Musashi2 sustains the mixed-lineage leukemia–driven stem cell regulatory program
Sun-Mi Park, Mithat Gönen, Ly Vu, Gerard Minuesa, Patrick Tivnan, Trevor S. Barlowe, James Taggart, Yuheng Lu, Raquel P. Deering, Nir Hacohen, Maria E. Figueroa, Elisabeth Paietta, Hugo F. Fernandez, Martin S. Tallman, Ari Melnick, Ross Levine, Christina Leslie, Christopher J. Lengner, Michael G. Kharas
Sun-Mi Park, Mithat Gönen, Ly Vu, Gerard Minuesa, Patrick Tivnan, Trevor S. Barlowe, James Taggart, Yuheng Lu, Raquel P. Deering, Nir Hacohen, Maria E. Figueroa, Elisabeth Paietta, Hugo F. Fernandez, Martin S. Tallman, Ari Melnick, Ross Levine, Christina Leslie, Christopher J. Lengner, Michael G. Kharas
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Research Article Oncology

Musashi2 sustains the mixed-lineage leukemia–driven stem cell regulatory program

Abstract

Leukemia stem cells (LSCs) are found in most aggressive myeloid diseases and contribute to therapeutic resistance. Leukemia cells exhibit a dysregulated developmental program as the result of genetic and epigenetic alterations. Overexpression of the RNA-binding protein Musashi2 (MSI2) has been previously shown to predict poor survival in leukemia. Here, we demonstrated that conditional deletion of Msi2 in the hematopoietic compartment results in delayed leukemogenesis, reduced disease burden, and a loss of LSC function in a murine leukemia model. Gene expression profiling of these Msi2-deficient animals revealed a loss of the hematopoietic/leukemic stem cell self-renewal program and an increase in the differentiation program. In acute myeloid leukemia patients, the presence of a gene signature that was similar to that observed in Msi2-deficent murine LSCs correlated with improved survival. We determined that MSI2 directly maintains the mixed-lineage leukemia (MLL) self-renewal program by interacting with and retaining efficient translation of Hoxa9, Myc, and Ikzf2 mRNAs. Moreover, depletion of MLL target Ikzf2 in LSCs reduced colony formation, decreased proliferation, and increased apoptosis. Our data provide evidence that MSI2 controls efficient translation of the oncogenic LSC self-renewal program and suggest MSI2 as a potential therapeutic target for myeloid leukemia.

Authors

Sun-Mi Park, Mithat Gönen, Ly Vu, Gerard Minuesa, Patrick Tivnan, Trevor S. Barlowe, James Taggart, Yuheng Lu, Raquel P. Deering, Nir Hacohen, Maria E. Figueroa, Elisabeth Paietta, Hugo F. Fernandez, Martin S. Tallman, Ari Melnick, Ross Levine, Christina Leslie, Christopher J. Lengner, Michael G. Kharas

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Figure 5

MSI2 binds and regulates the expression of Hoxa9, c-Myc, and Ikzf2.

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MSI2 binds and regulates the expression of Hoxa9, c-Myc, and Ikzf2. (A) ...
(A) Cumulative distribution function (CDF) showing mRNA fold enrichment of HITS-CLIP targets in the RNA sequencing data of Msi2fl/fl and Msi2Δ/Δ LSCs, Kolmogorov–Smirnov test. (B) Functional pathway overlap analysis from HITS-CLIP and RNA sequencing data from Msi2fl/fl and Msi2Δ/Δ LSCs and microarray data from Msi2Δ/Δ LSKs. MLL-related gene sets shared among LSKs and LSCs (Supplemental Table 6). (C) RNA immunoprecipitation of MSI2 showing binding to selected MLL targets in Msi2fl/fl (n = 4–7 independent RNA-IPs and clones), Msi2Δ/Δ (n = 3 clones), and MSI2O/E (n = 4 clones; MLL-AF9 leukemia cells derived from LSK transformed cells from MSI2 doxycycline transgenic mice and induced for 2 days with doxycycline) (fold change calculated over IgG for each gene SEM). Means and SEM, *P < 0.05, **P < 0.01, unpaired Student’s t test. Hebp2 was not assessed. (D) Quantitative PCR for selected MLL target genes in control and MSI2O/E leukemic cells at 48 hours of MSI2 induction. Data are the mean of 3 independent experiments. Control, n = 3; MSI2O/E, n = 6. Means and SEM, *P < 0.05, unpaired Student’s t test. (E) Immunoblot analysis for MLL targets and MSI2 in control and MSI2O/E leukemic cells, from 2 independent experiments. (F) Quantitative PCR after acute deletion of Msi2 at 68 hours after 4-hydroxytestosterone (4-OHT) treatment in Msi2fl/fl Cre-ER– and Msi2fl/fl Cre-ER+ LSCs (n = 3 Msi2fl/fl Cre– and n = 4 Msi2fl/fl Cre+), average and SEM. (G) Immunoblot analysis for MLL targets and deletion of Msi2 at 68 hours after 4-OHT treatment in Msi2fl/fl Cre-ER– and Msi2fl/fl Cre-ER+ leukemic cells, representative of 2 independent experiments.