Loss of FBXO11 Function Establishes a Stem Cell Program in Acute Myeloid Leukemia through Dysregulation of the Mitochondrial Protease LONP1
Notice bibliographique
Résumé
Despite advances in targeted therapies, survival for AML patients remains poor. Molecular classification systems attempt to stratify patients into risk groups, but 20-40% of patients do not fall under established genomic subgroups (Papaemmanuil et al., N Engl J Med 2016) We analyzed a total of 1,627 unique patient samples from the AML-PMP, TCGA, Beat AML, and TARGET datasets and identified loss-of-function mutations in ubiquitin ligase family genes in 10% of patient samples (n = 163). Among recurrently mutated genes, expression of FBXO11 was the most significantly reduced component of the SKP1-CUL1-Fbox (SCF) E3 ligase complex in AML samples compared to normal CD34+ hematopoietic stem/progenitor cells (HSPC). RNA sequencing (RNA-seq) following knockdown (KD) of FBXO11 by shRNA in cord blood derived CD34+ HSPCs resulted in enrichment of hematopoietic stem (HSC) and leukemia stem cell (LSC) signatures accompanied by a diminishment in hematopoietic progenitor cell signatures. Consistent with enrichment of a quiescent stem cell phenotype, cell cycle analysis showed that FBXO11 KD increased the G0 fraction of the CD34+ population. In liquid culture, FBXO11 KD HSPCs displayed myeloid-biased output. Fbxo11 KD in mouse HSPCs co-operated with AML1-ETO (RUNX1-RUNX1T1) fusion to generate serially transplantable AML in mice. The combination of FBXO11 KD with AML1-ETO and a KRASG12D mutant in human cord blood HSPC also promoted stem cell maintenance and myeloid malignancy in a human xenotransplant model, but AML1-ETO with KRASG12D without FBXO11 KD did not. Mass spectrometry analysis of proteins co-immunoprecipitating with FLAG-FBXO11 in myeloid cells identified the the mitochondria protease and chaperone protein LONP1. LONP1 protein expression remained unchanged with FBXO11 overexpression or CRISPR/Cas9-mediated targeting, suggesting that LONP1 is not targeted for degradation by the SCFFBXO11 complex. FBXO11 has been demonstrated to facilitate ubiquitination (Xu et al., Blood 2021) and neddylation (Abida et al., J Biol Chem 2007) of protein targets. FBXO11 expression resulted in activating K63 ubiquitination as well as neddylation of LONP1, but only ubiquitination inhibition resulted in reversal of an FBXO11-overexpression phenotype. Further, proteasome inhibitors also did not reverse the FBXO11-overexpression phenotype, consistent with FBXO11 mediating an activating ubiqutination of LONP1. FBXO11 or LONP1 KD in cord blood HSPCs promoted myeloid biased output and reduced mitochondria membrane potential, a characteristic of long-term HSCs (Mansell et al., Cell Stem Cell 2021). To validate these findings and direction of the FBXO11-LONP1 relationship, we performed RNA-seq on cord blood HSPCs following lentiviral transductions to combinatorially deplete or overexpress FBXO11 and LONP1. Unsupervised clustering of the samples demonstrated overexpression of LONP1 clustered with controls while knockdown of LONP1 reversed the effect of FBXO11 overexpression. We observed upregulation of a mitochondria respiratory defect gene set with depletion of FBXO11 or LONP1, indicating improper assembly of the electron transport chain, a reported consequence of impaired LONP1 function (Ghosh et al., Oncogene 2019). We also determined enrichment in several HSC and LSC genesets with KD of either FBXO11 or LONP1, while overexpression of FBXO11 downregulated these genesets. In contrast to normal HSCs which utilize glycolysis for energy production (Simsek et al., Cell Stem Cell 2010), LSCs from AML patients, are dependent on oxidative phosphorylation for energy metabolism but remain in a quiescent state (Jones et al., Cancer Cell 2018). Interestingly, FBXO11 depletion resulted in downregulated glycolysis genesets and upregulated quiescent stem cell signatures. Taken together, these findings suggest that FBXO11 depletion dysregulates LONP1 function to establish a mitochondrial state reminiscent of quiescent HSC. In summary, we demonstrate that the ubiquitin ligase family is frequently mutated in AML. We show that gene expression of a component of the SCF E3 ubiquitin ligase, FBXO11, is reduced in AML, and that FBXO11 depletion cooperates with other driver mutations to initate AML in mouse and human xenotransplant models. Finally, we establish a novel link between FBXO11 and LONP1, and provide evidence that loss of FBXO11 regulation of LONP1 promotes a myeloid-biased HSC/LSC-like state.
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Comment cette classification a été obtenuedéplier
Prédiction machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,000 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,001 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule source (Gemma direct ou Codex distillé), pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».