Background

Polycomb group (PcG) proteins are histone modifiers which control gene expression by assembling into large repressive complexes termed - Polycomb repressive complex (PRC); RING1B, core catalytic subunit of PRC1 that performs H2AK119 monoubiquitination leading to gene repression. The role of PRC1 complex during early neural specification in humans is unclear; we have tried to uncover the role of PRC1 in neuronal differentiation using human pluripotent stem cells as an in vitro model.

Results

We differentiated both human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) towards neural progenitor stage evident from the expression of NESTIN, TUJ1, NCAD, and PAX6. When we checked the total expression of RING1B and BMI1, we saw that they were significantly upregulated in differentiated neural progenitors compared to undifferentiated cells. Further, we used Chromatin Immunoprecipitation coupled with qPCR to determine the localization of RING1B, and the repressive histone modification H2AK119ub1 at the promoters of neuronal specific genes. We observed that RING1B localized to and catalyzed H2AK119ub1 modification at promoters of TUJ1, NCAM, and NESTIN during early differentiation and later RING1B was lost from its promoter leading their expression; while functional RING1B persisted significantly on mature neuronal genes such as IRX3, GSX2, SOX1, NEUROD1 and FOXG1 in neural progenitors.

Conclusion

The results of our study show that PRC1 catalytic component RING1B occupies neuronal gene promoters in human pluripotent stem cells and may prevent their precocious expression. However, when neuronal inductive signals are given, RING1B is not only removed from neuronal gene promoters, but the inhibitory H2AK119ub1 modification is also lost.

中文翻译：

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PRC1催化单元RING1B调节人多能干细胞的早期神经分化

背景

Polycomb group（PcG）蛋白是组蛋白修饰剂，可通过组装为大型抑制复合物来控制基因表达，这种复合物称为-Polycomb Repressed Complex（PRC）；RING1B，PRC1的核心催化亚基，执行H2AK119单泛素化，导致基因阻遏。在人类早期神经规范化过程中，PRC1复合物的作用尚不清楚。我们尝试使用人类多能干细胞作为体外模型来揭示PRC1在神经元分化中的作用。

结果

我们从NESTIN，TUJ1，NCAD和PAX6的表达来看，将人类胚胎干细胞（hESCs）和人类诱导的多能干细胞（hiPSCs）分化为神经祖细胞阶段。当我们检查RING1B和BMI1的总表达时，我们发现与未分化的细胞相比，它们在分化的神经祖细胞中明显上调。此外，我们将染色质免疫沉淀与qPCR结合使用来确定RING1B的定位以及神经元特异性基因启动子处的阻抑组蛋白修饰H2AK119ub1。我们观察到，RING1B定位并催化TUJ1，NCAM和NESTIN启动子处的H2AK119ub1修饰在早期分化过程中，后来RING1B从其启动子中丢失，导致其表达。而功能性RING1B则在神经祖细胞中的成熟神经元基因（例如IRX3，GSX2，SOX1，NEUROD1和FOXG1）上显着持久。

结论

我们的研究结果表明，PRC1催化成分RING1B在人多能干细胞中占据神经元基因启动子，并可能阻止其早熟表达。但是，当给出神经元诱导信号时，不仅从神经元基因启动子中去除了RING1B，而且抑制性H2AK119ub1修饰也丢失了。