene was up-regulated by 53.8% (P 0.05), suggesting that the intron of CART gene plays a negative role in the transcriptional regulation of CART gene through NRSE in its region. It was confirmed that the binding between NRSF and intron NRSE existed in the natural chromatin region.
Two, the regulatory effect of NRSF on CART gene transcription and its mechanism.
In the first part of this paper, we carried out a preliminary study on the NRSE-like sequence of the first intron region of the CART gene. In order to further explore how NRSF regulates the transcription of the gene through the promoter and intron sequences of the CART gene, we carried out a preliminary study on the NRSE-like sequence of the first intron region of the CART gene. Series of work. Electrophoretic mobility shift assay (EMSA) confirmed that the NRSE motif of CART gene promoter region and the first intron region could specifically bind to NRSF in vitro. Chromatin immunoprecipitation assay showed that NRSF and NRSE junction occurred in different NRSF expression cell lines. In order to understand how NRSF regulates the transcription of CART gene through promoter and intron sequences, we used CART promoter-luciferase reporter vectors P-Luc, P-Luc-tI, P-Lu. At the same time, we constructed the CART promoter-luciferase reporter vector pGL3-Basic-NRSEnon promoter (tP-Luc) and the luciferase reporter vector pGL3-Basic-NRSEnon promoter-N (tP-Luc-I), and the luciferase reporter vector pGL3-Basic-NRSEnon promoter-N (pGL3-Basic-NRSEnon Promoter-N) with or without NRSE motif deletion at the 3'end of the luciferase gene. RSEnon Intron (tP-Luc-tI), these reporter vectors were transfected into HeLa cells. The results of luciferase activity assay showed that CART gene intron and promoter cooperated with the negative transcriptional regulation of CART gene, and NRSF-NRSE in promoter region played a stronger negative transcriptional regulation on CART gene. Co-transfection Luciferase Report Similar results were also obtained in SK-N-SH cells with low NRSF expression plasmid pcDNA3.1-NRSF. In order to verify whether the negative transcriptional regulation of the CART gene promoter and the first intron depended on the NRSE elements in their respective regions, we originally contained the NRSE motif of the double-copy CART promoter (normal or normal). On the basis of the mutant sequence, a double-copy CART intron NRSE motif (normal or mutant) was inserted at the 3'-terminal of the luciferase gene. These luciferase reporter vectors were co-transfected into HeLa cells and SK-N-SH cells with the pRL-CMV plasmid containing marine luciferase, respectively. The results showed that the negative regulation of CART gene transcription by the promoter and the first intron region depended on the synergistic effect of NRSE elements in their respective regions.
Three, the positive and negative regulation system of CART gene expression and its mechanism of action.
To further investigate how the NRSF-NRSE negative regulatory system and the cAMP/PKA/CREB positive regulatory system competently regulate the expression of CART gene, we first co-transfected P-Luc-I and compared the plasmid pcDNA3.1-NRSF (100ng, 200ng, 400ng/2 *105 cells) with the plasmid pcDNA3.1 or the concentration gradient NRSF expression plasmid pcDNA3.1-NRSF (100ng, 200ng, 400ng/2 *105 cells) to detect the luciferase activity. The results showed that transfected pcDNA3.1-NRSF 400ng/2 *105 cells could inhibit the positive regulation of Forskolin/cAMP/PKA/CREB on CART, suggesting that the negative regulation of NRSF-NRSE induced by high level of NRSF expression could inhibit the function of positive regulation system of cAMP/PKA/CREB.
On this basis, we established an oxygen-glucose deprivation (OGD) model of neurons and carried out related experiments in combination with the function of CART peptides to reduce cell death induced by ischemia/hypoxia. In this model, we can further investigate whether changes in NRSF expression can cause changes in CART gene expression and affect the role of cAMP/PKA/CREB positive regulatory pathway.
SK-N-SH cells inoculated in 12-well plates were placed in a hypoxic glove box at 37 C, 0.3% O2, 95% N 2 for 3 hours to simulate the ischemia/hypoxia environment in vitro. The expression of NRSF and CART in the treated SK-N-SH cells was detected by RT-PCR and Western Blot after 24 h of reoxygenation, 48 h of reoxygenation, and the apoptosis rate was detected by flow cytometry. The results showed that the expression of NRSF was up-regulated and the expression of CART was down-regulated after reoxygenation in SK-N-SH cells treated with OGD, and the apoptosis rate was up-regulated with the prolongation of reoxygenation time. In the study of the interaction between cAMP/PKA/CREB and NRSF-NRSE regulatory system, we treated SK-N-SH cells with OGD for 3 hours. Comparing with the co-transfection of pcDNA3.1 or NRSF expression plasmid pcDNA3.1-NRSF, luciferase activity assay showed that OGD treatment could induce the up-regulation of NRSF expression, but could not inhibit the positive regulation of Forskolin/cAMP/PKA/CREB on CART. When exogenous transfection of NRSF expression plasmid 200 ng, 400 ng/2*105 cells The results of flow cytometry also indicated that the negative regulation of NRSF-NRSE induced by high level of NRSF expression could inhibit the positive regulation of cAMP/PKA/CREB.
In summary, we demonstrated that the NRSE-like motifs in the first intron of CART gene play a negative role in the transcriptional regulation of CART gene by specific binding to NRSF protein through bioinformatics analysis, electrophoretic mobility change assay, chromatin immunoprecipitation assay, and luciferase reporter system activity assay. CART gene promoter, the first intron, depends on the interaction of NRSE elements in their respective regions to play a synergistic role in the transcriptional repression of CART gene.
【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)位級別】：博士
【學(xué)位授予年份】：2011
【分類號】：R346

