Chromosomal translocations involving the Mixed Lineage Leukemia (MLL) gene lead to the expression of MLL fusion proteins and acute leukemia. MLL fusion protein-induced leukemia is aggressive, and often refractory to therapy, highlighting the importance of studying the pathogenesis of this disease. MLL fusion proteins upregulate wild-type MLL target genes, including HOX genes, and block hematopoietic differentiation, promoting leukemogenesis. However, the precise mechanism by which MLL fusion proteins upregulate HOX genes and block differentiation has been unclear. My thesis research shows that leukemia cells expressing the MLL fusion protein MLL-AF9 also express wild-type MLL from the non-translocated MLL allele. Wild-type MLL is required for MLL-AF9-mediated HOX gene upregulation and leukemogenesis. Menin, a nuclear DNA-binding protein, recruits both wild-type MLL and MLL-AF9 to HOX genes to activate their transcription, highlighting the central role of menin in this disease. We also found that menin blocks MLL-AF9 leukemia cell differentiation by promoting the expression of the polycomb group protein EZH2. EZH2 represses target genes of the pro-differentiation transcription factor C/EBP&alpha, a previously unknown mechanism for blocking differentiation. The cooperation between EZH2 and trithorax-associated menin is counter to the classical opposing roles of polycomb and trithorax proteins. These findings have uncovered novel insights into how menin and MLL-AF9 upregulate target genes and block leukemia cell differentiation, highlighting novel potential therapeutic targets for this disease.