Department of Pediatrics, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
PLoS Biol. 2012 Aug;10(8):e1001383. doi: 10.1371/journal.pbio.1001383. Epub 2012 Aug 28.
Erythropoietin (Epo)-induced Stat5 phosphorylation (p-Stat5) is essential for both basal erythropoiesis and for its acceleration during hypoxic stress. A key challenge lies in understanding how Stat5 signaling elicits distinct functions during basal and stress erythropoiesis. Here we asked whether these distinct functions might be specified by the dynamic behavior of the Stat5 signal. We used flow cytometry to analyze Stat5 phosphorylation dynamics in primary erythropoietic tissue in vivo and in vitro, identifying two signaling modalities. In later (basophilic) erythroblasts, Epo stimulation triggers a low intensity but decisive, binary (digital) p-Stat5 signal. In early erythroblasts the binary signal is superseded by a high-intensity graded (analog) p-Stat5 response. We elucidated the biological functions of binary and graded Stat5 signaling using the EpoR-HM mice, which express a "knocked-in" EpoR mutant lacking cytoplasmic phosphotyrosines. Strikingly, EpoR-HM mice are restricted to the binary signaling mode, which rescues these mice from fatal perinatal anemia by promoting binary survival decisions in erythroblasts. However, the absence of the graded p-Stat5 response in the EpoR-HM mice prevents them from accelerating red cell production in response to stress, including a failure to upregulate the transferrin receptor, which we show is a novel stress target. We found that Stat5 protein levels decline with erythroblast differentiation, governing the transition from high-intensity graded signaling in early erythroblasts to low-intensity binary signaling in later erythroblasts. Thus, using exogenous Stat5, we converted later erythroblasts into high-intensity graded signal transducers capable of eliciting a downstream stress response. Unlike the Stat5 protein, EpoR expression in erythroblasts does not limit the Stat5 signaling response, a non-Michaelian paradigm with therapeutic implications in myeloproliferative disease. Our findings show how the binary and graded modalities combine to generate high-fidelity Stat5 signaling over the entire basal and stress Epo range. They suggest that dynamic behavior may encode information during STAT signal transduction.
促红细胞生成素 (Epo) 诱导的 Stat5 磷酸化 (p-Stat5) 对于基础红细胞生成和缺氧应激期间的加速都至关重要。一个关键的挑战在于理解 Stat5 信号如何在基础和应激红细胞生成中引发不同的功能。在这里，我们想知道这些不同的功能是否可以通过 Stat5 信号的动态行为来指定。我们使用流式细胞术分析了体内和体外原代红细胞生成组织中的 Stat5 磷酸化动力学，确定了两种信号模式。在后期（碱性）红细胞中，Epo 刺激引发低强度但决定性的二元（数字）p-Stat5 信号。在早期红细胞中，二元信号被高强度分级（模拟）p-Stat5 反应所取代。我们使用表达缺乏细胞质磷酸酪氨酸的“敲入”EpoR 突变体的 EpoR-HM 小鼠阐明了二元和分级 Stat5 信号的生物学功能。引人注目的是，EpoR-HM 小鼠仅限于二元信号模式，该模式通过促进红细胞中的二元存活决策来挽救这些小鼠免于致命的围产期贫血。然而，EpoR-HM 小鼠中缺少分级 p-Stat5 反应阻止它们对压力做出反应，包括上调转铁蛋白受体的能力，我们证明这是一种新的应激靶标。我们发现 Stat5 蛋白水平随着红细胞分化而下降，控制了从早期红细胞中的高强度分级信号向后期红细胞中的低强度二元信号的转变。因此，我们使用外源性 Stat5 将后期红细胞转化为能够引发下游应激反应的高强度分级信号转导器。与 Stat5 蛋白不同，EpoR 在红细胞中的表达并不限制 Stat5 信号反应，这是一种非迈克尔型范例，在骨髓增生性疾病中具有治疗意义。我们的研究结果表明，二元和分级模式如何结合在整个基础和应激 Epo 范围内生成高保真 Stat5 信号。它们表明，动态行为可能在 STAT 信号转导过程中编码信息。
