State Key Laboratory of Medicinal Chemical Biology, College of Pharmacy, Nankai University, Tianjin 300350, China.
Swiss Federal Institute of Technology Zurich, Department of Biology, Institute of Molecular Health Sciences, Chair of RNAi and Genome Integrity, Zurich 8093, Switzerland.
Development. 2018 Jun 11;145(11):dev160531. doi: 10.1242/dev.160531.
Haploid embryonic stem cells (haESCs) have been extensively applied in forward and reverse genetic screening. However, a mammalian haploid somatic cell line is difficult to achieve because of spontaneous diploidization in differentiation. As a non-human primate species, monkeys are widely used in basic and pre-clinical research in which haploid cells are restricted to ESCs. Here, we report that rhesus monkey haESCs in an optimized culture medium show naïve-state pluripotency and stable haploidy. This model facilitated the derivation of haploid neural progenitor cells (haNPCs), which maintained haploidy and differentiation potential into neurons and glia for a long period High-throughput trapping mutations can be efficiently introduced into haNPCs via transposons. This system proves useful when identifying gene targets of neural toxicants via a proof-of-concept experiment. Using CRISPR/Cas9 editing, we confirmed that , from the candidate gene list, is a resistance gene of A803467 (a tetrodotoxin-like toxicant). This model is the first non-human primate haploid somatic cell line with proliferative ability, multipotency and an intact genome, thus providing a cellular resource for recessive genetic and potential drug screening.
单倍体胚胎干细胞 (haESCs) 已广泛应用于正向和反向遗传筛选。然而，由于分化过程中的自发二倍化，哺乳动物单倍体体细胞系难以实现。作为一种非人类灵长类动物，猴子广泛应用于基础和临床前研究，其中单倍体细胞仅限于胚胎干细胞。在这里，我们报告说，在优化的培养基中，恒河猴 haESCs 表现出原始状态的多能性和稳定的单倍性。该模型促进了单倍体神经祖细胞 (haNPCs) 的衍生，其单倍体和分化为神经元和神经胶质的潜力可以长期维持。通过转座子，可以有效地将高通量捕获突变引入 haNPCs。通过概念验证实验，当鉴定神经毒素的基因靶标时，该系统非常有用。使用 CRISPR/Cas9 编辑，我们证实从候选基因列表中，是 A803467（一种河豚毒素样毒素）的抗性基因。该模型是第一个具有增殖能力、多能性和完整基因组的非人类灵长类动物单倍体体细胞系，因此为隐性遗传和潜在药物筛选提供了细胞资源。
