MYOD1 GENE AND PROTEIN EXPRESSION IN PEDIATRIC TUMORS

编码小鼠MyoD 1蛋白的互补DNA（cDNA）被克隆到大肠杆菌中。 由A博士分离。Lasser和发现诱导间充质中的肌生成 细胞细菌融合蛋白的多克隆抗血清（兔）， MyoD 1序列被提出，并显示MYoD 1是磷蛋白 存在于增殖成肌细胞的细胞核中， 肌管，但不是在非肌肉细胞类型。MyoD 1蛋白被发现 含有与在所述蛋白质中的区域具有显著相似性的结构域（22个氨基酸）。 myc家族蛋白和预测的转录蛋白产物 果蝇中参与神经元决定的单位， 神经系统的发育。 基于上述关于MyoD 1蛋白同源性的信息， 由原始成肌细胞和神经转录单位共享， 知识：（1）某些肌原性组织（头部和颈部）起源于 胚胎学上来自神经嵴，和（2）儿童的某些肿瘤 含有原始神经和横纹肌母细胞成分 （外胚间充质瘤，平滑肌母细胞瘤），我们将研究表达 在RMS和PNET细胞系中的MYoD 1基因和蛋白质， 如果MyoD 1基因可以在不表达肌源性 表型，但有能力这样做。该研究将采用北方 并对MYOD 1基因和蛋白进行相应的Western印迹分析， 和使用针对MyoD 1的多克隆抗血清的免疫荧光 蛋白质以检测蛋白质的细胞学定位。质粒， 携带MYOD 1基因和抗血清将成为我们可用的博士。 A.Lasser已经联系并同意向我们提供 材料 此外，MyoD 1蛋白的存在和MyoD 1蛋白的表达与MyoD 1蛋白的表达相关。 基因在这些RMS细胞系治疗前后， 分化剂和TGF-β将通过免疫荧光进行评价， Western blotting和北方印迹，以确定是否 MyoD 1基因和产物可作为RMS分化标记。

A complementary DNA (cDNA) encoding for the mouse MyoD1 protein was isolated by Dr. A. Lasser and found to induce myogenesis in mesenchymal cells. Polyclonal antisera (rabbit) to bacterial fusion proteins containing the MyoD1 sequence were raised and showed that MYoD1 is a phosphoprotein present in the nuclei of proliferating myoblasts and differentiated myotubes, but not in nonmuscle cell types. The MyoD1 protein was found to contain a domain (22 amino acid) with marked similarity to a region in the myc-family proteins and to the predicted protein products of transcription units in Drosophila which are involved in neuronal determination and the development of nervous system. Based on the above information about the homologies of the MyoD1 protein shared by primitive myoblasts and neural transcription units, and on the knowledge that: (1) certain myogenous tissues (head and neck) originate embryologically from the neural crest, and (2) certain tumors of childhood contain both primitive neural and rhabdomyoblastic components (ectomesenchymoma, medullomyoblastoma), we will investigate the expression of the MYoD1 gene and protein in the RMS and PNET cell lines, to find out if the MyoD1 gene can be expressed in tissues not expressing a myogenous phenotype, but with the ability to do so. The study will employ Northern and Western blot analysis of the MYOD1 gene and protein correspondingly, and immunofluorescence using the polyclonal antisera against the MyoD1 protein to detect cytologic localization of the protein. The plasmids, carrying the MYOD1 gene and antisera will become available to us by Dr. A.Lasser who has already been contacted and agreed to provide us with the material. Moreover, the presence of the MyoD1 protein and the expression of the MyoD1 gene in these RMS cell lines before and after treatment with differentiating agents and TGF-b will be evaluated by immunofluorescence, Western blotting and Northern blotting correspondingly, to find out whether the MyoD1 gene and product can serve as a marker of differentiation in RMS.