EpoR & Stat5 regulation of ribosome biogenesis and protein synthesis in erythropoiesis

项目摘要蛋白质合成和核糖体生物合成是最耗能的两种 在一个细胞中的过程，导致它们通过严格的控制来调节。EPOR，细胞因子受体的成员 超家族及其下游调节因子Stat5对红细胞生成起着至关重要的作用。目前的提案 通过EPOR和Stat5信号对核糖体生物发生和蛋白质合成的新调节 红系发展的关键决策。在最近发表的研究中，我们发现EPOR/Stat5刺激的时间更短 和更多的早期红细胞周期，同时也促进更大的红细胞的形成， 在老鼠和人类身上，都会成熟为更大的红细胞。这一令人惊讶的发现表明，EPOR/Stat5 信号改变了细胞周期持续时间和细胞大小之间的关系，同时导致较短的 生物量的循环和异常快速的增长。在支持这一假说的初步数据中，单细胞RNA EPOR-/-和Stat5-/-胎儿肝脏测序显示核糖体生物发生和 翻译基因。此外，我们发现了依赖于EPOR/Stat5的rRNA速率的急剧尖峰 转录(~2.5倍)、蛋白质合成率(~4至6倍)和细胞大小的生长率(~3倍)， 与关键细胞命运的决定不谋而合。它发生在早期的红系祖细胞，即cfu-e转变。 从自我更新到红系终末分化(ETD)，成为红细胞。蛋白质含量的激增 此时的合成也与异常短的细胞周期相吻合。紧随CFU-e/ETD之后 过渡期，核糖体生物合成和蛋白质合成的速率开始一起下降到基线水平 随着细胞周期速度的下降，即使红细胞继续分裂和合成血红蛋白 以获得额外的3到5个细胞分裂。蛋白质合成峰值与核糖体的重合 CFU-e/ETD开关上细胞周期缩短的生物发生表明，这些过程是联系在一起的 可能在功能上与交换机相关。我们将研究EPOR/Stat5诱导的蛋白质尖峰 目的1：研究EPOR/Stat5在核糖体生物发生中诱导的尖峰信号。 蛋白质合成，确定调节这一峰值的细胞内信号通路，并识别 翻译率激增的潜在抄本子集。目标2：确定两者之间的因果关系 细胞周期、核糖体生物发生、蛋白质合成和细胞大小。细胞间的调控相互作用 哺乳动物细胞的周期持续时间、蛋白质合成率和细胞大小还不是很清楚 在发育和癌症中起关键作用。目的3：验证EPOR/Stat5诱导蛋白质尖峰的假设 合成是红系分化所必需的。我们将确定EPOR诱导的蛋白质峰值是否 合成是核糖体蛋白Rpl11缺陷小鼠的致命弱点，Rpl11是钻石黑粉的模型 贫血，潜在地解释了红系对核糖病的选择性敏感性。

Project Summary Protein synthesis and ribosome biogenesis are the two most energy -intensive processes in a cell, resulting in their regulation through tight controls. EpoR, a member of the cytokine receptor superfamily, and its downstream mediator Stat5, are essential for erythropoiesis. The current proposal addresses novel regulation of ribosome biogenesis and protein synthesis by EpoR and Stat5 signaling during a key erythroid developmental decision. In recently published work we found that EpoR/ Stat5 stimulate shorter and more numerous cycles in early erythroblasts, while also promoting formation of larger erythroblasts that mature into larger red cells, in both mice and humans. This surprising finding suggests that EpoR/ Stat5 signaling alters the relationship between cell cycle duration and cell size, simultaneously inducing shorter cycles and exceptionally fast growth in biomass. In preliminary data supporting this hypothesis, single-cell RNA sequencing of Epor-/- and Stat5-/- fetal livers show dysregulated expression of ribosome biogenesis and translation genes. Further, we identified an EpoR/ Stat5-dependent sharp spike in the rate of rRNA transcription (~2.5 fold), protein synthesis rate (~4 to 6 fold) and rate of growth in cell size (~3 fold), that coincides with a key cell fate decision. It takes place as early erythroid progenitors known as CFU-e transition from self-renewal to erythroid terminal differentiation (ETD), becoming erythroblasts. The spike in protein synthesis at this time also coincides with an unusually short cell cycle. Immediately following the CFU-e/ETD transition, the rates of ribosome biogenesis and protein synthesis begin to decline back to baseline together with the decline in cell cycle speed, even though erythroblasts continue to divide and synthesize hemoglobin for an additional 3 to 5 cell divisions. The coincidence in the spikes of protein synthesis and ribosome biogenesis with cell cycle shortening at the CFU-e/ETD switch suggests that these processes are linked and may be functionally relevant to the switch. We will investigate the EpoR/Stat5- induced spike in protein synthesis with the following aims: Aim 1: Investigate the EpoR /Stat5 -induced spike in ribosome biogenesis & protein synthesis, determining the intracellular signaling pathways that are mediating this spike and identifying a potential subset of transcripts whose translation rate spikes. Aim 2: Determine causal relationships between the cell cycle, ribosome biogenesis, protein synthesis and cell size. The regulatory interactions between cell cycle duration, protein synthesis rate and cell size in mammalian cells are not well understood and yet are critical in development and in cancer. Aim 3: Test the hypothesis that the EpoR /Stat5 -induced spike in protein synthesis is required for erythroid differentiation. We will determine whether the EpoR-induced spike in protein synthesis is an 'Achilles heel' in mice deficient in the ribosomal protein Rpl11, a model of Diamond Blackfan Anemia, potentially explaining the selective sensitivity of the erythroid lineage to ribosomopathies.