Degree of recruitment of DOT1L to MLL-AF9 defines level of H3K79 Di- and tri-methylation on target genes and transformation potential.
The 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.
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA 22908, USA.
Aravinda
A
Kuntimaddi
Oncology Institute, Department of Medicine, Loyola University Chicago, Maywood, IL 60153, USA.
Nicholas J
NJ
Achille
Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, VA 22908, USA.
Jeremy
J
Thorpe
Oncology Institute, Department of Medicine, Loyola University Chicago, Maywood, IL 60153, USA.
Alyson A
AA
Lokken
Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, VA 22908, USA.
Ritambhara
R
Singh
Oncology Institute, Department of Pediatrics, Loyola University Chicago, Maywood, IL 60153, USA.
Charles S
CS
Hemenway
Department of Biochemistry and Molecular Genetics, University of Virginia, Charlottesville, VA 22908, USA.
Mazhar
M
Adli
Oncology Institute, Department of Medicine, Loyola University Chicago, Maywood, IL 60153, USA. Electronic address: nzelezn@luc.edu.
Nancy J
NJ
Zeleznik-Le
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA 22908, USA. Electronic address: jhb4v@virginia.edu.
John H
JH
Bushweller
D000595:Amino Acid Sequence; D001665:Binding Sites; D047369:Chromatin Immunoprecipitation; D011495:Histone-Lysine N-Methyltransferase; D006657:Histones / Q000378:metabolism*; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008745:Methylation; D008780:Methyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D051788:Myeloid-Lymphoid Leukemia Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015514:Oncogene Proteins, Fusion / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D017422:Sequence Analysis, DNA
D006657:Histones; C474558:MLL-AF9 fusion protein, human; D015514:Oncogene Proteins, Fusion; D011994:Recombinant Proteins; D051788:Myeloid-Lymphoid Leukemia Protein; C473477:DOT1L protein, human; D008780:Methyltransferases; D011495:Histone-Lysine N-Methyltransferase