Activation of the maternally preset program of apoptosis by microinjection of 5‐aza‐2′‐deoxycytidine and 5‐methyl‐2′‐deoxycytidine‐5′‐triphosphate in Xenopus laevis embryos
Activation of the maternally preset program of apoptosis by microinjection of 5‐aza‐2′‐deoxycytidine and 5‐methyl‐2′‐deoxycytidine‐5′‐triphosphate in Xenopus laevis embryos
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在非洲爪蟾胚胎中显微注射 5-aza-2-脱氧胞苷和 5-甲基-2-脱氧胞苷-5-三磷酸激活母体预设的细胞凋亡程序
DOI：
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
K. Shiokawa
中科院分区：
文献类型：
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作者：
C. Kaito;M. Kai;T. Higo;E. Takayama;H. Fukamachi;K. Sekimizu;K. Shiokawa
本研究检查了非洲爪蟾受精卵显微注射 5-aza-2'-脱氧胞苷 (5-Aza-CdR)（诱导 DNA 低甲基化）和 5-甲基-2'-脱氧胞苷-5'-三磷酸（5-甲基-dCTP）（诱导 DNA 高甲基化）对胚胎发生的影响。注射了这些类似物中的任何一种的胚胎在囊胚中期之前都正常分裂，但在原肠胚早期经历了大量的细胞解离并停止发育。此处出现的解离细胞通过末端脱氧核糖核苷酸转移酶介导的脱氧尿苷三磷酸-地高辛缺口末端标记呈阳性，并含有带有浓缩染色质的碎片细胞核。这些细胞的 DNA 在电泳上形成了一个“梯子”。此外，通过共注射 Bcl-2 mRNA 和正常代谢物（分别为 CdR 和 dCTP），5-Aza-CdR 和 5-甲基-dCTP 诱导的细胞解离被推迟 2-3 小时。使用针对 5-甲基-胞嘧啶的特异性抗体，我们证实 5-Aza-CdR 会诱导低甲基化，而 5-甲基-dCTP 在细胞解离开始之前会诱导非洲爪蟾胚胎的高甲基化。放射性前体的掺入表明，在注射 5-Aza-CdR 和注射 5-甲基-dCTP 的胚胎中，DNA 和 RNA 的合成均受到显着抑制。这些结果表明，5-Aza-CdR 和 5-甲基-dCTP 可能通过改变 DNA 甲基化并抑制 DNA 复制和/或转录而掺入 DNA 并诱导细胞凋亡。
The present study examines the effects on embryogenesis of microinjecting Xenopus laevis fertilized eggs with 5‐aza‐2′‐deoxycytidine (5‐Aza‐CdR), which induces hypomethylation of DNA, and 5‐methyl‐2′‐ deoxycytidine‐5′‐triphosphate (5‐methyl‐dCTP), which induces hypermethylation of DNA. Embryos injected with either one of these analogs cleaved normally until the mid‐blastula stage, but underwent massive cell dissociation and stopped development at the early gastrula stage. Dissociated cells that appeared here were positive by terminal deoxyribonucleotidyl transferase‐mediated deoxyuridine triphosphate–digoxigenin nick end‐labeling and contained fragmented nuclei with condensed chromatin. The DNA from these cells formed a ‘ladder’ on electrophoresis. Furthermore, the induction of cell dissociation by 5‐Aza‐CdR and 5‐methyl‐dCTP was postponed by 2–3 h by co‐injection of Bcl‐2 mRNA and the normal metabolite (CdR and dCTP, respectively). Using a specific antibody against 5‐methyl‐cytosine, we confirmed that 5‐Aza‐CdR induces hypomethylation, whereas 5‐methyl‐dCTP induces hypermethylation in X. laevis embryos before the onset of cell dissociation. Incorporation of radioactive precursors revealed that synthesis of DNA, and also RNA, is inhibited significantly in both 5‐Aza‐CdR‐injected and 5‐methyl‐dCTP‐injected embryos. These results show that 5‐Aza‐CdR and 5‐methyl‐dCTP are incorporated into DNA and induce apoptosis, probably through alteration of DNA methylation coupled with inhibition of DNA replication and/or transcription.
非洲爪蟾原肠胚形成早期发育调节的细胞凋亡激活导致细胞周期间期细胞周期蛋白 A 降解。
DOI：
10.1242/dev.124.16.3185
发表时间：
1997
期刊：
Development (Cambridge, England)
影响因子：
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作者：
Stack,JH;Newport,JW
通讯作者：
Newport,JW
通过显微注射 5-甲基 dCTP 直接诱导海胆胚胎 DNA 高甲基化，刺激早期组蛋白基因表达并导致发育停滞。
影响因子：
2.7
作者：
Chen,J;Maxson,R;Jones,PA
通讯作者：
Jones,PA