Multipotent hematopoietic cell lines derived from C/EBP alpha(-/-) knockout mice display granulocyte macrophage-colony-stimulating factor, granulocyte-colony-stimulating factor, and retinoic acid-induced granulocytic differentiation

The transcription factor C/EBP alpha is an important mediator of granulocyt e differentiation and regulates the expression of multiple granulocyte-spec ific genes including the granulocyte-colony-stimulating factor (GCSE) recep tor, neutrophil elastase, and myeloperoxidase. Indeed C/EBP alpha knockout mice display a profound block in granulocyte differentiation. To study this block in granulocytic differentiation in more detail, retroviral vector-me diated transduction of a dominant-negative retinoic acid receptor was used to establish hematopoietic growth factor-dependent, lympho-myeloid progenit or cell lines from the fetal livers of both the C/EBP alpha knockout animal s (C/EBP alpha(-/-)) and their heterozygous littermates (C/EBP alpha(+/-)). Surprisingly, the C/EBP alpha(-/-) cell lines displayed significant sponta neous granulocytic differentiation, and this differentiation was markedly e nhanced when the cells were stimulated with granulocyte macrophage (GM)-CSF . This GM-CSF-mediated differentiation was associated with the up-regulatio n of G-CSF receptor mRNA, and the combination of GM-CSF and G-CSF generated more than 95% mature neutrophils in the C/EBP alpha(-/-) cultures. The add ition of all-trans retinoic acid also enhanced this granulocytic differenti ation of the cultured C/EBP alpha(-/-) cells, indicating that the activated retinoic acid receptors can enhance granulocytic differentiation through a molecular pathway that is independent of C/EBP alpha. These studies clearl y indicate that terminal granulocytic differentiation associated with the u p-regulation of C/EBP alpha -responsive genes can occur in the absence of C /EBP alpha, and they indicate the existence of multiple independent molecul ar pathways potentially used by primitive hematopoietic precursors that can lead to the development of mature granulocytes. (C) 2001 by The American S ociety of Hematology.