Examination of DNA methyltransferase expression in cloned embryos reveals an essential role for Dnmt1 in bovine development.
Examination of DNA methyltransferase expression in cloned embryos reveals an essential role for Dnmt1 in bovine development.
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DOI：
10.1002/mrd.21306
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发表时间：
2011-05
影响因子：
2.5
通讯作者：
Long, Charles R.
中科院分区：
文献类型：
--
作者：
Golding, Michael C.;Williamson, Gayle L.;Stroud, Todd K.;Westhusin, Mark E.;Long, Charles R.
在体细胞核移植(SCNT)的研究中，卵母细胞内的因子对移植的细胞核进行表观遗传重编程的能力对于克隆胚胎发育是至关重要的。然而，在重构胚胎中经常观察到X染色体失活、印记基因异常表达和基因组DNA高甲基化的异常模式，这表明在这一过程中存在异常。为了更好地理解SCNT重编程背后的表观遗传学事件，我们试图确定克隆胚胎中观察到的异常DNA甲基化水平是否是由于卵母细胞未能正确重编程胚胎和体细胞核之间的DNA甲基转移酶家族(DNMT)的转录和差异生化调节所致。为了解决这个问题，我们对通过体外受精(IVF)、孤雌激活和SCNT产生的牛着床前胚胎中的DNMT转录进行了实时定量。到8-细胞阶段，编码Dnmt1的转录本在克隆胚胎中显著下调；可能是对基因组高甲基化状态的反应，而从头开始的甲基转移酶保持着与体外受精和孤雌生殖对应的表达模式。使用短干扰RNA在体外受精胚胎中耗尽胚胎/母体Dnmt1转录本，虽然能够降低基因组DNA甲基化水平，但导致8/16细胞阶段的发育停滞。相比之下，来自稳定的、Dnmt1缺失的供体细胞系的SCNT胚胎发育到囊胚期，但未能足月妊娠。我们的结果表明Dnmt1在牛着床前发育过程中起着重要的作用，并提示该基因家族在SCNT胚胎中适当的转录重编程。
In studies of somatic cell nuclear transfer (SCNT), the ability of factors within the oocyte to epigenetically reprogram transferred nuclei is essential for clone embryonic development to proceed. However, irregular patterns of X-chromosome inactivation, abnormal expression of imprinted genes and genomic DNA hypermethylation are frequently observed in reconstructed embryos suggesting abnormalities in this process. To better understand the epigenetic events underlying SCNT reprogramming, we sought to determine whether the abnormal DNA methylation levels observed in cloned embryos result from a failure of the oocyte to properly reprogram transcription versus differential biochemical regulation of the DNA methyltransferase family of enzymes (DNMTs) between embryonic and somatic nuclei. To address this question, we conducted real time quantitation of Dnmt transcripts in bovine preimplantation embryos generated though in vitro fertilization (IVF), parthenogentic activation and SCNT. By the 8-Cell stage, transcripts encoding Dnmt1 become significantly down-regulated in cloned embryos; likely in response to the state of genomic hypermethylation, while the de novo methyltranserases maintain an expression pattern indistinguishable from their IVF and parthenote counterparts. Depletion of embryonic / maternal Dnmt1 transcripts within IVF embryos using short-interfering RNAs, while able to lower genomic DNA methylation levels, resulted in developmental arrest at the 8/16-cell stage. In contrast, SCNT embryos derived from a stable, Dnmt1-depleted donor cell line develop to blastocyst stage but failed to carry to term. Our results indicate an essential role for Dnmt1 during bovine preimplantation development and suggest proper transcriptional reprogramming of this gene family in SCNT embryos.
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影响因子：
1.2
作者：
Golding, MC;Westhusin, ME
通讯作者：
Westhusin, ME
影响因子：
3.6
作者：
Chung, YG;Ratnam, S;Latham, KE
通讯作者：
Latham, KE
影响因子：
64.5
作者：
Goldberg, Aaron D.;Allis, C. David;Bernstein, Emily
通讯作者：
Bernstein, Emily
DOI：
10.1089/15204559950020102
发表时间：
1999-01-01
期刊：
Cloning
影响因子：
--
作者：
De Sousa, P A;Winger, Q;Westhusin, M E
通讯作者：
Westhusin, M E
影响因子：
4.8
作者：
Kang, YK;Koo, DB;Han, YM
通讯作者：
Han, YM