【相似文獻】

相關(guān)期刊論文 前10條

1 李丹妮;趙越;;ERα的輔調(diào)節(jié)因子與乳腺癌關(guān)系的研究進展[J];生命科學(xué);2011年08期

2 孟希亭;林晨;梅佳;王海娟;馬飛;張金龍;張穎;錢海利;;活體成像系統(tǒng)檢測攜熒光素酶增殖缺陷型腺病毒在小鼠體內(nèi)的表達和分布[J];中國腫瘤生物治療雜志;2011年04期

3 劉竹;艾青;蘭歡;吉穎;楊正梅;何江宜;郝曉璐;宋方洲;卜友泉;;人ACER2基因啟動子的鑒定與初步分析[J];中國細胞生物學(xué)學(xué)報;2011年06期

4 高原;惠寧;劉善榮;;長鏈非編碼RNA的研究進展[J];第二軍醫(yī)大學(xué)學(xué)報;2011年07期

5 李小雷;伍志強;馬曉星;趙亞力;韓為東;;抑制LRP16基因的表達顯著下調(diào)TNF-α介導(dǎo)的NF-κB轉(zhuǎn)錄活性[J];中國生物化學(xué)與分子生物學(xué)報;2011年07期

6 ;第2個誘導(dǎo)腎癌的基因被發(fā)現(xiàn)[J];生物學(xué)通報;2011年03期

7 侯德富;關(guān)勇軍;關(guān)瑞;歐陽詠梅;余艷輝;陳主初;;人NPCEDRG基因啟動子的克隆及CCAAT/NFY結(jié)合位點初步分析[J];生物化學(xué)與生物物理進展;2011年08期

8 ;英科學(xué)家首次測出活體心臟能量水平[J];生命世界;2006年11期

9 李夏雨;沈守榮;武明花;李小玲;熊煒;盧建紅;周鳴;馬健;向娟娟;曾朝陽;向波;周艷宏;肖嵐;周厚德;范松青;李桂源;;多基因遺傳性腫瘤不同階段轉(zhuǎn)錄組學(xué)調(diào)控規(guī)律及其分子機制[J];中南大學(xué)學(xué)報(醫(yī)學(xué)版);2011年07期

