Abstract
Treatment of mammalian cells with 1,25-dihydroxyvitamin D3 (1,25D3) produces a G1 to S (G1/S) phase cell cycle block. In addition, it has been noted that a smaller proportion of cells accumulates in the G2/M compartment in 1,25D3-treated cultures. Since cyclins have a major influence on the regulation of cell cycle progression, we determined the expression of cyclins A and B as markers of the G2 phase and of cyclin E as the marker of G1/S transition. No increase in the steady-state levels of cyclin A or cyclin B mRNA was detected in the total cell population or in the cyclin B1 protein in the G2/M cell cycle compartment. In contrast, immunodetectable cyclin E protein was increased in cell cultures as a whole and specifically in the G2/M compartment cells. Determination of BrdU incorporation into DNA by flow cytometry showed marked inhibition of DNA replication in cells with DNA content higher than 4C, and autoradiography of 3H-TdR-pulsed cells showed that polynucleated cells did not replicate DNA after 96 h of treatment with 1,25D3 or analogs. Taken together, these experiments show that at least a portion of the G2/M compartment in 1,25D3-arrested cultures of HL60 cells represents G1 cells at a higher ploidy level, which are blocked from entering the high ploidy S phase.
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