Post-transcriptional regulation of bcl-2 in acute myeloblastic leukemia: significance for response to chemotherapy.

The blast stem cells of acute myeloblastic leukemia become more sensitive in culture to the chemotherapeutic agents cytosine arabinoside (Ara-C) and daunorubicin (DNR) when exposed to all-trans retinoic acid (ATRA) after drug. We have proposed that down regulation of bcl-2 by ATRA is part of the mechanism of sensitization. The hypothesis is based on reduced expression of bcl-2 mRNA, as seen in Northern blots, after ATRA. Nuclear run on experiments, however, failed to account completely for the effect at the transcriptional level. Accordingly, we looked for post-transcriptional effects of ATRA on bcl-2, using metabolic labelling of the protein to measure stability. We found that the half-life of bcl-2 protein is markedly shortened after treatment with ATRA. Hydrocortisone (HC) protects cells against the toxic effects of Ara-C or DNR when given before drug. HC does not alter bcl-2 expression at the level of mRNA; however, metabolic labelling shows that newly synthesized bcl-2 protein is stabilized in blast cells treated with HC. Response to Ara-C by growth factor responsive blast cells is influenced by the factor in the cultures; cells are more sensitive in cultures with G-CSF and less sensitive when GM-CSF is present. We compared two blast cell lines, OCI/AML-5, primarily responsive to GM-CSF, and OCI/AML-10, primarily responsive to G-CSF. Growth factor did not influence the stability of bcl-2 protein in either line. In contrast, Western blots showed that the amount of bcl-2 protein was greater in cultures with GM-CSF or GM-CSF in combination with G-CSF than in cultures with G-CSF or no added factor. This pattern was seen regardless of the mitogenic response to G-CSF or GM-CSF. We interpret our findings as indicating that bcl-2 protein is transcriptionally activated; that the stability of the protein is decreased after ATRA and increased after HC; that the amount of bcl-2 protein is greater in cultures with GM-CSF than in cultures with G-CSF, regardless of which factor gives the greater mitogenic response. We propose that these post-transcriptional modifications of transcriptionally activated bcl-2 account, in part, for the regulation of drug sensitivity by ATRA, HC and growth factors.