10 方煥;朱傳武;陳明;;hTERT轉(zhuǎn)錄調(diào)控及其在肝細胞癌治療中的研究進展[J];臨床肝膽病雜志;2011年09期

相關(guān)會議論文 前10條

1 徐衛(wèi)紅;高瑞蘭;陳小紅;錢煦岱;吳超群;;三七皂苷對造血細胞AP-1族轉(zhuǎn)錄調(diào)控蛋白NF-E2、c-Jun和c-Fos的誘導(dǎo)作用[A];第九屆全國實驗血液學(xué)會議論文摘要匯編[C];2003年

2 李園園;王壘;陸長德;;人增殖細胞核抗原與細胞周期相關(guān)的轉(zhuǎn)錄調(diào)控研究[A];華東六省一市生物化學(xué)與分子生物學(xué)會2003年學(xué)術(shù)交流會論文摘要集[C];2003年

3 楊蘭蘭;劉先俊;;NUAK1—NFkB通路新轉(zhuǎn)錄調(diào)控或共調(diào)控因子[A];重慶市生物化學(xué)與分子生物學(xué)學(xué)術(shù)會議論文摘要匯編[C];2009年

4 鮑永利;烏垠;于春雷;孟祥穎;孫穎;李玉新;;人多藥耐藥基因MDR1啟動子克隆及轉(zhuǎn)錄調(diào)控的初步研究[A];吉林省第六屆生命科學(xué)大型學(xué)術(shù)報告會論文集[C];2008年

5 高瑞蘭;馬逢順;吳超群;許家鸞;B.H.Chong;牛泱平;苗青;王京霞;金錦梅;;人參皂苷組分治療難治性血液病及對造血相關(guān)基因的轉(zhuǎn)錄調(diào)控[A];第四次全國中西醫(yī)結(jié)合中青年學(xué)術(shù)研討會論文集[C];2002年

6 徐衛(wèi)紅;高瑞蘭;陳小紅;鄭茵紅;;三七皂苷對多種造血相關(guān)轉(zhuǎn)錄調(diào)控蛋白的誘導(dǎo)作用[A];第六屆全國中西醫(yī)結(jié)合血液病學(xué)術(shù)會議論文匯編[C];2002年

7 劉兵;毛寧;;胚胎造血發(fā)育的轉(zhuǎn)錄調(diào)控[A];第九屆全國實驗血液學(xué)會議論文摘要匯編[C];2003年

8 孔輝;王瑩;朱慶林;余擎;Gabriele Mues;Rena N.D'Souza;;牙齒發(fā)育過程中Pax9和Msx1對BMP4轉(zhuǎn)錄調(diào)控作用的研究[A];全國第八次牙體牙髓病學(xué)學(xué)術(shù)會議論文匯編[C];2011年

9 高瑞蘭;馬逢順;吳超群;許家鸞;B.H.Chong;牛泱平;苗青;王京霞;金錦梅;;人參皂苷對造血相關(guān)基因的轉(zhuǎn)錄調(diào)控及治療難治性血液病[A];第九屆全國實驗血液學(xué)會議論文摘要匯編[C];2003年

10 高春芳;王皓;徐玲玲;趙文靜;孔憲濤;;人α1(Ⅰ)膠原基因轉(zhuǎn)錄調(diào)控研究[A];中國免疫學(xué)會第四屆學(xué)術(shù)大會會議議程及論文摘要集[C];2002年

相關(guān)重要報紙文章 前10條

1 衣曉峰;哈醫(yī)大一院系列研究阿爾茨海默病關(guān)聯(lián)基因[N];中國醫(yī)藥報;2007年

2 王春;弄潮正當(dāng)時[N];科技日報;2004年

3 王振嶺;誘發(fā)心肌肥厚新基因被發(fā)現(xiàn)[N];健康報;2005年

4 記者 陳勇 曉安;“改裝”艾滋病毒成克癌“導(dǎo)彈”[N];新華每日電訊;2005年

5 記者 李紅;讓科學(xué)和藝術(shù)相結(jié)合[N];科技日報;2000年

6 本報記者 李嬋;克隆豬身上發(fā)熒光[N];北京科技報;2004年

7 王坤;我國基因剔除研究取得重大進展[N];中國醫(yī)藥報;2000年

8 王世恩 特約記者 王坤;四醫(yī)大建立基因剔除動物模型[N];解放軍報;2000年

9 記者 周穎;中澳合作開發(fā)中草藥治療血液病取得進展[N];中國中醫(yī)藥報;2001年

10 通訊員 王坤;我國基因剔除技術(shù)取得重要進展[N];科技日報;2000年

相關(guān)博士學(xué)位論文 前10條

1 張靜;神經(jīng)元限制性沉默因子對CART基因轉(zhuǎn)錄的調(diào)控作用及其機制研究[D];中國人民解放軍軍事醫(yī)學(xué)科學(xué)院;2011年

