McGuigan A, Huxley C
Department of Biochemistry and Molecular Genetics, Imperial College School of Medicine at St Mary's, London, UK.
Nucleic Acids Res. 1996 Jun 15;24(12):2271-80. doi: 10.1093/nar/24.12.2271.
To determine whether yeast DNA can replicate or segregate in mammalian cells, we have transferred genomic DNA from the yeast Saccharomyces cerevisiae into mouse cells. Most of the lines contained stably integrated yeast DNA. However, in two of the lines, the yeast DNA was maintained as numerous small extrachromosomal elements which were still present after 26 cell divisions in selection but which were lost rapidly out of selection. This indicates that, although yeast DNA can replicate in mouse cells, the yeast centromere does not function to give segregation. In one cell line we observed a large novel chromosome consisting almost entirely of yeast DNA. This chromosome segregates well and contains mouse centromeric minor satellite DNA and variable amounts of major satellite DNA which probably comprise the functional centromere. The yeast DNA in the novel chromosome has a compacted chromatin structure which may be responsible for the efficient formation of anaphase bridges. Furthermore, yeast DNA integrated into mouse chromosomes forms constrictions at the point of integration. These features have previously been presumed to be hallmarks of centromeric function in transfection assays aimed at identifying putative centromeric DNA. Hence our results suggest caution be exercised in the interpretation of such assays.
为了确定酵母DNA能否在哺乳动物细胞中复制或分离，我们已将酿酒酵母的基因组DNA转入小鼠细胞。大多数细胞系含有稳定整合的酵母DNA。然而，在其中两个细胞系中，酵母DNA以大量小的染色体外元件形式存在，这些元件在选择条件下经过26次细胞分裂后仍然存在，但在无选择条件下会迅速丢失。这表明，尽管酵母DNA能在小鼠细胞中复制，但酵母着丝粒不能发挥分离功能。在一个细胞系中，我们观察到一条几乎完全由酵母DNA组成的大型新染色体。这条染色体分离良好，包含小鼠着丝粒小卫星DNA和数量可变的主要卫星DNA，这些可能构成了功能着丝粒。新染色体中的酵母DNA具有紧密的染色质结构，这可能是后期桥高效形成的原因。此外，整合到小鼠染色体中的酵母DNA在整合点处形成缢痕。在旨在鉴定假定着丝粒DNA的转染实验中，这些特征以前被认为是着丝粒功能的标志。因此，我们的结果表明，在解释此类实验时应谨慎。
