發(fā)布時(shí)間：2018-07-04 12:54

  本文選題：OsTCL1 + OsGL1��； 參考：《東北師范大學(xué)》2017年博士論文

【摘要】：細(xì)胞命運(yùn)決定的調(diào)控一直是發(fā)育生物學(xué)的核心問(wèn)題之一。擬南芥表皮毛和根毛的發(fā)生被當(dāng)作植物細(xì)胞命運(yùn)決定的重要研究模型。表皮毛是植物地上部分表皮組織特有的延伸出的毛狀附屬物,它能增加表皮厚度、減少水分和熱量的散失、減輕昆蟲和病原體侵害、減輕機(jī)械損傷等。擬南芥的表皮毛是一種特化的單細(xì)胞,廣泛分布在蓮座葉、莖生葉、莖和花瓣上。水稻的表皮毛分為三種:長(zhǎng)毛(macro hair)、微毛(micro hair)和腺毛(glandular hair)。無(wú)表皮毛是一種非常好的農(nóng)藝性狀,利于增加包裝效率并且減少收割、晾曬、包裝過(guò)程中粉塵給農(nóng)民帶來(lái)的過(guò)敏癥狀,因此無(wú)毛性狀被廣泛應(yīng)用到水稻種植當(dāng)中。根毛是植物根表皮細(xì)胞上特化的管狀突起。它能夠增加根與土壤的接觸面積,利于植物吸收土壤中的水分和營(yíng)養(yǎng),并對(duì)植物與土壤中微生物的相互作用起著重要作用。在擬南芥中,R2R3 MYB轉(zhuǎn)錄因子GLABRA1(GL1)、bHLH轉(zhuǎn)錄因子GLABRA3/ENHANCER OF GLABRA3(GL3/EGL3)和WDR蛋白TRANSPARENTTESTAGLABRA1(TTG1)構(gòu)成MBW轉(zhuǎn)錄激活復(fù)合體,激活GLABROUS2(GL2)的表達(dá),進(jìn)而誘導(dǎo)了表皮毛的發(fā)生。MBW轉(zhuǎn)錄激活復(fù)合體同時(shí)可以誘導(dǎo)包括TRICHOMELESS1(TCL1)、TCL2、TRYPTICHON(TRY)、CAPRICE(CPC)、ENHANCEROFTRYANDCPC1(ETC1)、ETC2和ETC3在內(nèi)的R3 MYB基因的表達(dá)。這些轉(zhuǎn)錄因子可以進(jìn)入鄰近的細(xì)胞當(dāng)中與GL1競(jìng)爭(zhēng)結(jié)合GL3,抑制轉(zhuǎn)錄激活復(fù)合體的功能,從而抑制了表皮毛的發(fā)生。調(diào)控?cái)M南芥根毛發(fā)生的機(jī)制與調(diào)控表皮毛發(fā)生的機(jī)制是類似的。只是在根細(xì)胞中,GL1被另一個(gè)R2R3 MYB轉(zhuǎn)錄因子WEREWOLF(WER)所替代。WER、GL3/EGL3和TTG1共同構(gòu)成了MBW轉(zhuǎn)錄激活復(fù)合體,啟動(dòng)下游GL2的表達(dá),進(jìn)而抑制根表皮細(xì)胞分化成根毛細(xì)胞。當(dāng)WER被R3 MYBs轉(zhuǎn)錄因子所代替,這種復(fù)合體不會(huì)激活GL2的表達(dá),使根表皮細(xì)胞分化成根毛細(xì)胞。然而,水稻表皮毛和根毛發(fā)生是否受到類似轉(zhuǎn)錄復(fù)合體的調(diào)控尚不清楚。本研究中,我們用擬南芥GL1、GL3、TTG1和TCL1的氨基酸全序列BLAST水稻的蛋白數(shù)據(jù)庫(kù),分別尋找到它們?cè)谒局械耐吹鞍譕sGL1(OsGL1A、OsGL1B、OsGL1C、OsGL1D、OsGL1E)、OsGL3(OsGL3A、OsGL3B、OsGL3C)、OsTTG1(OsTTG1A、OsTTG1B)、OsTCL1以及OsTCL2。我們的研究表明,OsGL1E、OsGL3B、OsTTG1A以及OsTCL1都能夠定位在細(xì)胞核中。此外,OsTTG1A和OsTCL1還能夠分別定位在細(xì)胞質(zhì)和細(xì)胞膜上。通過(guò)原生質(zhì)體實(shí)驗(yàn)證實(shí)OsGL3B是轉(zhuǎn)錄激活子,而OsTTG1A和OsTCL1有微弱的轉(zhuǎn)錄激活活性,OsGL1E沒(méi)有轉(zhuǎn)錄激活活性。在擬南芥原生質(zhì)體中,GL1、TTG1和TCL1可以與OsGL3B互作,OsGL1E、OsTTG1A和OsTCL1都能與GL3互作,也能與OsGL3B互作。以上結(jié)果暗示,水稻中的相應(yīng)轉(zhuǎn)錄因子也可以形成MBW復(fù)合體。我們以O(shè)sTCL1為例,研究其對(duì)擬南芥表皮毛和根毛發(fā)生的調(diào)控。用雙35S啟動(dòng)子調(diào)控OsTCL1的表達(dá)能夠抑制表皮毛的發(fā)生,同時(shí)還促進(jìn)根毛的發(fā)生。此外,利用tcl1、try和cpc突變體為背景的表型恢復(fù)實(shí)驗(yàn)的結(jié)果說(shuō)明,OsTCL1與TCL1的功能是類似的,但不完全一致。并且qRT-PCR結(jié)果顯示,在過(guò)量表達(dá)OsTCL1的轉(zhuǎn)基因擬南芥中,GL1的表達(dá)量大幅下降。以上結(jié)果表明,OsTCL1調(diào)控?cái)M南芥的表皮毛和根毛發(fā)生,很可能與擬南芥R3 MYB轉(zhuǎn)錄因子調(diào)控表皮毛和根毛發(fā)生的機(jī)制類似。另外,我們?cè)趖tg1突變體背景下過(guò)量表達(dá)OsTTG1A使突變體無(wú)表皮毛的表型得到恢復(fù),這證明了OsTTG1A與TTG1對(duì)擬南芥表皮毛發(fā)生有類似的調(diào)控作用。在水稻中,我們同樣通過(guò)過(guò)量表達(dá)OsTCL1,研究其對(duì)于水稻表皮毛和根毛發(fā)生的調(diào)控作用。但是我們并沒(méi)有發(fā)現(xiàn)明顯的表皮毛和根毛的表型變化。qRT-PCR結(jié)果顯示,與擬南芥中GL2的表達(dá)被明顯抑制不同,過(guò)量表達(dá)OsTCL1的轉(zhuǎn)基因水稻中OsGL2的表達(dá)量明顯升高。并且,OsGL3C的表達(dá)量也有明顯升高,這與過(guò)量表達(dá)TCL1或OsTCL1的擬南芥中,GL3的表達(dá)量沒(méi)有明顯變化也是不同的。以上結(jié)果暗示水稻很可能通過(guò)與擬南芥不同的機(jī)制調(diào)控表皮毛和根毛的發(fā)生。
