Notch signaling enhances survival and alters differentiation of 32D myeloblasts

The Notch transmembrane receptors play important roles in precursor surviva l and cell fate specification during hematopoiesis, To investigate the func tion of Notch and the signaling events activated by Notch in myeloid develo pment, me expressed truncated forms of Notch1 or Notch2 proteins that eithe r can or cannot activate the core binding factor 1 (CBF1) in 32D (clone 3) myeloblasts. 32D cells proliferate as blasts in the presence of the cytokin es, GM-CSF or IL-3, but they initiate differentiation and undergo granulopo iesis in the presence of granulocyte CSF (G-CSP). 32D cells expressing cons titutively active forms of Notch1 or Notch2 proteins that signal through th e CBF1 pathway maintained significantly higher numbers of viable cells and exhibited less cell death during G-CSF induction compared with controls, Th ey also displayed enhanced entry into granulopoiesis, and inhibited postmit otic terminal differentiation. In contrast, Notch1 constructs that either l acked sequences necessary for Coil binding or that failed to localize to th e nucleus had little effect. Elevated numbers of viable cells during G-CSF treatment were also observed in 32D cells overexpressing the basic helix-lo op-helix protein (bHLH), HES1, consistent with activation of the CBF1 pathw ay. Taken together, our data suggest that Notch signaling enhances 32D cell survival, promotes entry into granulopoiesis, and inhibits postmitotic dif ferentiation through a CBF1-dependent pathway.