Mutant NPM1 marginally impacts ribosome footprint in acute myeloid leukemia cells

EJHaem Pub Date : 2024-08-20 DOI:10.1002/jha2.996
Lorenzo Brunetti, Giulia Pianigiani, Michael C. Gundry, Margaret A. Goodell, Brunangelo Falini
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Abstract

Background

NPM1-mutated acute myeloid leukemia (AML) is the most frequent AML subtype. As wild-type NPM1 is known to orchestrate ribosome biogenesis, it has been hypothesized that altered translation may contribute to leukemogenesis and leukemia maintenance in NPM1-mutated AML. However, this hypothesis has never been investigated. We reasoned that if mutant NPM1 (NPM1c) directly impacts translation in leukemic cells, loss of NPM1c would result in acute changes in the ribosome footprint.

Methods

Here, we performed ribosome footprint profiling (Ribo-seq) and bulk messenger RNA (mRNA) sequencing in two NPM1-mutated cell lines engineered to express endogenous NPM1c fused to the FKBP (F36V) degron tag (degron cells).

Results and discussion

Incubation of degron cells with the small compound dTAG-13 enables highly specific degradation of NPM1c within 4 hours. As expected, RNA-sequencing data showed early loss of homeobox gene expression following NPM1c degradation, confirming the reliability of our model. In contrast, Ribo-seq data showed negligible changes in the ribosome footprint in both cell lines, implying that the presence of NPM1c does not influence ribosome abundance and positioning on mRNA. While it is predictable that NPM1c exerts its leukemogenic activity at multiple levels, ribosome footprint does not seem influenced by the presence of mutant NPM1.

Abstract Image

突变型 NPM1 对急性髓性白血病细胞中的核糖体足迹影响甚微
背景 NPM1 基因突变的急性髓性白血病(AML)是最常见的急性髓性白血病亚型。由于已知野生型 NPM1 能协调核糖体的生物生成,因此有人假设翻译的改变可能会导致 NPM1 突变的急性髓性白血病的白血病生成和白血病维持。然而,这一假设从未得到过研究。我们推断,如果突变型 NPM1(NPM1c)直接影响白血病细胞的翻译,那么 NPM1c 的缺失将导致核糖体足迹的急剧变化。 方法 在这里,我们在两个NPM1突变细胞系中进行了核糖体足迹分析(Ribo-seq)和大量信使RNA(mRNA)测序,这两个细胞系被设计为表达融合了FKBP(F36V)degron标签的内源性NPM1c(degron细胞)。 结果与讨论 用小化合物 dTAG-13 培养 degron 细胞,可在 4 小时内高度特异性地降解 NPM1c。正如预期的那样,RNA 序列数据显示 NPM1c 降解后同源染色体基因表达的早期损失,证实了我们模型的可靠性。与此相反,Ribo-seq 数据显示两种细胞系中核糖体足迹的变化可以忽略不计,这意味着 NPM1c 的存在不会影响核糖体的丰度和在 mRNA 上的定位。虽然可以预测 NPM1c 在多个水平上发挥其致白血病活性,但核糖体足迹似乎并不受突变 NPM1 存在的影响。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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