2 劉智慧;寡肽（OGP、HIV-1 tat～[47-57]）的合成、聚合及功能研究以及NRSF/REST在轉(zhuǎn)錄調(diào)控方面的研究[D];中國科學(xué)院研究生院（上海生命科學(xué)研究院）;2004年

3 李軍林;MINT蛋白同源二聚體形成及對其介導(dǎo)的轉(zhuǎn)錄抑制作用的拮抗[D];第四軍醫(yī)大學(xué);2005年

4 喬文濤;牛泡沫病毒3026反式激活因子Borf1的特征及作用機制初探[D];南開大學(xué);2004年

5 顏峻;Cyclin D3參與轉(zhuǎn)錄調(diào)控，p110C誘導(dǎo)凋亡及活性寡糖的研究[D];復(fù)旦大學(xué);2004年

6 王晶;裂殖酵母核糖體L32-2蛋白潛在轉(zhuǎn)錄調(diào)節(jié)調(diào)節(jié)作用的研究[D];南京師范大學(xué);2006年

7 盧建雄;營養(yǎng)和激素對原代培養(yǎng)大鼠脂肪細胞脂肪形成的調(diào)控及機理研究[D];西北農(nóng)林科技大學(xué);2005年

8 王繼;血管生成素與堿性成纖維生長因子之間相互作用關(guān)系及其分子機制的研究[D];東北師范大學(xué);2009年

9 朱慧芳;Y家族DNA聚合酶對化學(xué)致癌物MNNG應(yīng)答的轉(zhuǎn)錄調(diào)控研究[D];浙江大學(xué);2009年

10 秦瓏;整體調(diào)控子PhoP直接調(diào)控鼠疫耶爾森氏菌胞內(nèi)生存能力的研究[D];中國人民解放軍軍事醫(yī)學(xué)科學(xué)院;2006年

相關(guān)碩士學(xué)位論文 前10條

1 劉鳳鳴;人PDCD4啟動子的確定及其轉(zhuǎn)錄調(diào)控的初步研究[D];山東大學(xué);2010年

2 吳順泉;多個microRNAs通過直接作用于p21的3'UTR調(diào)控p21基因的表達[D];福建醫(yī)科大學(xué);2010年

3 王禮翠;HCV絲氨酸蛋白酶活性監(jiān)測體系的建立[D];中國人民解放軍軍事醫(yī)學(xué)科學(xué)院;2010年

4 翟春媛;豬TLR4基因的突變與功能分析[D];東北農(nóng)業(yè)大學(xué);2010年

5 劉成柏;用生物發(fā)光免疫技術(shù)檢測肝腫瘤標(biāo)志物AFP在肝癌早期診斷中的研究[D];吉林大學(xué);2005年

6 薛麗英;SV40增強子修飾的hTERT核心啟動子靶向轉(zhuǎn)錄熒光素酶載體的構(gòu)建和鑒定[D];天津醫(yī)科大學(xué);2005年

7 黃麗玲;水稻抗病相關(guān)基因OsDR2的分離克隆和功能鑒定[D];華中農(nóng)業(yè)大學(xué);2003年

8 唐建華;降脂藥物篩選模型的建立及其在決明子降脂作用機理研究中的應(yīng)用[D];四川大學(xué);2005年

9 劉艷杰;重組螢火蟲熒光素酶及其穩(wěn)定性研究[D];天津大學(xué);2010年

10 徐燕;熒光素酶作為報告基因的丙型肝炎病毒細胞感染系統(tǒng)的建立和應(yīng)用[D];復(fù)旦大學(xué);2011年

，

本文編號：2219090