Identification of a Highly Deregulated eIF4F Translation Initiation Complex in Drug-Resistant BCR-ABL + Cells By a Phospho-Proteomic Antibody Microarray
Notice bibliographique
Résumé
Abstract Protein-tyrosine kinase inhibitors (TKIs) have been effective for treatment of early stages of chronic myeloid leukemia (CML). However, BCR-ABL-dependent resistance mechanisms and TKI-unresponsive quiescent leukemic stem cells (LSCs) can result in drug resistance and disease relapse. We have demonstrated that Abelson helper integration site-1 (AHI-1) is a highly deregulated protein in CML LSCs. It interacts with BCR-ABL through the AHI-1 WD40 domain and with other proteins, like dynamin-2 (DNM2), through its SH3 domain, to enhance leukemic-initiating activities and TKI resistance. To uncover downstream effects of the AHI-1-BCR-ABL-DNM2 complex and its biological role in mediating TKI resistance in CML stem/progenitor cells, an advanced antibody microarray analysis was then used to investigate differences in the proteome and phosphorylation landscape of BCR-ABL + cells co-transduced with wild-type Ahi-1 or Ahi-1 SH3 Δ mutant in the presence or absence of imatinib (IM). The microarray simultaneously quantified the differences in expression and phosphorylation sites of proteins in multiple signaling pathways using 1325 antibodies in duplicate measurements, and each microarray analysis was performed in duplicate. Significant changes in antibody signals for protein expression or phosphorylation were determined using limma. This analysis revealed that the overexpression of wild-type Ahi-1 (WT Ahi-1) has a profound differential effect, compared to the SH3 domain deleted Ahi-1 (Ahi-1 SH3 Δ), on the expression and phosphorylation status of proteins in BCR-ABL + cells with and without IM treatment. BCR-ABL + cells co-expressing WT Ahi-1 had a greater number of significantly differential antibody signals (7 increases, 42 decreases) when compared to BCR-ABL + cells, while Ahi-1 SH3 Δ expressing cells resulted in fewer significantly differential antibody signals (12 increases, 2 decreases) compared to BCR-ABL + cells. IM treatment resulted in WT Ahi-1 expressing cells having the greatest number of significantly differential antibody signals (7 increases, 56 decreases) compared to BCR-ABL + cells (5 decreases) and those co-expressing Ahi-1 SH3 Δ (2 increases, 9 decreases). Pathway enrichment analysis, using gProfiler, identified that the targets with significantly increased differential antibody signal after IM treatment in WT Ahi-1 cells were related to the regulation of translation initiation complex (p>0.0001). Interestingly, our RNA-seq dataset analysis further identified several members of the eukaryotic initiation factor 4F (eIF4F) complex to be significantly upregulated in CD34 + CML patient cells compared to normal bone marrow, particularly eIF4G1, the scaffold protein of the eIF4F complex involved in translation initiation (2-fold, p=0.001), as well as mTOR, a key regulator that controls the assembly of the eIF4F complex. This finding prompted us to further explore the regulation of translation initiation and the members of the eIF4F complex in BCR-ABL + cells. Immunoblotting demonstrated that BCR-ABL + cells co-transduced with WT Ahi-1 showed increased expression of eIF4G1 (3-fold) and eIF4B (2-fold), a cofactor that regulates the helicase activity of the eIF4F complex, compared to BCR-ABL + cells. Additionally, Cyclin D3, a gene reported to be sensitive to eIF4F translational activity, was found to have slightly increased expression (1.4-fold) in WT Ahi-1 cells compared to BCR-ABL + cells. Most interestingly, these results were also demonstrated in IM-resistant CML cells as compared to IM-sensitive cells, with an increase in eIF4G1 expression (2-fold), phosphorylation of eIF4B (5-fold, p=0.003), and Cyclin D3 expression (4-fold, p<0.05). Mechanistically, eIF4G1 knockdown by shRNA impaired survival (5-fold, p<0.0001) and increases TKI sensitivity in IM-resistant cells (3-fold, p<0.05). A new protein interaction between eIF4G1 and the mRNA cap-binding protein eIF4E was further identified in these cells using a proximity ligation assay. Thus, we have uncovered that the eIF4F complex, the key regulator of the mRNA-ribosome recruitment phase of translation initiation, has increased activity in IM-resistant cells, which may contribute to the regulation of TKI resistance in CML. Disclosures No relevant conflicts of interest to declare.
Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.
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) | 0,002 | 0,001 |
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 ».