Abstract:
One of the key areas in stem cell research is the identification of factors capable of promoting the expansion of Neural Stem Cell/Progenitor Cells (NSPCs) and understanding their molecular mechanisms for future use in clinical settings. We previously identified Macrophage Migration Inhibitory Factor (MIF) as a novel factor that can support the proliferation and/or survival of NSPCs based on in vitro functional cloning strategy and revealed that MIF can support the proliferation of human brain tumor-initiating cells (BTICs). However, the detailed downstream signaling for the functions has largely remained unknown. Thus, in the present study, we newly identified translationally-controlled tumor protein-1 (TPT1), which is expressed in the ventricular zone of mouse embryonic brain, as a downstream target of MIF signaling in mouse and human NSPCs and human BTICs. Using gene manipulation (over or downregulation of TPT1) techniques including CRISPR/Cas9-mediated heterozygous gene disruption showed that TPT1 contributed to the regulation of cell proliferation/survival in mouse NSPCs, human embryonic stem cell (hESC) derived-NSPCs, human-induced pluripotent stem cells (hiPSCs) derived-NSPCs and BTICs. Furthermore, gene silencing of TPT1 caused defects in neuronal differentiation in the NSPCs in vitro. We also identified the MIF-CHD7-TPT1-SMO signaling axis in regulating hESC-NSPCs and BTICs proliferation. Intriguingly, TPT1suppressed the miR-338 gene, which targets SMO in hESC-NSPCs and BTICs. Finally, mice with implanted BTICs infected with lentivirus-TPT1 shRNA showed a longer overall survival than control. These results also open up new avenues for the development of glioma therapies based on the TPT1 signaling pathway.
摘要:
干细胞研究的关键领域之一是鉴定能够促进神经干细胞/祖细胞(NSPC)扩增的因子,并了解其分子机制,以便将来在临床环境中使用。我们先前基于体外功能克隆策略将巨噬细胞迁移抑制因子(MIF)确定为可以支持NSPCs增殖和/或存活的新因子,并揭示MIF可以支持人脑肿瘤起始细胞(BTIC)的增殖。然而,功能的详细下游信令在很大程度上仍然未知。因此,在本研究中,我们新发现了翻译控制的肿瘤蛋白-1(TPT1),在小鼠胚胎大脑的脑室区表达,作为小鼠和人NSPC和人BTIC中MIF信号的下游靶标。使用基因操作(TPT1的过度或下调)技术,包括CRISPR/Cas9介导的杂合基因破坏,表明TPT1有助于调节小鼠NSPC的细胞增殖/存活,人胚胎干细胞(hESC)衍生的NSPCs,人诱导多能干细胞(hiPSC)衍生的NSPC和BTIC。此外,TPT1基因沉默导致体外NSPCs神经元分化缺陷。我们还鉴定了MIF-CHD7-TPT1-SMO信号轴在调节hESC-NSPC和BTIC增殖中的作用。有趣的是,TPT1抑制了miR-338基因,以hESC-NSPC和BTIC中的SMO为目标。最后,植入感染慢病毒-TPT1shRNA的BTIC的小鼠显示出比对照更长的总生存期。这些结果也为开发基于TPT1信号通路的神经胶质瘤治疗开辟了新的途径。