Enhanced proliferative potential of hematopoietic cells expressing degradation-resistant c-Myb mutants.

The c-myb gene encodes a transcription factor required for proliferation, differentiation, and survival of hematopoietic cells. Expression of c-Myb is often increased in hematological malignancies, but the underlying mechanisms are poorly understood. We show here that c-Myb has a longer half-life (at least 2-fold) in BCR/ABL-expressing than in normal hematopoietic cells. Such enhanced stability was dependent on a phosphatidylinositol 3-kinase (PI-3K)/Akt/GSKIIIbeta pathway(s) as indicated by the suppression of c-Myb expression upon treatment with PI-3K inhibitors or co-expression with dominant negative Akt or constitutively active GSKIIIbeta. Moreover, inhibition of GSKIIIbeta by LiCl enhanced c-Myb expression in parental 32Dcl3 cells. Compared with wild type c-Myb, three mutants (delta(358-452), delta(389-418), and L389A/L396A c-Myb) of the leucine zipper domain had increased stability. However, only expression of delta(358-452) was not affected by inhibition of the PI-3K/Akt pathway and was not enhanced by a proteasome inhibitor, suggesting that leucine zipper-dependent and -independent mechanisms are involved in the regulation of c-Myb stability. Indeed, delta(389-418) carrying four lysine-to-alanine substitutions (delta(389-418) K387A/K428A/K442A/K445A) was as stable as delta(358-452) c-Myb. Compared with full-length c-Myb, constitutive expression of delta(358-452) and delta(389-418) c-Myb in Lin-Sca-1+ mouse marrow cells increased cytokine-dependent primary and secondary colony formation. In K562 cells, expression of delta(358-452), delta(389-418), and L389A/L396A c-Myb led to enhanced proliferation after STI571 treatment. Thus, enhanced stability of c-Myb by activation of PI-3K-dependent pathway(s) might contribute to the higher proliferative potential of BCR/ABL-expressing and, perhaps, other leukemic cells.