Differentiation of mouse erythroleukemia cells is blocked by late up-regulation of a c-myb transgene.

Abstract

During chemically induced differentiation of Friend virus-infected mouse erythroleukemia (MEL) cell lines, there is a biphasic down-regulation of the c-myb proto-oncogene. A plasmid containing a murine c-myb cDNA controlled by a mouse metallothionein I promoter was transfected into the C19 MEL cell line. For six transfected clones, it was found that expression of the exogenous c-myb mRNA could be up-regulated by the addition of 120 FIM ZnCl2 and that the N,N'-hexamethylenebisacetamide-induced differentiation of these transfectants was inhibited in proportion to the level of exogenous c-myb mRNA expression. By adding or removing ZnC12 at different times during the induction process, it was possible to show that up-regulation of exogenous c-myb limited to the first 2 days of induction had little or no effect on differentiation. In contrast, continuous expression of exogenous c-myb beginning at any time during the period of induction blocked further differentiation. These results suggest that during HMBA induction of MEL cells, the early down-regulation of c-myb mRNA is not necessary for terminal differentiation, whereas the down-regulation of c-myb at a later time is necessary.

Expression of BCR – ABL in M1 myeloid leukemia cells induces differentiation without arresting proliferation

“…Curiously, M1/SCL cells were partially inhibited in their clonal suppression and di erentiation in response to LIF and Oncostatin-M but were indistinguishable from control M1 cultures when exposed to IL-6 (Tanigawa et al, 1995). Consistent with their e ects on M1 differentiation, constitutive expression of c-MYB and c-MYC also inhibited the di erentiation and growth arrest of murine erythroleukemia cells treated with DMSO (Coppola and Cole, 1986;Dmitrovsky et al, 1986;Prochownik and Kukowska, 1986;Clarke et al, 1988;McClinton et al, 1990). Based on these studies, one may speculate that the inappropriately high levels of c-MYC (and c-MYB, data not shown) mRNA expressed by M1.210 cells played a role in inhibiting cytokine-induced growth arrest.…”

The carbonic anhydrase I locus contains a c-Myb target promoter and modulates differentiation of murine erythroleukemia cells

“…c-Myb is downregulated during HMBA induced differentiation and forced expression of c-Myb can block differentiation (Gonda and Metcalf, 1984;Marks et al, 1987;Clarke et al, 1988;McClinton et al, 1990). To develop a model that would allow the identification of Myb target genes, we have generated stable transfectants in the C19 MEL cell line that carry a dominant interfering Myb transgene (MEnT).…”

“…The C19 MEL cell line is a thymidine kinase-negative subclone of the 745 murine erythroleukemia cell line (McClinton et al, 1990) and was grown in Roswell Park Memorial Institute (RPMI)-1640 medium supplemented with 10% fetal bovine serum and 2 mM glutamine. Clone 17 cells are stable transfectants of the C19 MEL cell line that contain a dominant interfering Myb cDNA (MEnT) under the inducible transcriptional control of a mouse metallothionein I promoter and were cultured in the same medium containing G418 at a concentration of 400 mg/ml (Chen et al, 2002).…”

“…Filters were exposed to a phosphor screen and visualized by using a phosphorImager system. CAI MEL cell stable transfectants and protein blotting Stable transfections into C19 MEL cells were performed using lipofectin (Invitrogen) as we have previously described in detail (McClinton et al, 1990). Stable transfectants were cultured in growth medium containing 400 mg/ml G418 in the absence or presence of 100 mM ZnCl 2 for 16 h. Approximately 2 Â 10 7 cells were harvested and sonicated in 1 ml YRIPA lysis buffer as we have previously described (Aziz et al, 1995), and 100 mg of whole-cell lysates were resolved by sodium dodecylsulfate-10% polyacrylamide gel electrophoresis (SDS-PAGE).…”

“…Expression of c-Myb is downregulated during MEL cell differentiation, and forced expression of c-Myb blocks chemically induced MEL cell differentiation (McClinton et al, 1990). To gain insight into the mechanism of cMyb-regulated differentiation in MEL cells, we developed C19 MEL cell lines that carry an inducible dominant interfering Myb protein (MEnT) and induction of MEnT expression provides a sufficient signal for MEL cells to differentiate (Chen et al, 2002).…”