Koziol Magdalena J, Garrett Nigel, Gurdon J B
Wellcome Trust Cancer Research UK Gurdon Institute, University of Cambridge, Cambridge CB2 1QN, UK.
Curr Biol. 2007 May 1;17(9):801-7. doi: 10.1016/j.cub.2007.03.062. Epub 2007 Apr 19.
Nuclear transfer to eggs or oocytes provides a potential route for cell-replacement therapies because oocytes directly reprogram transplanted mammalian somatic-cell nuclei such that they have an embryo-like pattern of gene expression. This includes a large increase in the mRNA level of the stem-cell marker gene oct4. We have developed a novel procedure to identify new proteins that greatly increase the level of oct4 mRNA upon nuclear transfer. We have isolated Xenopus oocyte proteins that bind to the regulatory region of the mouse oct4 gene and identified these by mass spectrometry. The proteins include the retinoic-acid-receptor gamma, a known repressor of oct4 transcription, and Tpt1, a cancer-associated factor. The depletion of transcripts of retinoic-acid receptor gamma from oocytes increases oct4 and nanog transcription as expected, and depletion of tpt1 transcripts in oocytes reduces oct4 and nanog transcription in injected HeLa nuclei. An elevation of tpt1 transcripts in oocytes results in an earlier activation of oct4 transcription. Therefore, we identify a novel role for tpt1 in activating pluripotency genes upon nuclear transfer. Our results help to elucidate the mechanism by which somatic-cell nuclei are reprogrammed to have an embryo-like pattern of gene expression.
将细胞核移植到卵子或卵母细胞中为细胞替代疗法提供了一条潜在途径，因为卵母细胞可直接对移植的哺乳动物体细胞核进行重编程，使其具有类似胚胎的基因表达模式。这包括干细胞标记基因oct4的mRNA水平大幅增加。我们开发了一种新方法来鉴定那些在细胞核移植后能大幅提高oct4 mRNA水平的新蛋白质。我们分离出了与小鼠oct4基因调控区域结合的非洲爪蟾卵母细胞蛋白，并通过质谱法对其进行了鉴定。这些蛋白质包括视黄酸受体γ（一种已知的oct4转录抑制因子）和Tpt1（一种与癌症相关的因子）。从卵母细胞中去除视黄酸受体γ的转录本如预期那样增加了oct4和nanog的转录，而在卵母细胞中去除tpt1转录本则降低了注射到HeLa细胞核中的oct4和nanog的转录。卵母细胞中tpt1转录本的增加导致oct4转录的更早激活。因此，我们确定了tpt1在细胞核移植后激活多能性基因方面的新作用。我们的结果有助于阐明体细胞核被重编程以具有类似胚胎基因表达模式的机制。
