National Engineering Research Center for Breeding Swine Industry, College of Animal Science, South China Agricultural University, Guangzhou, Guangdong, China.
Guangdong Provincial Key Laboratory of Agro-animal Genomics and Molecular Breeding, College of Animal Science, South China Agricultural University, Guangzhou, Guangdong, China.
Mol Reprod Dev. 2021 Mar;88(3):228-237. doi: 10.1002/mrd.23460. Epub 2021 Mar 1.
Ectopic expression of Xist on the putative active X chromosome is a primary cause of the low developmental efficiency of cloned mouse and pig embryos. Suppression of abnormal Xist expression via gene knockout or RNA interference (RNAi) can significantly enhance the developmental competence of cloned mouse and pig embryos. RLIM is a Xist expression activator, whereas REX1 is an Xist transcription inhibitor, as RLIM triggers Xist expression by mediating the proteasomal degradation of REX1 to induce imprinted and random X chromosome inactivation in mice. This study aimed to test whether the knockdown of RLIM and overexpression of REX1 can repress aberrant Xist expression and improve the developmental ability of cloned male pig embryos. Results showed that injection of anti-RLIM small interfering RNA significantly decreased Xist messenger RNA abundance, increased REX1 protein level, and enhanced the preimplantation development of cloned male porcine embryos. These positive effects were not observed in cloned male pig embryos injected with REX1 expression plasmid, which might be due to the low expression efficiency of injected REX1 plasmid and/or the short half-life of expressed REX1 protein. The findings from this study indicated that RLIM participated in the ectopic activation of Xist expression in cloned pig embryos by targeting REX1 degradation. Furthermore, this study provided a new method to improve cloned pig embryo development by the inhibition of Xist expression via RNAi of RLIM.
Xist 在假定的活性 X 染色体上的异位表达是克隆鼠和猪胚胎发育效率低的主要原因。通过基因敲除或 RNA 干扰 (RNAi) 抑制异常 Xist 表达可以显著提高克隆鼠和猪胚胎的发育能力。RLIM 是 Xist 表达的激活剂，而 REX1 是 Xist 转录抑制剂，因为 RLIM 通过介导 REX1 的蛋白酶体降解来触发 Xist 表达，从而在小鼠中诱导印迹和随机 X 染色体失活。本研究旨在测试 RLIM 的敲低和 REX1 的过表达是否可以抑制异常 Xist 表达并提高克隆雄性猪胚胎的发育能力。结果表明，注射抗 RLIM 小干扰 RNA 显著降低了 Xist 信使 RNA 的丰度，增加了 REX1 蛋白水平，并增强了克隆雄性猪胚胎的植入前发育。在注射 REX1 表达质粒的克隆雄性猪胚胎中未观察到这些积极影响，这可能是由于注射的 REX1 质粒表达效率低和/或表达的 REX1 蛋白半衰期短所致。这项研究的结果表明，RLIM 通过靶向 REX1 降解参与了克隆猪胚胎中 Xist 表达的异位激活。此外，本研究通过 RNAi 抑制 RLIM 抑制 Xist 表达提供了一种提高克隆猪胚胎发育的新方法。
