Mechanisms of MLL Fusion Protein-Mediated Leukemic Transformation.

Abstract

Leukemogenic MLL fusion proteins, including MLL-AF4, MLL-AF9, and MLL-ENL, transform through up regulated expression of HOX genes and the HOX cofactor MEIS1. How they lead to the aberrant activation of these genes is not completely understood. In this study, we identified proteins recruited by one of the most common MLL fusion proteins, MLL-AF9, through purification of proteins that interact with the transforming domain of AF9 in myeloblastic M1 cells. Consistent with earlier purifications of ENL and AF4 from 293 cells, the 90 amino acid C-terminal domain associates with many of the most common MLL translocation partners including Enl, Af4, Laf4, Af5q31 and Af10. This complex, previously termed EAP for Elongation Assisting Proteins, contains the RNA polymerase II C-terminal domain kinase Cdk9/Cyclin T1/T2 (pTEFb), the histone H3 lysine 79 methyltransferase Dot1l, and the Polycomb proteins Pc3 and Ring1b. Chromatin immunoprecipitation (ChIP) studies show that myeloid cells transformed by MLL fusion proteins show higher levels and a broader distribution of EAP components at leukemogenic loci. Inhibition of EAP components pTEFb and Dot1l show that both contribute significantly to activation of Hoxa9 and Meis1 expression. EAP is dynamically associated with the Hoxa9 and Meis1 loci in hematopoietic cells and rapidly dissociates during induction of differentiation. However, in the presence of MLL fusion proteins, EAP dissociation from target loci is prevented. These data suggest that MLL fusion proteins deregulate key target genes by excessive recruitment and impaired dissociation of EAP from target loci.