RÉSUMÉ
C/EBPα (p42 and p30 isoforms) is commonly dysregulated in cancer via the action of oncogenes, and specifically in acute myeloid leukaemia (AML) by mutation. Elevated TRIB2 leads to the degradation of C/EBPα p42, leaving p30 intact in AML. Whether this relationship is a cooperative event in AML transformation is not known and the molecular mechanism involved remains elusive. Using mouse genetics, our data reveal that in the complete absence of C/EBPα, TRIB2 was unable to induce AML. Only in the presence of C/EBPα p42 and p30, were TRIB2 and p30 able to cooperate to decrease the latency of disease. We demonstrate that the molecular mechanism involved in the degradation of C/EBPα p42 requires site-specific direct interaction between TRIB2 and C/EBPα p42 for the K48-specific ubiquitin-dependent proteasomal degradation of C/EBPα p42. This interaction and ubiquitination is dependent on a critical C terminal lysine residue on C/EBPα. We show effective targeting of this pathway pharmacologically using proteasome inhibitors in TRIB2-positive AML cells. Together, our data show that excess p30 cooperated with TRIB2 only in the presence of p42 to accelerate AML, and the direct interaction and degradation of C/EBPα p42 is required for TRIB2-mediated AML.
Sujet(s)
Protéine alpha liant les séquences stimulatrices de type CCAAT/génétique , Calcium-Calmodulin-Dependent Protein Kinases/génétique , Protéines et peptides de signalisation intracellulaire/génétique , Leucémie aigüe myéloïde/génétique , Isoformes de protéines/génétique , Animaux , Protéine alpha liant les séquences stimulatrices de type CCAAT/antagonistes et inhibiteurs , Calcium-Calmodulin-Dependent Protein Kinases/antagonistes et inhibiteurs , Différenciation cellulaire/effets des médicaments et des substances chimiques , Prolifération cellulaire/effets des médicaments et des substances chimiques , Humains , Protéines et peptides de signalisation intracellulaire/antagonistes et inhibiteurs , Leucémie aigüe myéloïde/traitement médicamenteux , Leucémie aigüe myéloïde/anatomopathologie , Souris , Mutation , Inhibiteurs du protéasome/administration et posologie , Isoformes de protéines/biosynthèseRÉSUMÉ
Bcr-Abl is a constitutively active tyrosine kinase involved in the development and progression of chronic myeloid leukaemia (CML). It has been demonstrated that Bcr-Abl-positive cells can be uniquely resistant to apoptosis induced by different types of stimuli, but the mechanism by which this is achieved is not defined. In this study we have investigated how cells expressing high expression levels of Bcr-Abl may gain resistance to cytotoxic drugs. We have established cell lines expressing low and high expression levels of Bcr-Abl. Cells expressing elevated Bcr-Abl are resistant to cytotoxic drugs. In drug-sensitive 32D-parental and low Bcr-Abl expressing cells, pro-apoptotic Bcl-2 family members, Bax and Bad translocate from the cytosol to the mitochondrion following a cytotoxic insult. In contrast, high Bcr-Abl expression prevents the early translocation of these pro-apoptotic proteins to the mitochondrion, mitochondrial membrane potential is retained and caspases are inactive. We also demonstrate that IL-3 can contribute to drug resistance in low Bcr-Abl expressing cells, however, independent inhibition of IL-3 activated pathways (PI3K/AKT and Jak/STAT) does not sensitise cells to apoptosis. This study demonstrates that the subcellular translocation of Bax and Bad can be regulated by elevated Bcr-Abl expression and this may be a key event in the abrogation of an apoptotic response following a cytotoxic insult.
Sujet(s)
Protéines de transport/métabolisme , Protéines de fusion bcr-abl/biosynthèse , Cellules souches hématopoïétiques/métabolisme , Mitochondries/métabolisme , Protéines proto-oncogènes/métabolisme , Antinéoplasiques d'origine végétale/pharmacologie , Apoptose , Caspase-3 , Caspases/métabolisme , Cytoplasme/métabolisme , Résistance aux médicaments antinéoplasiques , Antienzymes/pharmacologie , Étoposide/pharmacologie , Protéines de fusion bcr-abl/génétique , Cellules souches hématopoïétiques/anatomopathologie , Humains , Immunotransfert , Interleukine-3/pharmacologie , Kinase Janus-2 , Potentiels de membrane , Phosphorylation , Protein-tyrosine kinases/antagonistes et inhibiteurs , Protéines proto-oncogènes c-bcl-2/métabolisme , Tyrosine/métabolisme , Protéine Bax , Protéine Bad , Protéine bcl-XRÉSUMÉ
A characteristic feature of chronic myeloid leukaemia (CML) is the inevitable advancement from a treatable chronic phase to a fatal, drug-resistant stage referred to as blast crisis. The molecular mechanisms responsible for this disease transition remain unknown. As increased expression of Bcr-Abl has been associated with blast crisis CML, we have established transfectants in 32D cells that express low and high levels of Bcr-Abl, and assessed their drug sensitivity. Cells with high Bcr-Abl expression levels are resistant to conventional cytotoxic drugs, and also require higher levels of STI571 (an inhibitor of Bcr-Abl), to induce cell death. Co-treatment with cytotoxic drugs and STI571 increased the sensitivity of the drug-resistant cells. Despite the drug-resistant phenotype, high Bcr-Abl levels concomitantly increased the expression of p53, p21, Bax and down-regulated Bcl-2. These cells maintain a survival advantage irrespective of a reduced proportion of cycling cells and the pro-apoptotic shift in gene expression. In addition, the level of Bcr-Abl expression (high or low) does not alter the growth factor independence and elevated Bcl-xL expression observed. Our study indicates that drug resistance can be primarily attained by increased Bcr-Abl expression, and highlights the potential of therapy which combines STI571 with conventional cytotoxic drugs.
Sujet(s)
Résistance aux médicaments antinéoplasiques , Protéines de fusion bcr-abl/biosynthèse , Cellules souches hématopoïétiques/effets des médicaments et des substances chimiques , Antinéoplasiques/pharmacologie , Apoptose/effets des médicaments et des substances chimiques , Apoptose/physiologie , Benzamides , Cycle cellulaire/effets des médicaments et des substances chimiques , Division cellulaire/effets des médicaments et des substances chimiques , Cellules cultivées , Résistance aux médicaments antinéoplasiques/génétique , Antienzymes/pharmacologie , Protéines de fusion bcr-abl/métabolisme , Cellules souches hématopoïétiques/cytologie , Cellules souches hématopoïétiques/enzymologie , Humains , Mésilate d'imatinib , Interleukine-3/pharmacologie , Phénotype , Pipérazines/pharmacologie , Protein-tyrosine kinases/antagonistes et inhibiteurs , Protein-tyrosine kinases/pharmacologie , Pyrimidines/pharmacologie , TransfectionRÉSUMÉ
Chronic myeloid leukemia (CML) is a disease of the hematopoietic system, characterized by the presence of the Bcr-Abl oncoprotein. The main characteristics of this disease include adhesion independence, growth factor independence, and resistance to apoptosis. Loss or mutation of the tumor suppressor gene, p53, is one of the most frequent secondary mutations in CML blast crisis. The transition between chronic phase and blast crisis is associated with increased resistance to apoptosis correlating with poor prognosis. This review focuses on the involvement of these two oncoproteins in the development and progression of the apoptotic-resistant phenotype in CML.