Functional mutagenesis of AML1/RUNX1 and PEBP2 beta/CBF beta define distinct, non-overlapping sites for DNA recognition and heterodimerization by the Runt domain.
The Runt domain family of transcription factors play key roles in transcriptional regulation of definitive hematopoiesis and osteogenesis. This transcription factor family is characterized by a DNA-binding alpha-subunit harboring the Runt domain and a secondary subunit, beta, which binds to the Runt domain and enhances its interaction with DNA. Missense mutations in the Runt domain from either the blood or bone-related gene product are associated with the onset of acute human leukemia as well as a disease of skeletal patterning known as cleidocranial dysplasia. NMR "footprinting" analysis of Runt domain/beta/DNA ternary complexes in solution previously identified the likely residues that form the heterodimerization and DNA-binding surfaces of the Runt domain. Functional mutagenesis at 37 positions in the Runt domain or beta confirms the original identification of these interaction surfaces and reveals that the heterodimerization and DNA-binding surfaces of the Runt domain occur at distinct, non-overlapping sites within the domain. The analysis of an additional 21 disease-related missense mutations identified from patients with either blood or bone disease demonstrates that the primary defect in these patients is a failure in DNA-recognition by the Runt domain. The molecular basis for the DNA-binding defect is analyzed in the context of the three-dimensional structure of the Runt domain in binary and ternary protein/DNA complexes.
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The Rockefeller University, 1230 York Avenue, Box 42, New York, NY 10021, USA.
Werner
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D019943:Amino Acid Substitution / Q000235:genetics; D001665:Binding Sites; D002942:Circular Dichroism; D002973:Cleidocranial Dysplasia / Q000235:genetics; D050676:Core Binding Factor Alpha 2 Subunit; D050658:Core Binding Factor beta Subunit; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D029721:Drosophila Proteins; D006801:Humans; D007951:Leukemia, Myeloid / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D016296:Mutagenesis / Q000235:genetics; D009687:Nuclear Proteins; D010641:Phenotype; D017354:Point Mutation / Q000235:genetics*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D050676:Core Binding Factor Alpha 2 Subunit; D050658:Core Binding Factor beta Subunit; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D009687:Nuclear Proteins; D021122:Protein Subunits; D011518:Proto-Oncogene Proteins; C493728:RUNX1 protein, human; D050656:Transcription Factor AP-2; D014157:Transcription Factors; C080833:run protein, Drosophila; D004247:DNA