Chimeric proteins joining the histone methyltransferase MLL with various fusion 
partners trigger distinctive lymphoid and myeloid leukemias. Here, we 
immunopurified proteins associated with ENL, a protein commonly fused to MLL. 
Identification of these ENL-associated proteins (EAPs) by mass spectrometry 
revealed enzymes with a known role in transcriptional elongation (RNA polymerase 
II C-terminal domain kinase [RNAPolII CTD] positive transcription elongation 
factor b [pTEFb]), and in chromatin modification (histone-H3 methyltransferase 
DOT1L) as well as other frequent MLL partners (AF4, AF5q31, and LAF4), and 
polycomb group members (RING1, CBX8, and BCoR). The composition of EAP was 
further verified by coimmunoprecipitation, 2-hybrid analysis, pull-down, and 
colocalization experiments. Purified EAP showed a histone H3 lysine 79-specific 
methylase activity, displayed a robust RNAPolII CTD kinase function, and 
counteracted the effect of the pTEFb inhibitor 
5,6-dichloro-benzimidazole-riboside. In vivo, an ENL knock-down diminished 
genome-wide as well as gene-specific H3K79 dimethylation, reduced global run-on 
elongation, and inhibited transient transcriptional reporter activity. According 
to structure-function data, DOT1L recruitment was important for transformation 
by the MLL-ENL fusion derivative. These results suggest a function of ENL in 
histone modification and transcriptional elongation.