Acute lymphoblastic leukemia (ALL) is a leading cause of cancer-related death in children and adolescents, and cure rates for relapsed/refractory ALL remain dismal, highlighting the need for novel targeted therapies. To identify genome-wide metabolic-stress regulated genes, we used RNA-sequencing in ALL cells treated with AICAR, an AMPK activator. RNA-sequencing identified the immediate early genes (IEGs) as a subset of genes downregulated by AICAR. We show that AICAR-induced IEGs downregulation was blocked by an adenosine uptake inhibitor indicating AICAR was responsible for IEGs reprogramming. Using pharmacologic and genetic models we established this mechanism was AMPK-independent. Further investigations using kinase assays, PKD/PKC inhibitors and rescue experiments, demonstrated that AICAR directly inhibited PKD kinase activity and identified PKD as responsible for IEGs downregulation. Mechanistically, PKD inhibition suppressed phosphorylation and nuclear export of class IIa HDACs, which lowered histone H3 acetylation and decreased NFκB(p65) recruitment to IEGs promoters. Finally, PKD inhibition induced apoptosis via DUSP1/DUSP6 downregulation eliciting a DNA damage response. More importantly, ALL patient cells exhibited the same PKD-HDACs-IEGs–mediated mechanism. As proof of principle of the therapeutic potential of targeting PKD, we established the in vivo relevance of our findings using an NSG ALL mouse model. In conclusion, we identified a previously unreported PKD-dependent survival mechanism in response to AICAR-induced cellular stress in ALL through regulation of DUSPs and IEGs' expression. Implications: PKD mediates early transcriptional responses in ALL cells as an adaptive survival mechanism to overcome cellular stress.

中文翻译：

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在急性淋巴细胞白血病中通过 IIA 类组蛋白脱乙酰酶对即刻早期基因的蛋白激酶 D 依赖性下调

急性淋巴细胞白血病 (ALL) 是儿童和青少年癌症相关死亡的主要原因，复发/难治性 ALL 的治愈率仍然很低，这凸显了对新型靶向治疗的需求。为了鉴定全基因组代谢应激调节基因，我们在用 AMPK 激活剂 AICAR 处理的所有细胞中使用 RNA 测序。RNA 测序将直接早期基因 (IEG) 鉴定为 AICAR 下调的基因子集。我们表明，AICAR 诱导的 IEG 下调被腺苷摄取抑制剂阻断，表明 AICAR 负责 IEG 重编程。使用药理学和遗传模型，我们建立了这种机制是 AMPK 独立的。使用激酶测定、PKD/PKC 抑制剂和救援实验进行进一步研究，证明 AICAR 直接抑制 PKD 激酶活性并确定 PKD 是导致 IEG 下调的原因。从机制上讲，PKD 抑制抑制了 IIa 类 HDAC 的磷酸化和核输出，从而降低了组蛋白 H3 乙酰化并减少了 NFκB(p65) 向 IEG 启动子的募集。最后，PKD 抑制通过 DUSP1/DUSP6 下调诱导细胞凋亡，从而引发 DNA 损伤反应。更重要的是，所有患者细胞都表现出相同的 PKD-HDACs-IEGs 介导的机制。作为靶向 PKD 治疗潜力的原理证明，我们使用 NSG ALL 小鼠模型建立了我们的研究结果的体内相关性。总之，我们通过调节 DUSP 和 IEG 的表达，确定了一种以前未报道的 PKD 依赖性生存机制，以响应 AICAR 诱导的 ALL 细胞应激。