Inositol requiring mutant 80 (INO80) is a chromatin remodeler that regulates pluripotency maintenance of embryonic stem cells and reprogramming of somatic cells into pluripotent stem cells. However, the roles and mechanisms of INO80 in porcine pre-implantation embryonic development remain largely unknown. Here, we show that INO80 modulates trophectoderm epithelium permeability to promote porcine blastocyst development. The INO80 protein is highly expressed in the nuclei during morula-to-blastocyst transition. Functional studies revealed that RNA interference (RNAi)-mediated knockdown of INO80 severely blocks blastocyst formation and disrupts lineage allocation between the inner cell mass and trophectoderm. Mechanistically, single-embryo RNA sequencing revealed that INO80 regulates multiple genes, which are important for lineage specification, tight junction assembly, and fluid accumulation. Consistent with the altered expression of key genes required for tight junction assembly, a permeability assay showed that paracellular sealing is defective in the trophectoderm epithelium of INO80 knockdown blastocysts. Importantly, aggregation of 8-cell embryos from the control and INO80 knockdown groups restores blastocyst development and lineage allocation via direct complementation of the defective trophectoderm epithelium. Taken together, these results demonstrate that INO80 promotes blastocyst development by regulating the expression of key genes required for lineage specification, tight junction assembly, and fluid accumulation.
INO80蛋白是一种染色质重塑因子，参与调控胚胎干细胞多能性的维持以及体细胞诱导为多能干细胞的重编程过程。然而，INO80在猪着床前胚胎发育中的作用及机制尚不清楚。该研究中，我们证实INO80调节滋养层细胞的渗透能力以促进猪囊胚发育。在桑椹胚向囊胚转变过程，INO80蛋白在胚胎细胞核中呈高表达。功能研究证明RNA干扰介导的INO80敲低导致囊胚形成失败，并破坏囊胚中内细胞团和滋养层细胞谱系的分布。单胚胎RNA测序分析揭示INO80调控许多与囊胚细胞谱系分化、紧密连接组装和囊胚腔液体积累等基因的表达。细胞渗透性分析证实INO80敲低囊胚滋养层细胞之间的细胞封闭发生缺陷。对照组和INO80敲低组8-细胞胚胎的聚合试验表明，通过补偿缺陷型滋养层细胞可以诱导INO80敲低胚胎发育成囊胚并恢复正常的细胞谱系分布。因此，以上研究结果证明染色质重塑因子INO80通过调控细胞谱系特化、紧密连接组装和液体聚集等关键基因的表达以促进囊胚发育。.
Keywords: Blastocyst; INO80; Permeability; Tight junction; Trophectoderm.