The N6-methyladenosine (m6A)-forming enzyme METTL3 controls myeloid differentiation of normal hematopoietic and leukemia cells

Abstract

N6-methyladenosine (m6A) is an abundant nucleotide modification in mRNA that is required for the differentiation of mouse embryonic stem cells. However, it remains unknown whether the m6A modification controls the differentiation of normal and/or malignant myeloid hematopoietic cells. Here we show that shRNA-mediated depletion of the m6A-forming enzyme METTL3 in human hematopoietic stem/progenitor cells (HSPCs) promotes cell differentiation, coupled with reduced cell proliferation. Conversely, overexpression of wild-type METTL3, but not of a catalytically inactive form of METTL3, inhibits cell differentiation and increases cell growth. METTL3 mRNA and protein are expressed more abundantly in acute myeloid leukemia (AML) cells than in healthy HSPCs or other types of tumor cells. Furthermore, METTL3 depletion in human myeloid leukemia cell lines induces cell differentiation and apoptosis and delays leukemia progression in recipient mice in vivo. Single-nucleotide-resolution mapping of m6A coupled with ribosome profiling reveals that m6A promotes the translation of c-MYC, BCL2 and PTEN mRNAs in the human acute myeloid leukemia MOLM-13 cell line. Moreover, loss of METTL3 leads to increased levels of phosphorylated AKT, which contributes to the differentiation-promoting effects of METTL3 depletion. Overall, these results provide a rationale for the therapeutic targeting of METTL3 in myeloid leukemia.

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Figure 1: m6A inhibits myeloid differentiation of HSPCs.
Figure 2: m6A promotes leukemogenesis.
Figure 3: m6A is required for maintaining the translation of target mRNAs that control cell fate.
Figure 4: m6A directly controls expression of c-MYC, BCL-2 and PTEN.

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Acknowledgements

We thank D. Bachovchin (MSKCC, New York) for the MOLM-13 constitutively expressing Cas9 cell line, and C. Vakoc (Cold Spring Harbor Laboratory, New York) for the constitutively expressing Cas9-RN2c cell line. We would like to thank the members of the Scaltriti laboratory (MSKCC, New York) for providing us with PI3K/AKT inhibitors. We thank the Weill Cornell Medicine Epigenomics Core for their assistance with sequencing. M.G.K. was supported by the US National Institutes of Health National Institute of Diabetes Digestive and Kidney Diseases Career Development Award, NIDDK NIH R01-DK101989-01A1, NCI 1R01CA193842-01, Kimmel Scholar Award, V-Scholar Award, Geoffrey Beene Award, Leukemia Lymphoma Society Career Development Award and Alex's Lemonade Stand A Award. This work was also supported by a Tri-Institutional Stem Cell Award (M.G.K. and S.R.J.), R01CA186702 (S.R.J.), T32CA062948 (B.F.P.), Ruth L. Kirschstein National Research Service Award 1F32CA22104-01 (B.F.P.), a Damon Runyon-Sohn Pediatric Cancer Fellowship Award DRSG10-14 (L.P.V.), and the American-Italian Cancer Foundation (S.Z.). The research was funded in part through the NIH/NCI Cancer Support Core Grant P30 CA08748 MGK. The RPPA core facility is funded by NCI #CA16672.

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L.P.V. led the project, performed experiments, analyzed data and wrote the manuscript. B.F.P., Y.C., D.N. and S.Z. performed experiments, analyzed data and edited the manuscript. G.M., T.C. and A.C. provided experimental supports. C.E.M., Y.S. and M.M. performed and analyzed MeRIP-seq experiments on patient-derived samples. J.S., C.F., M.P., F.E.G.-B., A.M., V.M.K. and M.C. provided patient samples. S.R.J. supervised the project and wrote the manuscript. M.G.K. directed the project, analyzed data and wrote the manuscript.

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Correspondence to Samie R Jaffrey or Michael G Kharas.

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Vu, L., Pickering, B., Cheng, Y. et al. The N6-methyladenosine (m6A)-forming enzyme METTL3 controls myeloid differentiation of normal hematopoietic and leukemia cells. Nat Med 23, 1369–1376 (2017). https://doi.org/10.1038/nm.4416

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