Understanding the regulatory networks for germ cell fate specification is necessary to developing strategies for improving the efficiency of germ cell production in vitro. In this study, we developed a coupled screening strategy that took advantage of an arrayed bi-molecular fluorescence complementation (BiFC) platform for protein-protein interaction screens and epiblast-like cell (EpiLC)-induction assays using reporter mouse embryonic stem cells (mESCs). Investigation of candidate interaction partners of core human pluripotent factors OCT4, NANOG, KLF4 and SOX2 in EpiLC differentiation assays identified novel primordial germ cell (PGC)-inducing factors including BEN-domain (BEND/Bend) family members. Through RNA-seq, ChIP-seq, and ATAC-seq analyses, we showed that Bend5 worked together with Bend4 and helped mark chromatin boundaries to promote EpiLC induction in vitro. Our findings suggest that BEND/Bend proteins represent a new family of transcriptional modulators and chromatin boundary factors that participate in gene expression regulation during early germline development.

了解生殖细胞命运规范的调控网络对于制定提高体外生殖细胞生产效率的策略是必要的。在这项研究中，我们开发了一种耦合筛选策略，该策略利用阵列双分子荧光互补 (BiFC) 平台进行蛋白质-蛋白质相互作用筛选和使用报告小鼠胚胎干细胞 (mESC) 进行外胚层样细胞 (EpiLC) 诱导测定。 ）。在 EpiLC 分化测定中对核心人类多能因子 OCT4、NANOG、KLF4 和 SOX2 的候选相互作用伙伴进行研究，确定了新型原始生殖细胞 (PGC) 诱导因子，包括 BEN 结构域 (BEND/Bend) 家族成员。通过 RNA-seq、ChIP-seq 和 ATAC-seq 分析，我们表明 Bend5 与 Bend4 协同作用，帮助标记染色质边界，从而促进 EpiLC体外诱导。我们的研究结果表明，BEND/Bend 蛋白代表了一个新的转录调节剂和染色质边界因子家族，参与早期种系发育过程中的基因表达调控。