Abstract
ETS1 is the archetype of the ETS transcription factor (TF) family. ETS TFs share a DNA-binding domain, the ETS domain. All ETS TFs recognize a core GGA(A/T) binding site, and thus ETS TFs are found to redundantly regulate the same genes. However, each ETS TF has unique targets as well. One prevailing hypotheses explaining this duality is that protein-protein interactions, including homodimerization, allow each ETS TF to display distinct behavior. The behavior of ETS1 is further regulated by autoinhibition. Autoinhibition apparently modulates ETS1 DNA binding affinity, but the mechanism of this inhibition is not completely understood. We sought to characterize the relationship between DNA binding and ETS1 homodimer formation. We find that ETS1 interrogates DNA and forms dimers even when the DNA does not contain an ETS recognition sequence. Mutational studies also link nonspecific DNA backbone contacts with dimer formation, in addition to providing a new role for the recognition helix of ETS1 in maintaining the autoinhibited state. Finally, in showing that residues in the DNA recognition helix affect autoinhibition, we define a new function of ETS1 autoinhibition: maintenance of a monomeric state in the absence of DNA. The conservation of relevant amino acid residues across all ETS TFs indicates that the mechanisms of nonspecific DNA interrogation and protein oligomer formation elucidated here may be common to all ETS proteins that autoinhibit.
Highlights
The transcription factor ETS1 controls its DNA binding activity through an autoinhibitory module
We find that when increasing concentrations of ETS1⌬280 are added to radiolabeled DNA containing either the MMP3 or p53 ETS binding sites (EBS), ETS1⌬280 forms a single complex with each DNA
Consistent with the suggestion that the identity of the bases that flank the EBS core consensus sequence are important in determining ETS1 DNA affinity, we find that despite having identical core sequences, the dissociation constants (KD) of the complexes between ETS1⌬280 and MMP3 or p53 differ by 30-fold (Table 1)
Summary
The transcription factor ETS1 controls its DNA binding activity through an autoinhibitory module. Mutational studies link nonspecific DNA backbone contacts with dimer formation, in addition to providing a new role for the recognition helix of ETS1 in maintaining the autoinhibited state. We are interested in exploring how binding site sequence affects oligomeric form and autoinhibitory function together to regulate the DNA binding activities of transcription factors. To this end, because there is an exact correspondence between DNA binding affinity and transcriptional regulatory activity of ETS family members [1], we are investigating the form and function of ETS1, the prototypical ETS protein. Autoinhibition was previously thought to only involve allosteric hindrance of a separate region of the ETS domain
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