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Acute myeloid leukemia

Opposing effects of NPM1wt and NPM1c mutants on AKT signaling in AML

Leukemia volume 34, pages 1172–1176 (2020)Cite this article

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Nucleophosmin (NPM1), a ubiquitously expressed phosphoprotein that shuttles between the nucleus and cytoplasm, is implicated in multiple cellular processes including ribosomal biogenesis, cell cycle regulation, centrosome duplication and maintenance of genome stability [1]. NPM1 regulates the TP53-MDM2-ARF axis through its physical interaction with each of these proteins. NPM1 transcriptionally regulates TP53, and stabilizes TP53 protein through direct physical association [2] as well as through inhibition of MDM2 in the nucleolus, thereby protecting TP53 from MDM2-mediated ubiquitination [3]. Protein kinase B (PKB/AKT) has been shown to regulate NPM1-mediated ARF nucleolar localization and stability by phosphorylating NPM1 on serine 48 [4]. Moreover, AKT physically interacts with NPM1 and protects it from ubiquitination-mediated degradation [5]. Recurrent NPM1 mutations are found in approximately one-third of patients with acute myeloid leukemia (AML). These mutations create a frameshift that converts the carboxyl-terminal nucleolar localization signal of wild-type NPM1 (NPM1wt) into a nuclear export signal; the mutant proteins are referred to as NPM1c due to their cytoplasmic localization [6].

To address whether NPM1wt affects AKT signaling, we stably knocked down NPM1 in human embryonic kidney-derived HEK 293T cells using lentiviral shRNAs (shNPM1). NPM1 depletion led to a significant increase in AKT phosphorylation at S473 and T308 (Fig. 1c and Fig. S3). We also observed increased phosphorylation of AKT downstream targets including S6K at T389, and S6 at S235/236 and S240/244 (Figs. S3 and S4). Similar results were observed in the triple-negative breast cancer line MDA-MB-231 upon NPM1 knock down (Fig. S5). These results suggest that while AKT-NPM1 interaction stabilizes NPM1, this interaction also suppresses AKT phosphorylation and reduces its downstream signaling.

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Acknowledgements

We gratefully acknowledge the technical and administrative assistance from Jason Ho, Jian Liu, Youqi Han, Andrea Arruda, Narmin Ibrahimova, and Genna Luciani. We thank G. S. Vassiliou and P. P. Pandolfi for respectively providing Npm1loxp-cA/+ mice and NPM1c antibody. We thank Dr D. Herlyn of the Wistar Institute for the generous gift of lentiviral shNPM1 plasmids. We give thanks to Dr C. J. Sherr for the generous provision of the pcDNA3.1-FLAG-full length NPM1 and NPM1 deletion mutant constructs. The Minden lab is funded by grants from the Canadian Institute for Cancer Research, Ontario Institute for Cancer Research, the Orsino Chair in Leukemia Research, and the Wendy Eisen Foundation. The Stambolic lab and the Zacksenhaus lab are funded by grants from Canadian Cancer Society. We are very grateful to Muriel Matthews and Eric Mills for their help with the editing of the manuscript.

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Authors and Affiliations

  1. Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada

    Zhuo Ren, Tali Melkman, Li Meng, Tak W. Mak, Vuk Stambolic, Mark D. Minden & Jiance Atom Wang

  2. Toronto General Research Institute, University Health Network, Toronto, ON, Canada

    Mariusz Shrestha & Eldad Zacksenhaus

  3. Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada

    Mariusz Shrestha & Eldad Zacksenhaus

  4. The Campbell Family Institute for Breast Cancer Research at Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada

    Takashi Sakamoto, Rob A. Cairns & Tak W. Mak

  5. Department of Medicine and Medical Biophysics, University of Toronto, Toronto, ON, Canada

    Eldad Zacksenhaus, Tak W. Mak, Vuk Stambolic, Mark D. Minden & Jiance Atom Wang

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  1. Zhuo Ren
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Contributions

ZR, MDM, EZ, MS, TWM, RAC and TS developed the concepts and designed the experiments. ZR, MS, TS and JAW performed the experiments. ZR, MS, TS, JAW, TM, LM, RAC, EZ, MDM and VS analyzed data. ZR, MS, TS, EZ, MDM and VS wrote the manuscript.

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Correspondence to Mark D. Minden.

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Ren, Z., Shrestha, M., Sakamoto, T. et al. Opposing effects of NPM1wt and NPM1c mutants on AKT signaling in AML. Leukemia 34, 1172–1176 (2020). https://doi.org/10.1038/s41375-019-0621-7

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