REGULATION OF THE SYNTHESIS OF BCL-2 PROTEIN BY GROWTH-FACTORS

The sensitivity of AML blast stem cells can be measured in cell cultur e, using a clonogenic assay to determine survival after each of a grad ed series of drug concentrations. For cytosine arabinoside, the dose-r esponse curve is a simple negative exponential that can be described b y a D-10 value, a measure of slope. This D-10 value can be affected by regulatory molecules added to the cultures. All-trans retinoic acid ( ATRA) usually sensitizes cells, while hydrocortisone (HC) is protectiv e. Growth factor responsive cells are more Ara-C sensitive in G-CSF th an in GM-CSF or IL-3. The proto-oncogene bcl-2 may be part of the mech anism by which drug sensitivity is regulated. Previous work has shown that ATRA decreases bcl-2 RNA expression and the half-life of the prot ein; in contrast, the protein from cells treated with HC is more stabl e than controls. Growth factors were not shown to change either expres sion of bcl-2 RNA or the stability of its protein. In this paper, we d escribe experiments where OCI/AML-1 cells were grown in G-CSF and then transferred to medium containing both G-CSF and the GM-CSF-IL-3 fusio n protein pIXY. Steady-state levels of bcl-2 protein were measured by Western blot and synthesis by incorporation of S-35 methionine into pr otein. We observed that both measures doubled within 12-24 h after tra nsfer from G-CSF in G-CSF with pIXY, but promptly returned to the prev ious state when pIXY was withdrawn. We conclude that growth factors re gulate that activity of bcl-2 post-transcriptionally by altering the r ate of synthesis of the protein.