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FBW7 inhibits myeloid differentiation in acute myeloid leukemia via GSK3-dependent ubiquitination of PU.1
Molecular Cancer Research ( IF 5.2 ) Pub Date : 2020-11-13 , DOI: 10.1158/1541-7786.mcr-20-0268
Mukul Mishra 1 , Gatha Thacker 1 , Akshay Sharma 1 , Anil Kumar Singh 1 , Vishal Upadhyay 1 , Sabyasachi Sanyal 2, 3 , Shailendra Prasad Verma 4 , Anil Kumar Tripathi 4, 5 , Madan Lal Brahma Bhatt 4 , Arun Kumar Trivedi 1, 3
Affiliation  

GSK3β, an ubiquitously expressed serine/threonine kinase is reported to be overexpressed and hyperactivated in cancers including Acute Myeloid Leukemia where it promotes self-renewal, growth and survival of AML cells. Therefore, GSK3β inhibition results in AML cell growth inhibition and myeloid differentiation. Here we identified master transcription factor PU.1 of monocyte-macrophage differentiation pathway as potential GSK3β target. We demonstrate that GSK3β phosphorylates PU.1 at Ser41 and Ser140 leading to its recognition and subsequent ubiquitin-mediated degradation by E3 ubiquitin ligase FBW7. This GSK3-dependent degradation of PU.1 by FBW7 inhibited monocyte-macrophage differentiation. We further showed that a phospho-deficient PU.1 mutant (PU.1-S41,S140A) neither bound to FBW7 nor was degraded by it. Consequently, PU.1-S41,S140A retained its transactivation, DNA binding ability and promoted monocyte-macrophage differentiation of U937 cells even without PMA treatment. We further showed that FBW7 overexpression inhibited both PMA as well as MCSF-induced macrophage differentiation of myeloid cell lines and PBMCs from healthy volunteers respectively. Contrarily, FBW7 depletion promoted differentiation of these cells even without any inducer suggesting for a robust role of GSK3β-FBW7 axis in negatively regulating myeloid differentiation. Furthermore, we also recapitulated these findings in PBMCs isolated from leukemia patients where FBW7 over expression markedly inhibited endogenous PU.1 protein levels. In addition, PBMCs also showed enhanced differentiation when treated with M-CSF and GSK3 inhibitor (SB216763) together compared to M-CSF treatment alone. Implications: Our data demonstrate a plausible mechanism behind PU.1 restoration and induction of myeloid differentiation upon GSK3β inhibition and further substantiates potential of GSK3β as a therapeutic target in AML.

中文翻译:

FBW7 通过 GSK3 依赖的 PU 泛素化抑制急性髓系白血病的髓系分化。1

据报道,GSK3β 是一种普遍表达的丝氨酸/苏氨酸激酶,在包括急性髓性白血病在内的癌症中过度表达和过度活化,在这种情况下,它促进了 AML 细胞的自我更新、生长和存活。因此,GSK3β 抑制导致 AML 细胞生长抑制和骨髓分化。在这里,我们将单核细胞-巨噬细胞分化途径的主转录因子 PU.1 鉴定为潜在的 GSK3β 靶标。我们证明 GSK3β 在 Ser41 和 Ser140 处磷酸化 PU.1,导致其被 E3 泛素连接酶 FBW7 识别和随后泛素介导的降解。FBW7 对 PU.1 的这种 GSK3 依赖性降解抑制了单核细胞-巨噬细胞分化。我们进一步表明,缺乏磷的 PU.1 突变体(PU.1-S41,S140A)既不与 FBW7 结合,也不被它降解。因此,PU.1-S41,即使没有 PMA 处理,S140A 仍保留其反式激活、DNA 结合能力并促进 U937 细胞的单核细胞-巨噬细胞分化。我们进一步表明,FBW7 过表达分别抑制了来自健康志愿者的骨髓细胞系和 PBMC 的 PMA 和 MCSF 诱导的巨噬细胞分化。相反,即使没有任何诱导剂,FBW7 耗竭也促进了这些细胞的分化,这表明 GSK3β-FBW7 轴在负调节骨髓分化中的强大作用。此外,我们还在从白血病患者分离的 PBMC 中概括了这些发现,其中 FBW7 过表达显着抑制了内源性 PU.1 蛋白水平。此外,与单独的 M-CSF 治疗相比,当用 M-CSF 和 GSK3 抑制剂 (SB216763) 一起治疗时,PBMC 也显示出增强的分化。含义:
更新日期:2020-11-13
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