Multiple GTF2I-like repeats of general transcription factor 3 exhibit DNA binding properties. Evidence for a common origin as a sequence-specific DNA interaction module.
A hallmark of general transcription factor 3 (GTF3) is the presence of multiple GTF2I-like repeats that were suggested to mediate protein-protein interactions. However, we have recently demonstrated that repeat 4 is necessary and sufficient for binding of GTF3 to the bicoid-like motif of the Troponin I slow enhancer. Given the sequence similarity between different GTF2I-like repeats we hypothesized that DNA binding might be a common property of this domain type. We subjected five repeats of GTF3 to random oligonucleotide selection (SELEX) to assess their DNA binding potentials. We delineated the consensus sequence G(TC)G(A)GATTA(G)BG(A) for repeat 4 and showed that binding sites for GTF3 in enhancers for Troponin I and homeobox c8 (HOXc8) are in very good agreement with this motif. SELEX selections for repeats 5 and 2 enriched for oligonucleotides that were also bound by R4, suggesting that they share common sequence preferences, whereas repeat 3 exhibited relaxed sequence requirements for DNA binding. No binding was observed for repeat 1. We also show that GTF2I-like repeats 4 and 6 of transcription factor II-I (TFII-I) exhibit modest DNA binding properties. Lastly, we identified several amino acids of GTF3 repeat 4 required for high affinity protein-DNA interaction. Based on the ability of many repeats to bind DNA in vitro, we suggest that GTF2I-like domains evolved by duplication and diversification of a prototypic DNA-binding ancestor.
Section on Molecular Neurobiology, NICHD, National Institutes of Health, Bethesda, Maryland 20892, USA. vullhord@mail.nih.gov
Detlef
D
Vullhorst
Andres
A
Buonanno
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004742:Enhancer Elements, Genetic; D019143:Evolution, Molecular; D051379:Mice; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D020449:Repetitive Sequences, Amino Acid; D016415:Sequence Alignment; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D019473:Transcription Factors, TFII / Q000235:genetics*
D004268:DNA-Binding Proteins; C486330:Gtf2i protein, mouse; C486558:Gtf2ird1 protein, mouse; D009124:Muscle Proteins; D009687:Nuclear Proteins; D015534:Trans-Activators; D019473:Transcription Factors, TFII