Regulation of gene expression using single-chain, monomeric, ligand dependent polypeptide switches
AssigneeThe Scripps Research Institute
InventorsCarlos F. Barbas, III, Roger Beerli, Ulrich Schopfer
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Abstract
Single chain, monomeric polypeptide gene switches are provided. The gene switches include ligand binding domains and at least one functional domain. Preferred functional domains are DNA binding domains and transcriptional regulating domains. Methods of regulating gene function using the switches are also provided.
Claims102
An isolated chimeric polypeptide comprising a first functional domain of a protein which binds a selected DNA sequence, wherein the first functional domain is operatively linked to the N-terminus of two ligand binding domains from nuclear hormone receptors, wherein the DNA binding domain comprises at least one zinc finger DNA binding motif, wherein the chimeric polypeptide binds the ligands bound by the ligand binding domains of the nuclear hormone receptors and binds the selected DNA sequence bound by the first functional domain of the protein.
An isolated chimeric polypeptide comprising:
- (a) a DNA binding domain having from two to six zinc finger DNA binding motifs which domain binds a selected DNA sequence;
- (b) a first ligand binding domain from a retinoid X receptor operatively linked to the DNA binding domain, and a second ligand binding domain from an ecdysone receptor operatively linked to the first ligand binding domain with a peptide spacer of from 18 to 36 amino acid residues;
- and (c) a transcriptional regulating domain operatively linked to the second ligand binding domain, wherein the chimeric polypeptide binds the selected DNA sequence, binds a ligand bound by the retinoid X receptor, binds a ligand bound by the ecdysone receptor, and transcriptionally regulates DNA that is operatively linked to the selected DNA sequence as a result of the presence of the transcriptional regulating domain in the chimeric polypeptide.
An isolated chimeric polypeptide comprising:
- (a) a DNA binding domain having from three to six zinc finger DNA binding motifs which domain binds a selected DNA sequence;
- (b) a first ligand binding domain from a progesterone receptor operatively linked to the DNA binding domain, and a second ligand binding domain from a progesterone receptor linked to the first ligand binding domain with a peptide spacer of from 18 to 36 amino acid residues;
- and (c) a transcriptional regulating domain operatively linked to the second ligand binding domain, wherein the chimeric polypeptide binds the selected DNA sequence, binds a ligand bound by progesterone receptor, and transcriptionally regulates DNA that is operatively linked to the selected DNA sequence as a result of the presence of the transcriptional regulating domain in the chimeric polypeptide.
An isolated chimeric polypeptide comprising a first functional domain of a protein which binds a selected DNA sequence, wherein the first functional domain is operatively linked to the N-terminus of two ligand binding domains for the same ligand, wherein at least one of the ligand binding domains is from a nuclear hormone receptor, wherein the chimeric polypeptide binds the ligands bound by the ligand binding domains and binds the selected DNA sequence bound by the first functional domain of the protein.
Dependencies are inferred from the claim text.