Identification of the gene coding for the Endo B murine cytokeratin and its methylated, stable inactive state in mouse nonepithelial cells.
Identification of the gene coding for the Endo B murine cytokeratin and its methylated, stable inactive state in mouse nonepithelial cells.
复制标题
鉴定编码 Endo B 鼠细胞角蛋白的基因及其在小鼠非上皮细胞中的甲基化、稳定失活状态。
DOI：
10.1101/gad.2.5.505
复制
发表时间：
1988
影响因子：
10.5
通讯作者：
Ceceña,G
中科院分区：
文献类型：
--
作者：
Oshima,RG;Trevor,K;Shevinsky,LH;Ryder,OA;Ceceña,G
Endo B型角蛋白中间丝蛋白在小鼠发育的4- 8细胞阶段首次表达。在成人中，其表达仅限于多种简单上皮细胞类型。为了研究Endo B受限表达的机制，通过Southern杂交分析，从小鼠基因组中发现的5个不同的Endo B基因中鉴定了编码Endo B的基因，并克隆了其中4个基因的全部或部分。核运行实验表明，Endo B的表达在转录水平上受到调控。活性基因的5'端被称为Endo β 1，在不表达Endo B的成纤维细胞和成肌细胞中被发现高度甲基化，呈相对耐核酸酶的染色质构象，但在内胚层细胞或F9胚胎癌细胞中被发现甲基化不足，对核酸酶消化相对敏感。Endo B β 1基因在成纤维细胞中的失活状态似乎非常稳定，因为由表达同源人蛋白、角蛋白18和小鼠成纤维细胞的HeLa细胞融合形成的体细胞杂种继续表达角蛋白18，但不激活Endo B的表达。同样，小鼠内胚层细胞和成纤维细胞的融合产生的杂交种不会抑制Endo B的表达。这些结果表明，Endo B转录受到两种不同机制的限制。在成纤维细胞或成肌细胞等体细胞中，表达可能受到甲基化和稳定的非允许转录状态的限制。然而，在胚胎癌细胞中，Endo B β 1基因是低甲基化的，处于一个相对核酸酶敏感的构象中，但它受到一个额外的负调控机制的限制。
The Endo B type-I keratin intermediate filament protein is first expressed at the 4- to 8-cell stage of mouse development. In the adult, its expression is restricted to a variety of simple epithelial cell types. To investigate the mechanisms responsible for the restricted expression of Endo B, the gene coding for Endo B has been identified from among the five different Endo B genes found in the mouse genome by Southern hybridization analysis and cloning all or part of four of the genes. Nuclear run-on experiments demonstrate that Endo B expression is regulated at the level of transcription. The 5' end of the active gene, designated Endo beta 1, was found to be highly methylated and in a relatively nuclease-resistant chromatin conformation in fibroblasts and myoblasts that do not express Endo B, but undermethylated and relatively sensitive to nuclease digestion in endodermal cells or F9 embryonal carcinoma cells. The inactive state of the Endo B beta 1 gene in fibroblast appears to be very stable, because somatic cell hybrids formed by the fusion of HeLa cells, which express the homologous human protein, keratin 18, and mouse fibroblasts, continue to express keratin 18 but do not activate Endo B expression. Similarly, the fusion of mouse endodermal cells and fibroblasts results in hybrids that do not extinguish Endo B expression. These results suggest that Endo B transcription is limited by two different mechanisms. In somatic cells such as fibroblasts or myoblasts, expression may be restricted by methylation and a stable, nonpermissive transcriptional state. However, in embryonal carcinoma cells, the Endo B beta 1 gene is undermethylated and in a relatively nuclease-sensitive conformation, but it is restricted by an additional, negative regulatory mechanism.