It is well established that c-myc expression is down-regulated during induction of differentiation in various leukemia cell lines including HL-60 human myeloid leukemia cells. Previously we have shown that c-myc protein expression was specifically down-regulated in G0/G1 phase of the cell cycle during induction of granulocytic differentiation with dimetylsulfoxide (DMSO) in HL-60 cells. In the present study, we examined whether c-myc protein expression is also down-regulated in G0/G1 phase during monocytic differentiation of HL-60 cells. HL-60 cells can be induced to differentiate along monocytic lineage by treatment with 1,25 dihydroxyvitamin D3 (D3). C-myc protein expression was down-regulated after 72h of treatment with D3 in HL-60 cells. Simultaneous analysis of protein level and cell cycle using a two-color flow cytometry showed that the down-regulation of c-myc protein expression occurred specifically in G0/G1 phase. Down-regulation of c-myc protein expression in G0/G1 phase was more clearly shown by using synchronized cell population that was arrested at G0/G1 phase by serum starvation. Furthermore, G0/G1-synchronized cells showed a higher susceptibility to differentiation in response to D3. These results suggest that c-myc protein is down-regulated in G0/G1-specific phase during not only granulocytic differentiation but also monocytic differentiation, and down-regulation of c-myc protein expression in G0/G1 phase is involved in the program of differentiation of HL-60 cells.