ONLY LATE, NONMITOTIC STAGES OF GRANULOCYTE DIFFERENTIATION IN 32DCL3CELLS ARE BLOCKED BY ECTOPIC EXPRESSION OF MURINE C-MYB AND ITS TRUNCATED FORMS

In murine leukemia virus-induced myeloid leukemias, insertional mutage nesis of the c-myb locus has been shown to occur frequently. Proto-onc ogene activation is achieved in most leukemias by integration of murin e leukemia virus upstream of exons 3 or 4 or by integration into exon 9 with consequent truncation of the protein. The present study investi gates the effect of ectopic expression of full-length c-myb or c-myb c ontaining amino- or carboxyl-terminal truncations (minus 47 and 248 am ino acids, respectively) on granulocyte differentiation in vitro. Reco mbinant myb retroviruses were used to infect an interleukin 3-dependen t progenitor cell line, 32Dcl3, which undergoes terminal differentiati on to mature neutrophilic granulocytes in the presence of granulocyte colony-stimulating factor. Overexpression of c-myb did not abrogate th e interleukin 3 dependency of the parental cell line. However, cells e xpressing all forms of c-myb were blocked at an intermediate stage of granulocyte differentiation and continued to proliferate in the presen ce of granulocyte colony-stimulating factor. After 14 days in medium w ith granulocyte colony-stimulating factor, myb-expressing cultures pre dominantly consisted of promyelocytes with some myelocytes and almost undetectable numbers of neutrophilic granulocytes. This suggested that early stages of granulocyte differentiation were not inhibited, a fin ding that was further supported by the induction of myeloperoxidase, a biochemical marker of promyelocytes. Interestingly, the expression of lactoferrin, known to be a marker of late stages of granulocyte diffe rentiation, was completely inhibited in the cells infected with myb vi ruses. It was concluded that c-myb expression blocked granulocyte diff erentiation to the terminal mitotic stages and that deletion of the NH 2-terminal 47 amino acids and/or the COOH-terminal 248 amino acids of c-myb neither enhanced nor diminished this effect.