Uncovering the HOXA9 Translational Regulatory Complex That Promotes AML Leukemic Stem Cells
Bibliographic record
Abstract
HOXA9 is a master transcription factor of hematopoiesis and essential for maintaining self-renewal of leukemia stem cells (LSCs). Although mechanisms associated with its transcriptional regulation are extensively studied, how Hoxa9 mRNA translation is controlled remain poorly understood. We have previously identified the RNA binding protein (RBP) MUSASHI-2 (MSI2) as a central regulator of myeloid LSCs by controlling translation of crucial transcription factors including Hoxa9 (Park et al. JCI, 2015; Nguyen et al. Nature Comm, 2020). However, the mechanisms by which HOXA9 translation is modulated and the role of MSI2 remain unknown. To characterize the HOXA9 3'UTR mRNA regulon, we used it as a bait to perform an RNA-pull down followed by mass spectrometry (MS) with control or MSI2 depleted cells. To determine the direct MSI2's protein interactors on HOXA9, we also performed MSI2 co-immunoprecipitation (co-IP) MS in control or MSI2 deficient AML cells. We identified 10 RBPs that directly bind to MSI2 and are dependent on MSI2 for their binding to the HOXA9-3'UTR mRNA. Among these, we focused on SYNCRIP, a known MSI2 interactor (Vu et al. Nat Gen, 2017), HNRNPC and MOV10, of which binding ability on HOXA9 3'UTR was most reduced and enhanced, respectively, upon MSI2 loss. Reciprocal co-IP confirmed the interactions between these RBPs with MSI2. We then reasoned that if these RBPs are in the same regulatory complex with MSI2, they might phenocopy MSI2 loss of function in LSCs. To understand how each selected factor impacts LSC function and gene control, we first assessed MSI2's global regulation by performing integrative omic studies including MSI2-HyperTRIBE, RNA-seq and proteomics in MLL-AF9 murine LSCs that have acutely deleted (68hrs) of Msi2. We found that MSI2 binding promotes translation of 264 genes while suppresses 8 genes; and only 10 genes affected at mRNA level. These data found that MSI2 mediates translation of the self-renewal program in LSCs. We then found that SYNCRIP is required for both leukemia initiation and maintenance in the MLL-AF9 driven AML model with the Syncrip conditional knockout mice. Serial transplantation assays of Syncrip deleted cells show a continued delay in leukemogenesis in vivo, suggesting SYNCRIP is required for functional LSC activity. To identify SYNCRIP's targets, we performed SYNCRIP-HyperTRIBE and iCLIP, and found that majority (~64%) of SYNCRIP targets overlapped with MSI2 targets and their binding sites are proximity in LSCs. SYNCRIP knockout transcriptome enriched for HOXA9-MEIS1 targets, and MSI2 deficient HSC signature. These data suggest that SYNCRIP is required for LSC function by co-regulating HOXA9-associated program with MSI2. Furthermore, depleting HNRNPC or MOV10 reduces AML cell proliferation, colony formation and increases apoptosis. Both HNRNPC and MOV10 depletion significantly decreased HOXA9 protein abundance. HNRNPC knockout cells exhibited delayed leukemia development in vivo and serial plating assays shows that this RBP is required for maintaining LSC' self-renewal. Importantly, higher HNRNPC or MOV10 expression associated with a worse overall survival in AML patients (p=0.0026 for HNRNPC; p=0.0101 for MOV10). ENRICHR analysis shows HNRNPC correlatedly expressed genes in human AML patients enriched for HOXA9, MOV10 and MSI2 signature. These data suggest that HNRNPC and MOV10 co-regulate leukemia HOXA9-associated program with MSI2. We also discovered that these RBPs collaborate in the translational regulation mediated by MSI2. SYNCRIP is necessary for MSI2 recruitment to the HOXA9 3'UTR based on reduced MSI2 binding after SYNCRIP depletion (MSI2-HYPERTRIBE and MSI2-RNA-IP). Luciferase reporter assays with HOXA9-3'UTR found that knockdown each one of the components in the MSI2-SYNCRIP-HNRNPC-MOV10 complex alone significantly decreased luciferase signal. Moreover, loss of each of the core components SYNCRIP and HNRNPC exacerbated translation inhibition caused by MSI2 depletion and attenuated the increase in translation resulted from MSI2 overexpression. Our data suggest that SYNCRIP and HNRNPC facilitate MSI2-mediated translation regulation of HOXA9. Overall, our study uncovered a MSI2-associated RBP complex co-regulating HOXA9 translation and LSC program in AML. We revealed that SYNCRIP is required for LSC function, and HNRNPC and MOV10 as novel vulnerabilities in AML.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.002 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".