Institute of Stem Cell and Translational Cancer Research, Chang Gung Memorial Hospital at Linkou, Taoyuan, Taiwan.
Genomics Research Center, Academia Sinica, Taipei, Taiwan.
Stem Cells. 2018 Oct;36(10):1514-1524. doi: 10.1002/stem.2862. Epub 2018 Jul 29.
Stem cell surface markers may facilitate a better understanding of stem cell biology through molecular function studies or serve as tools to monitor the differentiation status and behavior of stem cells in culture or tissue. Thus, it is important to identify additional novel stem cell markers. We used glycoproteomics to discover surface glycoproteins on human embryonic stem cells (hESCs) that may be useful stem cell markers. We found that a surface glycoprotein, leucine-rich repeat neuronal protein 1 (LRRN1), is expressed abundantly on the surface of hESCs before differentiation into embryoid bodies (EBs). Silencing of LRRN1 with short hairpin RNA (shLRRN1) in hESCs resulted in decreased capacity of self-renewal, and skewed differentiation toward endoderm/mesoderm lineages in vitro and in vivo. Meanwhile, the protein expression levels of the pluripotency factors OCT4, NANOG, and SOX2 were reduced. Interestingly, the mRNA levels of these pluripotency factors were not affected in LRRN1 silenced cells, but protein half-lives were substantially shortened. Furthermore, we found LRRN1 silencing led to nuclear export and proteasomal degradation of all three pluripotency factors. In addition, the effects on nuclear export were mediated by AKT phosphorylation. These results suggest that LRRN1 plays an important role in maintaining the protein stability of pluripotency factors through AKT phosphorylation, thus maintaining hESC self-renewal capacity and pluripotency. Overall, we found that LRRN1 contributes to pluripotency of hESC by preventing translocation of OCT4, NANOG, and SOX2 from nucleus to cytoplasm, thereby lessening their post-translational modification and degradation. Stem Cells 2018;36:1514-1524.
干细胞表面标志物可通过分子功能研究促进对干细胞生物学的更好理解，或作为工具来监测培养物或组织中干细胞的分化状态和行为。因此，识别其他新的干细胞标志物非常重要。我们使用糖蛋白质组学来发现人胚胎干细胞（hESC）表面的糖蛋白，这些糖蛋白可能是有用的干细胞标志物。我们发现，一种表面糖蛋白，富含亮氨酸重复神经元蛋白 1（LRRN1），在 hESC 分化为胚状体（EB）之前大量表达在表面。用短发夹 RNA（shLRRN1）沉默 hESC 中的 LRRN1 会导致自我更新能力下降，并且体外和体内偏向内胚层/中胚层谱系分化。同时，多能性因子 OCT4、NANOG 和 SOX2 的蛋白表达水平降低。有趣的是，LRRN1 沉默细胞中这些多能性因子的 mRNA 水平不受影响，但蛋白半衰期大大缩短。此外，我们发现 LRRN1 沉默导致所有三种多能性因子的核输出和蛋白酶体降解。此外，核输出的影响是由 AKT 磷酸化介导的。这些结果表明，LRRN1 通过 AKT 磷酸化在维持多能性因子的蛋白稳定性方面发挥重要作用，从而维持 hESC 的自我更新能力和多能性。总的来说，我们发现 LRRN1 通过防止 OCT4、NANOG 和 SOX2 从细胞核易位到细胞质，从而减少它们的翻译后修饰和降解，从而对 hESC 的多能性做出贡献。干细胞 2018;36:1514-1524。
