Abstract
In acute myeloid leukemias (AMLs) with t(8;21), the transcription factor AML1 is juxtaposed to the zinc finger nuclear protein ETO (Eight-Twenty-One), resulting in transcriptional repression of AML1 target genes. ETO has been shown to interact with corepressors, such as N-CoR and mSin3A to form complexes containing histone deacetylases. To define regions of ETO required for maximal repressor activity, we analyzed amino-terminal deletions in a transcriptional repression assay. We found that ETO mutants lacking the first 236 amino acids were not affected in their repressor activity, whereas a further deletion of 85 amino acids drastically reduced repressor function and high molecular weight complex formation. This latter mutant can still homodimerize and bind to N-CoR but shows only weak binding to mSin3A. Furthermore, we could show that a "core repressor domain" comprising nervy homology region 2 and its amino- and carboxyl-terminal flanking sequences recruits mSin3A and induces transcriptional repression. These results suggest that mSin3A and N-CoR bind to ETO independently and that both binding sites cooperate to maximize ETO-mediated transcriptional repression. Thus, ETO has a modular structure, and the interaction between the individual elements is essential for the formation of a stable repressor complex and efficient transcriptional repression.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acids / chemistry
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Cell Line
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Core Binding Factor Alpha 2 Subunit
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DNA-Binding Proteins / antagonists & inhibitors
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DNA-Binding Proteins / chemistry*
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DNA-Binding Proteins / genetics
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Dimerization
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Electrophoresis, Polyacrylamide Gel
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Gene Deletion
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Glutathione Transferase / metabolism
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Humans
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Leukemia, Myeloid, Acute / genetics
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Leukemia, Myeloid, Acute / metabolism
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Mutagenesis, Site-Directed
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Nuclear Proteins / metabolism
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Nuclear Receptor Co-Repressor 1
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Plasmids / metabolism
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Precipitin Tests
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Protein Binding
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Protein Structure, Tertiary
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Proto-Oncogene Proteins*
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RUNX1 Translocation Partner 1 Protein
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Recombinant Fusion Proteins / metabolism
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Repressor Proteins / metabolism
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Sin3 Histone Deacetylase and Corepressor Complex
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Transcription Factors / antagonists & inhibitors
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Transcription Factors / chemistry*
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Transcription Factors / genetics
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Transcription, Genetic*
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Transfection
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Zinc Fingers
Substances
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Amino Acids
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Core Binding Factor Alpha 2 Subunit
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DNA-Binding Proteins
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NCOR1 protein, human
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Nuclear Proteins
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Nuclear Receptor Co-Repressor 1
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Proto-Oncogene Proteins
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RUNX1 Translocation Partner 1 Protein
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RUNX1 protein, human
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RUNX1T1 protein, human
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Recombinant Fusion Proteins
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Repressor Proteins
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SIN3A transcription factor
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Transcription Factors
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Glutathione Transferase
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Sin3 Histone Deacetylase and Corepressor Complex