Spadafora C, Riccardi P
J Mol Biol. 1985 Dec 20;186(4):743-58. doi: 10.1016/0022-2836(85)90394-8.
The chromatin structure of the ribosomal DNA in Xenopus laevis was studied by micrococcal nuclease digestions of blood, liver and embryonic cell nuclei. We have found that BglI-restricted DNA from micrococcal nuclease-digested blood cell nuclei has an increased electrophoretic mobility compared to the undigested control. Micrococcal nuclease digestion of liver cell nuclei causes a very slight shift in mobility, only in the region of the spacer containing the "Bam Islands". In contrast, the mobility of ribosomal DNA in chromatin of embryonic cells, under identical digestion conditions, remains unaffected by the nuclease activity. Denaturing gels or ligase action on the nuclease-treated DNA abolishes the differences in the electrophoretic mobility. Ionic strength and ethidium bromide influence the relative electrophoretic migration of the two DNA fragment populations, suggesting that secondary structure may play an important role in the observed phenomena. In addition, restriction analysis under native electrophoretic conditions of DNA prepared from blood, liver and embryonic cells shows that blood cell DNA restriction fragments always have a faster mobility than the corresponding fragments of liver and embryo cell DNA. We therefore propose that nicking activity by micrococcal nuclease modifies the electrophoretic mobility of an unusual DNA conformation, present in blood cell, and to a lesser extent, in liver cell ribosomal chromatin. A possible function for these structures is discussed. The differences of the ribosomal chromatin structures in adult and embryonic tissues may reflect the potential of the genes to be expressed.
通过对非洲爪蟾血液、肝脏和胚胎细胞核进行微球菌核酸酶消化,研究了其核糖体DNA的染色质结构。我们发现,与未消化的对照相比,微球菌核酸酶消化的血细胞细胞核中经BglI酶切的DNA具有更高的电泳迁移率。肝细胞细胞核的微球菌核酸酶消化仅在包含“Bam岛”的间隔区引起迁移率的非常轻微变化。相反,在相同消化条件下,胚胎细胞染色质中核糖体DNA的迁移率不受核酸酶活性影响。对核酸酶处理后的DNA进行变性凝胶电泳或连接酶处理,可消除电泳迁移率的差异。离子强度和溴化乙锭影响两个DNA片段群体的相对电泳迁移,表明二级结构可能在观察到的现象中起重要作用。此外,在天然电泳条件下对血液、肝脏和胚胎细胞制备的DNA进行限制性分析表明,血细胞DNA限制性片段的迁移率总是比肝细胞和胚胎细胞DNA的相应片段更快。因此,我们提出微球菌核酸酶的切口活性改变了血细胞中存在的一种异常DNA构象的电泳迁移率,在肝细胞核糖体染色质中程度较小。讨论了这些结构可能的功能。成体和胚胎组织中核糖体染色质结构的差异可能反映了基因表达的潜力。