Histone H2A.X Phosphorylation Activates Caspase-Induced Chromatin Condensation in Late Stage Erythropoiesis

Condensation of chromatin prior to enucleation is an essential component of terminal erythroid maturation, and defects in this process are associated with inefficient erythropoiesis and anemia. However, the mechanisms involved in this phenomenon are not well understood. Here, we identify a novel role for the histone variant H2A.X in erythropoiesis. We find in multiple model systems that this histone is essential for normal maturation and the loss of H2A.X in erythroid cells results in dysregulation in expression of erythroid specific genes as well a nuclear condensation defect. In Addition, we demonstrate that erythroid maturation is characterized by phosphorylation at both S139 and Y142 during late stage erythropoiesis. Knockout of the kinase BAZ1B/WSTF results in loss of Y142 phosphorylation and similar to loss of H2A.X, a defect in nuclear condensation. In this study, we present a model in which post-translational modifications on histone H2A.X during terminal erythroid maturation leads to Caspase-Induced Chromatin Condensation (CICC). In this novel pathway, although apoptosis is specifically suppressed, aspects of the apoptotic pathway remain active to drive terminal erythroid maturation. Suggesting that terminal erythroid maturation is indeed is a unique form of apoptosis. Overall design: RNA-Seq of H2A.X and BAZ1B WT and KO HUDEP-2 cells at D0 and D6 of differentiation.

去核前的染色质浓缩是终末红细胞成熟的核心环节，该过程的缺陷与低效红细胞生成及贫血密切相关。然而，目前学界对这一过程背后的分子机制尚缺乏充分认知。本研究揭示了组蛋白变体H2A.X（histone variant H2A.X）在红细胞生成中的全新功能。我们在多种模型系统中发现，该组蛋白对正常红细胞成熟至关重要；红细胞中H2A.X的缺失会导致红细胞特异性基因表达失调，同时引发染色质浓缩缺陷。此外，本研究证实，晚期红细胞生成阶段的红细胞成熟过程会伴随S139与Y142两位点的磷酸化修饰。敲除激酶BAZ1B/WSTF会导致Y142位点磷酸化的丢失，其表型与H2A.X缺失相似，同样引发染色质浓缩缺陷。本研究提出全新模型：终末红细胞成熟过程中，组蛋白H2A.X的翻译后修饰会介导半胱天冬酶依赖性染色质浓缩（Caspase-Induced Chromatin Condensation, CICC）。在该新型通路中，尽管细胞凋亡被特异性抑制，但凋亡通路的部分环节仍保持活性以驱动终末红细胞成熟，这表明终末红细胞成熟实为一种独特的细胞凋亡形式。实验整体设计：对分化第0天（D0）与第6天（D6）的H2A.X、BAZ1B野生型（wild type, WT）及敲除型（knockout, KO）HUDEP-2细胞进行RNA测序（RNA-Seq）。