Degree of Recruitment of DOT1L to MLL-AF9 Defines Level of H3K79 Di- and Tri-methylation on Target Genes and Transformation Potential

• DOT1L has three interaction sites with AF9
• DOT1L-AF9 complex forms a mixed alpha-beta structure similar to the AF4-AF9 complex
• Blocking DOT1L binding to MLL-AF9 causes loss of serial replating capability
• Differential H3K79me2 and me3 loss is observed on subsets of MLL-AF9 target genes

SummaryThe MLL gene is a common target of chromosomal translocations found in human leukemia. MLL-fusion leukemia has a consistently poor outcome. One of the most common translocation partners is AF9 (MLLT3). MLL-AF9 recruits DOT1L, a histone 3 lysine 79 methyltransferase (H3K79me1/me2/me3), leading to aberrant gene transcription. We show that DOT1L has three AF9 binding sites and present the nuclear magnetic resonance (NMR) solution structure of a DOT1L-AF9 complex. We generate structure-guided point mutations and find that they have graded effects on recruitment of DOT1L to MLL-AF9. Chromatin immunoprecipitation sequencing (ChIP-seq) analyses of H3K79me2 and H3K79me3 show that graded reduction of the DOT1L interaction with MLL-AF9 results in differential loss of H3K79me2 and me3 at MLL-AF9 target genes. Furthermore, the degree of DOT1L recruitment is linked to the level of MLL-AF9 hematopoietic transformation.

Graphical AbstractFigure optionsDownload as PowerPoint slide