Context-dependent modification of PFKFB3 in hematopoietic stem cells promotes anaerobic glycolysis and ensures stress hematopoiesis
Context-dependent modification of PFKFB3 in hematopoietic stem cells promotes anaerobic glycolysis and ensures stress hematopoiesis
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DOI：
10.1101/2023.03.16.532898
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发表时间：
2023-11
期刊：
影响因子：
7.7
通讯作者：
Shintaro Watanuki;Hiroshi Kobayashi;Yuki Sugiura;M. Yamamoto;Daiki Karigane;K. Shiroshita;Yuriko Sorimachi;Shinya Fujita;T. Morikawa;S. Koide;M. Oshima;Akira Nishiyama;Koichi Murakami;Miho Haraguchi;Shinpei Tamaki;Takehiro Yamamoto;Tomohiro Yabushita;Yosuke Tanaka;Go Nagamatsu;Hiroaki Honda;Shinichiro Okamoto;N. Goda;Tomohiko Tamura;A. Nakamura-Ishizu;M. Suematsu;Atsushi Iwama;Toshio Suda;K. Takubo
中科院分区：
文献类型：
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作者：
Shintaro Watanuki;Hiroshi Kobayashi;Yuki Sugiura;M. Yamamoto;Daiki Karigane;K. Shiroshita;Yuriko Sorimachi;Shinya Fujita;T. Morikawa;S. Koide;M. Oshima;Akira Nishiyama;Koichi Murakami;Miho Haraguchi;Shinpei Tamaki;Takehiro Yamamoto;Tomohiro Yabushita;Yosuke Tanaka;Go Nagamatsu;Hiroaki Honda;Shinichiro Okamoto;N. Goda;Tomohiko Tamura;A. Nakamura-Ishizu;M. Suematsu;Atsushi Iwama;Toshio Suda;K. Takubo
代谢途径是可塑性的，在应对压力或干扰时会迅速改变。目前的代谢谱技术需要裂解许多细胞，这使得跟踪稀有细胞(如造血干细胞)在应激后随时间的代谢变化变得复杂。在这里，我们的目标是确定定义在稳态和应激条件下HSCs糖酵解代谢差异的关键代谢酶，并阐明它们的调节机制。通过利用高灵敏度葡萄糖示踪和数学模型对葡萄糖代谢的~(13)C代谢通量进行定量分析，我们发现HSC在增殖和氧化磷酸化(OXPHOS)抑制过程中激活糖酵解限速酶磷酸果糖激酶(PFK)。实时测量单个HSC中的三磷酸腺苷(ATP)水平表明，增殖应激或OXPHOS抑制通过在几秒钟内增加PFKFB3的活性来加速糖酵解，PFKFB3是调节变构PFK激活剂的酶，以满足ATP的要求。此外，不同的应激通过依赖于PRMT1的甲基化和依赖于AMPK的磷酸化在OXPHOS抑制期间激活了PFKFB3。过表达PFKFB3可诱导HSC增殖并促进分化细胞的产生，而抑制或缺失PFKFB3则抑制其增殖和分化。本研究揭示了HSC糖酵解代谢在应激状态下维持造血的灵活和多层次调节，为更好地了解稀有造血细胞的生理代谢提供了技术支持。结合同位素示踪、数学建模和单细胞ATP分析的关键点可以实现对血细胞新陈代谢的高分辨率评估。在应激状态下，HSCs迅速加速糖酵解，以满足ATP的需求，并通过上下文依赖的PFKFB3激活来维持造血。
Metabolic pathways are plastic and rapidly change in response to stress or perturbation. Current metabolic profiling techniques require lysis of many cells, complicating the tracking of metabolic changes over time after stress in rare cells such as hematopoietic stem cells (HSCs). Here, we aimed to identify the key metabolic enzymes that define differences in glycolytic metabolism between steady-state and stress conditions in HSCs and elucidate their regulatory mechanisms. Through quantitative 13C metabolic flux analysis of glucose metabolism using high-sensitivity glucose tracing and mathematical modeling, we found that HSCs activate the glycolytic rate-limiting enzyme phosphofructokinase (PFK) during proliferation and oxidative phosphorylation (OXPHOS) inhibition. Real-time measurement of adenosine triphosphate (ATP) levels in single HSCs demonstrated that proliferative stress or OXPHOS inhibition led to accelerated glycolysis via increased activity of PFKFB3, the enzyme regulating an allosteric PFK activator, within seconds to meet ATP requirements. Furthermore, varying stresses differentially activated PFKFB3 via PRMT1-dependent methylation during proliferative stress and via AMPK-dependent phosphorylation during OXPHOS inhibition. Overexpression of Pfkfb3 induced HSC proliferation and promoted differentiated cell production, whereas inhibition or loss of Pfkfb3 suppressed them. This study reveals the flexible and multilayered regulation of HSC glycolytic metabolism to sustain hematopoiesis under stress and provides techniques to better understand the physiological metabolism of rare hematopoietic cells. Key Points Combined isotope tracing, mathematical modeling, and single cell ATP analysis enable high-resolution evaluation of blood cell metabolism. Under stress, HSCs quickly accelerate glycolysis to meet ATP demands and maintain hematopoiesis via context-dependent PFKFB3 activation.