Jaenisch Rudolf, Hochedlinger Konrad, Eggan Kevin
Whitehead Institute, Massachusetts Institute of Technology, 9 Cambridge Center, Cambridge, MA 02142, USA.
Novartis Found Symp. 2005;265:107-18; discussion 118-28.
The full-term development of sheep, cows, goats, pigs and mice has been achieved through the transfer of somatic cell nuclei into enucleated oocytes. Despite these successes, mammalian cloning remains an inefficient process, with a preponderance of reconstructed embryos failing at early- to mid-gestation stages of development. The small percentage of conceptuses that survive to term are characterized by a high mortality rate and frequently display grossly increased placental and birth weights. It is likely that inappropriate expression of key developmental genes may contribute to lethality of cloned embryos. One of the most interesting issues of nuclear cloning is the question of genomic reprogramming, i.e. whether successful cloning requires the resetting of epigenetic modifications which are characteristic of the adult donor nucleus. Processes such as X-inactivation and genomic imprinting are known to depend on epigenetic modifications of the genome. The classical nuclear transfer experiments with frogs have suggested that the source of the donor nucleus affects the phenotype of the clone. We have, using expression profiling, compared gene expression in clones derived from embryonic stem (ES) cells and from somatic donor cell nuclei and find substantial gene dysregulation. Our results suggest that faulty reprogramming is caused by the nuclear cloning procedure itself. In addition, the type of donor nucleus contributes to the abnormal expression pattern seen in cloned animals. One of the major unresolved issues has been whether nuclei of terminally differentiated cells can be reprogrammed by transfer into the oocyte. To address this question we have derived monoclonal mice from B and T cells and used the genetic rearrangements of the immunoglobulin and T cell receptor genes to retrospectively verify the differentiation state of the donor nucleus. Finally, we discuss our recent studies on the reprogramming of nuclei from terminally differentiated neurons and from cancer cells.
通过将体细胞核转移到去核卵母细胞中，已经实现了绵羊、奶牛、山羊、猪和小鼠的足月发育。尽管取得了这些成功，但哺乳动物克隆仍然是一个效率低下的过程，大量重构胚胎在妊娠早期至中期发育阶段失败。存活至足月的少数胎儿具有高死亡率的特征，并且经常表现出胎盘和出生体重显著增加。关键发育基因的不适当表达可能导致克隆胚胎的致死性。核克隆最有趣的问题之一是基因组重编程问题，即成功克隆是否需要重置成年供体细胞核特有的表观遗传修饰。已知诸如X染色体失活和基因组印记等过程依赖于基因组的表观遗传修饰。经典的青蛙核移植实验表明，供体细胞核的来源会影响克隆体的表型。我们通过表达谱分析，比较了源自胚胎干细胞（ES）和体供体细胞细胞核的克隆体中的基因表达，发现了大量的基因失调。我们的结果表明，错误的重编程是由核克隆程序本身引起的。此外，供体细胞核的类型导致了克隆动物中出现的异常表达模式。一个主要未解决的问题是终末分化细胞的核是否可以通过转移到卵母细胞中进行重编程。为了解决这个问题，我们从B细胞和T细胞中获得了单克隆小鼠，并利用免疫球蛋白和T细胞受体基因的基因重排来回顾性验证供体细胞核的分化状态。最后，我们讨论了我们最近关于终末分化神经元和癌细胞核重编程的研究。
