A New AHI-1-DNM2-BCR-ABL Complex Regulates Endocytosis Processes in Chronic Myeloid Leukemia
Notice bibliographique
Résumé
Abstract Tyrosine kinase inhibitors (TKIs) have been introduced into clinical practice with remarkable effects on chronic phase CML. However, early relapses, acquired drug resistance, and persistence of leukemic stem cells remain problematic. Improved treatment approaches to target other key molecular elements active in CML stem/progenitor cells are needed. One candidate is AHI-1 (Abelson helper integration site-1), an oncogene that is highly deregulated in CML stem cells. It harbors two key domains, SH3 and WD40-repeat, which are known important mediators of protein-protein interactions. The AHI-1-mediated protein complex containing BCR-ABL and JAK2 has been shown to mediate transforming activity and TKI-response/resistance of CML stem/progenitor cells. We have recently identified Dynamin-2 (DNM2) as another AHI-1 interacting protein using the AHI-1 SH3 domain as protein ‘bait’ in immunoprecipitation/mass spectrometry. DNM2, a large GTPase, is mainly involved in the trafficking processes such as endocytosis, and is activated through tyrosine phosphorylation. Its role in the mediation of CML stem cell functions is unknown. We have now demonstrated that transcript levels of DNM2 are significantly increased in pre-treatment CD34+ stem/progenitor cells from CML patients who were classified retrospectively, after IM therapy, as IM-responders (n=11) and IM-nonresponders (n=15) as compared to CD34+ normal bone marrow cells (n=7, p=0.013 and 0.037). In addition, DNM2 is more highly expressed in CML stem cells (CD34+CD38-) and progenitor cells (CD34+CD38+) than more mature cells (CD34- , 2-fold). Co-immunopreciptation with mutant forms of AHI-1 and DNM2 (HA-AHI-1, HA-AHI-1 SH3Δ, Myc-DNM2 and Myc-DNM2 PRDΔ) in 293T cells indicated that the PRD domain of DNM2 is mainly responsible for the interaction between AHI-1 and DMN2. Co-localization analysis using confocal microscopy further demonstrated that the interaction between full-length AHI-1 and DNM2 occurs in a “punctate dot” pattern throughout the whole cytoplasm; in contrast, the co-localization signals were significantly disrupted in cells co-transfected with AHI-1 and DNM2 mutants. Interestingly, in AHI-1 SH3D mutant cells, AHI-1was located in the nucleus, suggesting that the SH3 domain of AHI-1 is required for AHI-1 cytoplasmic retention. Moreover, AHI-1 was observed to co-localize with EEA-1 (early endosome marker) and LAMP-1 (late endosome marker) in cells co-transfected with full-length AHI-1 and DNM2, but not in AHI-1 and DNM2 mutant cells. Increased transferrin signals were also demonstrated in AHI-1 and DNM2 co-transfected cells compared to their mutant cells, using transferrin uptake assays, suggesting that the interaction between DNM2 and AHI-1 indeed increases the kinetics and efficiency of endocytosis. These results were further confirmed in BCR-ABL-transduced and BCR-ABL/AHI-1 co-transduced hematopoietic cells that are relatively resistant to TKI-induced apoptosis. Particularly, the co-localization signals between DNM2 and AHI-1 were stronger in BCR-ABL/AHI-1 co-transduced cells, and the transferrin uptake was also more efficient as compared to BCR-ABL-transduced cells. On the other hand, transferrin uptake was reduced in CML cells with knockdown of DNM2 and these cells also demonstrated reduced proliferation and increased sensitivity to IM treatment compared to control cells. Importantly, we further identified a new protein interaction between DNM2 and BCR-ABL in both BCR-ABL and BCR-ABL/AHI-1 co-transduced cells and this interaction is enhanced in BCR-ABL/AHI-1 co-transduced cells using co-IP/Western analysis. To the best of our knowledge, this is the first study to implicate this new AHI-1-DNM2-BCR-ABL complex in the deregulation of endocytosis signaling in CML, which may play an unusual role in regulation of the cellular properties of primitive CML cells, including their response/resistance to TKI, by aberrantly disrupting critical endocytosis processes in CML. Disclosures No relevant conflicts of interest to declare.
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Prédiction machine sur la base complète
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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 ».