ELL-associated factor 2 (EAF2), a functional homolog of EAF1 with alternative ELL binding properties.
The (11;19)(q23;p13.1) translocation in acute leukemia results in the formation of an MLL-ELL fusion protein. ELL is an RNA polymerase II elongation factor that interacts with the recently identified EAF1 protein. To characterize the normal functions of ELL and its aberrant activities when fused to MLL, we isolated a second protein that interacts with ELL named EAF2 for ELL Associated Factor 2. EAF2 is highly homologous to EAF1, with 58% identity and 74% amino acid conservation. Using specific antibodies generated to EAF2, we coimmunoprecipitated ELL and EAF2 from multiple cell lines. Confocal microscopy revealed that endogenous EAF2 and ELL colocalized in a nuclear speckled pattern. Database comparisons with EAF2 identified a region with a high content of serine, aspartic acid, and glutamic acid residues that is conserved with EAF1 and exhibited amino acid similarity with several translocation partner proteins of MLL, including AF4 and ENL. We found that EAF2 and EAF1 both contain transcriptional activation domains within this region. Using retroviral bone marrow transduction, we observed that a heterologous fusion of EAF2 to MLL immortalized hematopoietic progenitor cells. In contrast to EAF1, EAF2 does not bind to the carboxy-terminus of ELL. We identified a protein-protein interaction domain within the amino-terminus of ELL that binds to both EAF1 and EAF2. This amino-terminal interaction domain is disrupted in the formation of the MLL-ELL fusion protein. Thus, MLL-ELL retains an interaction domain for EAF1 but not for EAF2. Taken together, these data suggest that MLL-ELL may disrupt the normal protein-protein interactions of ELL.
Simone
Federico
F
University of Chicago, Section of Hematology/Oncology, Chicago, IL 60637, USA.
Luo
Roger T
RT
Polak
Paul E
PE
Kaberlein
Joseph J
JJ
Thirman
Michael J
MJ
D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000032:analysis; D001665:Binding Sites; D015152:Blotting, Northern; D001853:Bone Marrow / Q000378:metabolism; D002467:Cell Nucleus / Q000737:chemistry; D002471:Cell Transformation, Neoplastic; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000235:genetics; D018698:Glutamic Acid / Q000032:analysis; D006367:HeLa Cells; D006412:Hematopoietic Stem Cells / Q000473:pathology; D011495:Histone-Lysine N-Methyltransferase; D006801:Humans; D007163:Immunosorbent Techniques; D015470:Leukemia, Myeloid, Acute / Q000473:pathology; D051379:Mice; D008969:Molecular Sequence Data; D051788:Myeloid-Lymphoid Leukemia Protein; D015514:Oncogene Proteins, Fusion / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D011519:Proto-Oncogenes; D011993:Recombinant Fusion Proteins; D012190:Retroviridae / Q000235:genetics; D017385:Sequence Homology; D012694:Serine / Q000032:analysis; D014157:Transcription Factors / Q000032:analysis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D035602:Transcriptional Elongation Factors; D014162:Transfection
D004268:DNA-Binding Proteins; C431383:EAF1 protein, human; C472771:EAF2 protein, human; C447613:ELL2 protein, human; C497422:KMT2A protein, human; C472772:MLL-ELL fusion protein, human; D015514:Oncogene Proteins, Fusion; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D051788:Myeloid-Lymphoid Leukemia Protein; D001224:Aspartic Acid; D018698:Glutamic Acid; D012694:Serine; D011495:Histone-Lysine N-Methyltransferase; C497423:Kmt2a protein, mouse