Kisliakova T V, Kustova M E, Lianguzova M S, Malashcheva A B, Strunnikova M A, Suchkova I O, Pospelov V A, Patkin E L
Institute of Cytology RAS, St. Petersburg.
Tsitologiia. 2000;42(11):1060-8.
Our previous study demonstrated the high incidence of non-induced DNA single strand breaks (SSB) in preimplantation mouse embryo genom (Patkin et al., 1994). F9 mouse teratocarcinoma cell line is an in vitro model for early embryonal differentiation, since F9 cells remind in many respects the inner cell mass cells of mouse blastocyst and are capable of differentiation under retinoic acid (RA) and dibutyryl cAMP (db-cAMP) treatment. Using gap filling reaction of F9 metaphase chromosomes and single-cell DNA electrophoresis, we have observed multiple SSB in undifferentiated F9 cells as well as in F9 cells at the early steps of RA-induced differentiation (days of RA treatment), but not in terminally differentiated F9 cells and in mouse embryonal fibroblasts. Rad51 nuclear protein that binds specifically single stranded DNA is highly expressed in all cells of undifferentiated F9 population and is not expressed in terminally differentiated F9 population. Multiple SSB could lead to enhanced rate of sister chromatid exchanges (SCE) in F9 cells. In undifferentiated F9 population the level of SCE was 9.6 +/- 0.44 per metaphase, that was not higher than in NIH 3T3 cell line. However, RA treatment for 48 h led to rising the SCE level up to 16.68 +/- 0.72 followed by its decrease to the initial rate by 72 h of RA treatment. Since the enhanced level of SSB in undifferentiated F9 cells and in mouse blastocyst does not normally lead to chromosomal instability, we consider SSB to be a natural consequence of fast-going DNA replication in these cells.
我们之前的研究表明，在植入前小鼠胚胎基因组中，非诱导性DNA单链断裂（SSB）的发生率很高（帕特金等人，1994年）。F9小鼠畸胎瘤细胞系是早期胚胎分化的体外模型，因为F9细胞在许多方面类似于小鼠囊胚的内细胞团细胞，并且在视黄酸（RA）和二丁酰环磷腺苷（db-cAMP）处理下能够分化。利用F9中期染色体的缺口填补反应和单细胞DNA电泳，我们在未分化的F9细胞以及RA诱导分化早期阶段（RA处理天数）的F9细胞中观察到多个SSB，但在终末分化的F9细胞和小鼠胚胎成纤维细胞中未观察到。特异性结合单链DNA的Rad51核蛋白在未分化的F9群体的所有细胞中高表达，而在终末分化的F9群体中不表达。多个SSB可能导致F9细胞中姐妹染色单体交换（SCE）率增加。在未分化的F9群体中，每个中期的SCE水平为9.6±0.44，不高于NIH 3T3细胞系。然而，RA处理48小时导致SCE水平升至16.68±0.72，随后在RA处理72小时时降至初始水平。由于未分化的F9细胞和小鼠囊胚中SSB水平的升高通常不会导致染色体不稳定，我们认为SSB是这些细胞中快速进行的DNA复制的自然结果。
