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    Please use this identifier to cite or link to this item: http://ir.lib.ncu.edu.tw/handle/987654321/6521


    Title: 觀察肌肉生成轉錄因子如何調控 M- 和N- cadherin 表現;The regulation of M- and N-cadherin expression by myogenic factors
    Authors: 蕭勝斌;Sheng-Pin Hsiao
    Contributors: 生命科學研究所
    Keywords: 聚合酶鏈鎖反應;肌纖維母細胞;N-cadherin;M-cadherin;MyoD;promoter
    Date: 2008-07-08
    Issue Date: 2009-09-22 10:21:03 (UTC+8)
    Publisher: 國立中央大學圖書館
    Abstract: M-和N-cadherin 都是屬於鈣離子依賴型細胞與細胞間黏著分子家族的一員,M-cadherin主要表現在正在發育生成中的骨骼肌中,且在之前已被證實其在肌肉的終極分化階段扮演著非常重要的腳色尤其是在肌纖維母細胞進行融合的時期特別的明顯。藉由N-cadherin的細胞與細胞間的黏著作用同樣的也在骨骼肌的生成過程中有著顯要的地位。 MyoD、Myogenine、Myf-5和 MRF4 這四個轉錄因子同時都具有Basic-loop-helix motif 且同屬於肌肉調節因子(Muscle Regulatory Factor , MRF)。 MyoD和Myf-5在整個胚胎發育中整個肌肉生成的全部步驟中都是不可缺少的,但是Myogenin 和MRF4則不然,他們是在肌肉生成的後期也就是終極分化時才有顯著的影響。 之前的研究指出在MyoD-/- 肌原性細胞中,並無法行最後的終極分化且M-cadherin的表現量也有很明顯的下降。 我們也知道當細胞缺乏M-cadherin時 N-cadherin 會互補的表現出來而維持細胞正常的功能。 在我們的研究中我們要了解究竟這些肌肉生成調節因子是如何去調控M-和N-cadherin promoter的表現。 首先我製備了M-cadherin promoter質體並且發現MyoD會藉由結合到M-cadherin promoter上而去活化M-cadherin的表現。 接下來我們突變了M-cadherin core promoter上面的 E-box更正確的定義出MyoD及Myogenin 的結合位置。同時我們也觀察及比較MRFs對於N-cadherin的調控是否有不同於M-cadherin調控的部份,發現 MRFs 以及 MEF-2C 對於N-和M-cadherin 活性的調控並不相同,所以之後我們要找出為什麼會不同以及這種不同在生理上代表了什麼意義,未來我們會用ChIP assay (chromatin-immunoprecipitation ) 分析MRF調控M-cadherin和N-cadherin 的生理意義。 M- and N-cadherin are members of the Ca2+ dependent cell-cell adhesion molecule family. M-cadherin is predominantly expressed in developing skeletal muscles and has been implicated in terminal myogenic differentiation, particularly in myoblast fusion. N-cadherin-mediated cell-cell adhesion also plays an important role in skeletal myogenesis. MyoD, Myf-5, Myogenine and MRF-4 are basic helix-loop-helix (bHLH) transctiption factors are that form the Myogenic regulatory factors (MRFs) family. MyoD and Myf-5 are required for proper myogenesis during embryogenesis. In contrast, myogenine and MRF-4 are important only for terminal myogenic differentiation. In MyoD-/- myOblast terminal myogenic differentiation is impaired and the expression of M-cadherin is also reduced. We also known that N-cadherin can largely compensate for lack of M-cadherin. In our study we will want to know how myogenic factors regulate M- and N-cadherin expression. Firstly we cloned M-cadherin promoter and found that MyoD could activate M-cadherin by binding to it’s promoter. Then we mutated the putative E-boxs of M-cadherin promoter to define the MRF binding sites. At the same time we cloned and analysed N-cadherin promoter and compare it’s regulation by MRFs with that of M-cadherin. We found that the regulation of M- and N-Cadherin by MRFs and MEF-2C were different. In the future we will use CHIP assay to analyze the physiology meaning in vivo of this regulation by MRFs.
    Appears in Collections:[生命科學研究所 ] 博碩士論文

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