[Abstract]:The regulation of cell fate determination is one of the core problems in developmental biology. The occurrence of the surface fur and root hairs of Arabidopsis is regarded as an important research model for the fate of plant cells. The surface fur is a special hair like appendage of the epidermal tissue of the plant. It can increase the thickness of the epidermis and reduce the loss of water and heat. The epidermis of Arabidopsis is a special single cell, widely distributed in the lotus leaves, stems, stems and petals. The epidermis of rice is divided into three kinds: long hair (macro hair), micro hair and glandular hair. No surface fur is a very good agronomic character. Increasing the efficiency of packaging and reducing the allergic symptoms caused by dust in the process of harvesting, drying and packing, so the hairless characters are widely used in rice planting. The root hair is a special tubular protuberance on the root epidermal cells of plants. It can increase the contact area between the root and the soil, and benefit the plant to absorb water and nutrition in the soil. In Arabidopsis, the R2R3 MYB transcription factor GLABRA1 (GL1), the bHLH transcription factor GLABRA3/ENHANCER OF GLABRA3 (GL3/EGL3) and WDR protein TRANSPARENTTESTAGLABRA1 (TTG1) constitute the transcriptional activation complex, which activates the expression of the transcription factor, thus inducing the hair of the epidermis. The transcriptional activation complex of.MBW can also induce the expression of TRICHOMELESS1 (TCL1), TCL2, TRYPTICHON (TRY), CAPRICE (CPC), ENHANCEROFTRYANDCPC1 (ETC1), ETC2 and ETC3, which can enter the adjacent cells to compete with the cells to inhibit the function of the transcriptional activation complex and thus inhibit the function of the transcriptional activation complex. The mechanism of regulation of the root hairs of Arabidopsis is similar to that in the regulatory table. Only in the root cells, GL1 is replaced by another R2R3 MYB transcription factor WEREWOLF (WER), and GL3/EGL3 and TTG1 co constitute the MBW transcriptional activation complex, initiating the expression of the downstream GL2 and inhibiting the root epidermal cells. It is transformed into root hair cells. When WER is replaced by R3 MYBs transcription factors, this complex does not activate GL2 expression and causes root epidermal cells to differentiate into root hair cells. However, it is not clear whether the hair and root hair of rice is regulated by similar transcriptional complexes. In this study, we used the amino acid full order of Arabidopsis