Identification of Meis1 Target Genes Involved in the Induction of AML in Collaboration with NUP98-HOXD13.

Meis1, a homeodomain-containing HOX cofactor, cooperates with multiple native and NUP98-HOX fusion proteins to accelerate the onset of AML. We have recently shown that enforced overexpression of Meis1 in NUP98-HOX transduced bone marrow (bm) cells leads to a marked elevation in the tyrosine receptor molecule, Flt3 transcription and subsequent activation of Flt3 signaling pathways (Blood. Aug 1, 2006). Furthermore, overexpression of wildtype Flt3 is sufficient to collaborate with NUP98-HOX fusions to induce AML in mice, albeit with a longer disease latency than NUP98-HOX+Meis1-mediated AML. To further study the molecular mechanisms underlying Meis1’s strong pro-leukemic effect together with HOX proteins, we conducted a structure-function analysis of Meis1 in the context of NUP98-HOXD13 (ND13) leukemogenesis. We show that the homeodomain (ΔHD, amino acids 272–335) is required for Meis1 collaboration with ND13 while the N-terminal domain (ΔN, amino acids 1–67) is dispensable for leukemogenesis but decelerates the disease onset. Interestingly, primary bm cells transduced with ND13-Meis1ΔHD or ND13-Meis1ΔN revealed no significant upregulation of Flt3 mRNA levels in either case. These results suggest that Meis1 triggers additional Flt3-independent pathways to accelerate leukemia development. Thus, the retained leukemogenic properties of ND13-Meis1ΔN and the longer latency observed for NUP98-HOX fusions and Flt3 overexpression to induce leukemia point to additional roles for Meis1 in induction of leukemia independent of its ability to upregulate Flt3. To search for these additional Meis1-induced leukemic pathway(s), we used the Affymetrix GeneChip MOE430 to compare the gene expression profiles of bm cells transduced with ND13, ND13+Meis1, ND13+Meis1ΔHD and Meis1ΔN. Interestingly, the gene array results indicate that only a relatively small number of genes (72 increased and 64 decreased) differed significantly between the non-leukemic Meis1ΔHD and the leukemic Meis1 bm cells and these genes are all strong candidates as to be direct Meis1 target genes. A significant number of these genes are involved in cytokine receptor pathways as revealed by gene ontology analysis and, furthermore, promoter analysis revealed the presence of putative Hox, Meis1 and Pbx binding sites in several of these. The gene array results also indicate that only a small fraction of genes differed significantly between the non-leukemic Meis1ΔHD and the leukemic Meis1ΔN bm cells suggesting that these genes could be involved in Flt3-independent Meis1 leukemogenic activity.