Byrne J A, Simonsson S, Gurdon J B
Wellcome Cancer Research U.K. Institute, Tennis Court Road, Cambridge CB2 1QR, United Kingdom.
Proc Natl Acad Sci U S A. 2002 Apr 30;99(9):6059-63. doi: 10.1073/pnas.082112099. Epub 2002 Apr 23.
Nuclear transplantation is one of the very few ways by which the genetic content and capacity for nuclear reprogramming can be assessed in individual cells of differentiated somatic tissues. No more than 6% of the cells of differentiated tissues have thus far been shown to have nuclei that can be reprogrammed to elicit the formation of unrelated cell types. In Amphibia, about 25% of such nuclear transfers form morphologically abnormal partial blastulae that die within 24 h. We have investigated the genetic content and capacity for reprogramming of those nuclei that generate partial blastulae, using as donors the intestinal epithelium cells of feeding Xenopus larvae. We have analyzed single nuclear transplant embryos obtained directly from intestinal tissue, thereby avoiding any genetic or epigenetic changes that might accumulate during cell culture. The expression of the intestine-specific gene intestinal fatty acid binding protein is extinguished by at least 10(4) times, within a few hours of nuclear transplantation. At the same time several genes that are normally expressed only in early embryos are very strongly activated in nuclear transplant embryos, but to an unregulated extent. Remarkably, cells from intestine-derived partial blastulae, when grafted to normal host embryos, contribute to several host tissues and participate in the normal 100-fold increase in axial muscle over several months. Thus, cells of defective cloned embryos unable to survive for more than 1 day can be reprogrammed to participate in new directions of differentiation and to maintain indefinite growth, despite the abnormal expression of early genes.
核移植是极少数能够在分化的体细胞组织的单个细胞中评估遗传内容和核重编程能力的方法之一。到目前为止，已显示分化组织中不超过6%的细胞具有可重编程以引发无关细胞类型形成的细胞核。在两栖动物中，约25%的此类核移植形成形态异常的部分囊胚，这些囊胚在24小时内死亡。我们使用正在进食的非洲爪蟾幼体的肠上皮细胞作为供体，研究了那些产生部分囊胚的细胞核的遗传内容和重编程能力。我们分析了直接从肠组织获得的单核移植胚胎，从而避免了细胞培养过程中可能积累的任何遗传或表观遗传变化。在核移植后的几个小时内，肠特异性基因肠脂肪酸结合蛋白的表达至少降低了10^4倍。与此同时，几个通常仅在早期胚胎中表达的基因在核移植胚胎中被强烈激活，但激活程度不受调控。值得注意的是，来自肠源部分囊胚的细胞移植到正常宿主胚胎中时，会参与多个宿主组织，并在几个月内使轴向肌肉正常增加100倍。因此，尽管早期基因表达异常，但无法存活超过1天的有缺陷克隆胚胎的细胞仍可被重编程，以参与新的分化方向并维持无限生长。