GL1, GL3, TTG1, and TCL1. The protein database of BLAST rice is listed to find the homologous protein OsGL1 (OsGL1A, OsGL1B, OsGL1C, OsGL1D, OsGL1E) in rice, OsGL3 (OsGL3A, OsGL3B, OsGL3C), and our research shows that all of them can be located in the nucleus. TCL1 can also be located on the cytoplasm and cell membrane respectively. Through protoplast experiments, OsGL3B is a transcriptional activator, and OsTTG1A and OsTCL1 have weak transcriptional activation activity, and OsGL1E has no transcriptional activation activity. In Arabidopsis protoplasts, GL1, TTG1 and TCL1 can interact with OsGL3B, OsGL1E, OsTTG1A, and OsTCL1. The above results suggest that the corresponding transcription factors in rice can also form a MBW complex. We take OsTCL1 as an example to study the regulation of the hair and root hair of Arabidopsis. The expression of OsTCL1 with double 35S promoter can inhibit the occurrence of hair and also promote the occurrence of root hair. In addition, TCL1, try and C are used. The results of the phenotypic recovery experiment with the PC mutant showed that the function of OsTCL1 and TCL1 was similar, but not exactly consistent. And qRT-PCR results showed that the expression of GL1 in the transgenic Arabidopsis thaliana with excessive expression of OsTCL1 decreased significantly. The results showed that OsTCL1 regulates the hair and root hair of the Arabidopsis thaliana, which is likely to be with the pseudo south. The mechanism of R3 MYB transcription factor of mustard regulates the mechanism of epidermal hair and root hair. In addition, our overexpression of OsTTG1A under the TTG1 mutant background restores the phenotype of the mutant without the epidermis, which proves that OsTTG1A and TTG1 have similar regulatory effects on the formation of the Arabidopsis thaliana fur. In rice, we also overexpress OsTCL1, We studied the regulation of the hair and root hair of rice. But we did not find the obvious phenotypic changes of the surface fur and root hair..qRT-PCR results showed that the expression of GL2 in the Arabidopsis was significantly inhibited, and the amount of OsGL2 in the transgenic rice with excessive expression of OsTCL1 was significantly increased. And the expression of OsGL3C was also expressed. There was a significant increase in the expression of GL3 in Arabidopsis thaliana, which overexpressed TCL1 or OsTCL1. The results suggested that rice is likely to regulate the formation of fur and root hairs by different mechanisms of Arabidopsis.
【學(xué)位授予單位】：東北師范大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：Q943.2

【參考文獻(xiàn)】

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1 王諾菡;于霽雯;吳Z，

本文編號(hào)：2096146