Identification of key factors conquering developmental arrest of somatic cell cloned embryos by combining embryo biopsy and single-cell sequencing.
Identification of key factors conquering developmental arrest of somatic cell cloned embryos by combining embryo biopsy and single-cell sequencing.
复制标题
通过胚胎活检和单细胞测序相结合，鉴定克服体细胞克隆胚胎发育停滞的关键因素。
DOI：
10.1038/celldisc.2016.10
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发表时间：
2016
期刊：
影响因子：
33.5
通讯作者：
Gao S
中科院分区：
文献类型：
--
作者：
Liu W;Liu X;Wang C;Gao Y;Gao R;Kou X;Zhao Y;Li J;Wu Y;Xiu W;Wang S;Yin J;Liu W;Cai T;Wang H;Zhang Y;Gao S
分化的体细胞可以通过体细胞核移植重新编程为全能胚胎。然而，大多数克隆胚胎在早期停滞，其潜在的分子机制在很大程度上仍未被探索。在这里，我们首先开发了一个体细胞核移植胚胎活组织检查系统，该系统可以在两个或四个细胞阶段，使我们能够准确地追踪被活检胚胎的发育命运。然后，通过对不同发育命运的体细胞核移植胚胎进行单细胞转录组测序，我们确定了组蛋白H3赖氨酸9三甲基化去甲基化酶Kdm4b的失活是克隆胚胎两细胞停滞的屏障。此外，我们通过单细胞分析发现，另一个组蛋白去甲基酶Kdm5b的失活是克隆胚胎在四细胞期停滞的原因。联合注射Kdm4b和Kdm5b可以恢复体细胞核移植胚胎的转录水平，显著提高囊胚发育率(95%以上)和克隆小鼠的产量。因此，我们的研究为确定导致体细胞克隆胚胎发育停滞的关键因素提供了一种有效的途径。
Differentiated somatic cells can be reprogrammed into totipotent embryos through somatic cell nuclear transfer. However, most cloned embryos arrest at early stages and the underlying molecular mechanism remains largely unexplored. Here, we first developed a somatic cell nuclear transfer embryo biopsy system at two- or four-cell stage, which allows us to trace the developmental fate of the biopsied embryos precisely. Then, through single-cell transcriptome sequencing of somatic cell nuclear transfer embryos with different developmental fates, we identified that inactivation of Kdm4b, a histone H3 lysine 9 trimethylation demethylase, functions as a barrier for two-cell arrest of cloned embryos. Moreover, we discovered that inactivation of another histone demethylase Kdm5b accounts for the arrest of cloned embryos at the four-cell stage through single-cell analysis. Co-injection of Kdm4b and Kdm5b can restore transcriptional profiles of somatic cell nuclear transfer embryos and greatly improve the blastocyst development (over 95%) as well as the production of cloned mice. Our study therefore provides an effective approach to identify key factors responsible for the developmental arrest of somatic cell cloned embryos.