Dysfunction of AML1/Runx1, a transcription factor, plays a crucial role in the development of many types of leukemia. Additional events are often required for AML1 dysfunction to induce full-blown leukemia; however, a mechanistic basis of their cooperation is still elusive. Here, we investigated the effect of AML1 deficiency on the development of MLL-ENL leukemia in mice. Aml1 excised bone marrow cells lead to MLL-ENL leukemia with shorter duration than Aml1 intact cells in vivo. Although the number of MLL-ENL leukemia-initiating cells is not affected by loss of AML1, the proliferation of leukemic cells is enhanced in Aml1-excised MLL-ENL leukemic mice. We found that the enhanced proliferation is the result of repression of p19^ARF that is directly regulated by AML1 in MLL-ENL leukemic cells. We also found that down-regulation of p19^ARF induces the accelerated onset of MLL-ENL leukemia, suggesting that p19^ARF is a major target of AML1 in MLL-ENL leukemia. These results provide a new insight into a role for AML1 in the progression of leukemia.