A Differential Screen for Ligand-Regulated Genes: Identification of HoxA10 as a Target of Vitamin D(3) Induction in Myeloid Leukemic Cells

1,25-Dihydroxyvitamin D(3) [1,25(OH)(2)D(3)], the hormonal ligand for vitamin D(3), is a potent inducer of myeloid-leukemic-cell differentiation. Such cells differentiate exclusively into monocytes/macrophages in response to this ligand. Since 1,25(OH)(2)D(3) transduces its hormone signal through the vitamin D(3) receptor (VDR), a ligand-modulated transcription factor and member of the nuclear hormone receptor superfamily, we sought to identify direct VDR target genes induced during this differentiation process. To do so, we applied a modified differential screen with a nascent-RNA purification strategy using biases for immediate-early-response genes induced by 1,25(OH)(2)D(3) in the myelomonocytic cell line U937. Using this screen, we had previously identified p21(Waf1/Cip1) as a gene transcriptionally induced by 1,25(OH)(2)D(3) and demonstrated that this induction facilitates the differentiation of U937 cells into monocytes/macrophages (24). Here, we describe in detail our differential screen strategy and the identification and isolation of 20 1,25(OH)(2)D(3)-inducible genes or unknown cDNAs by means of this screen. One gene newly identified as a target of VDR regulation in myeloid cells is the homeobox HoxA10 gene. HoxA10 protein may act as a general regulator of cell growth, since overexpression of HoxA10 facilitated the differentiation of U937 cells into monocytes/macrophages independent of 1,25(OH)(2)D(3) and acted to strongly inhibit the growth of the breast cancer cell line MCF-7 by arresting these cells in G(1).