Caged-degron-based molecular feedback circuits and methods of using the same

A molecular feedback circuit, the circuit comprising:

• a signaling protein that, when activated by an input of a signaling pathway, drives an output of the signaling pathway, wherein the signaling protein comprises a degradation-based switch polypeptide;
• and
• a regulatory nucleic acid sequence responsive to the output and operably linked to a nucleic acid sequence encoding a key polypeptide,
• wherein the output induces expression of the key polypeptide in a eukaryotic cell,
• wherein the degradation-based switch polypeptide comprises an amino acid sequence selected from a group consisting of the amino acid sequences of SEQ ID NOs:
• 1-25,
• wherein the amino acid sequence of the degradation-based switch polypeptide comprises:
• a first domain comprising a degron;
• a second domain comprising five alpha helices;
• and
• a third domain comprising an alpha helix,
• wherein, in the absence of the key polypeptide, the second and third domains form a three dimensional structure comprising a six helix bundle to sequester the degron and prevent the degron from triggering degradation of the signaling protein, and
• wherein the key polypeptide comprises an amino acid sequence selected from a group consisting of the amino acid sequences of SEQ ID NOs:
• 46-49, and
• wherein the amino acid sequence of the key polypeptide comprises an alpha helix that binds to the second domain with a binding affinity that is higher than the binding affinity of the third domain binding to the second domain, and, when expressed in a eukaryotic cell and bound to the second domain, modifies the three dimensional structure of the degradation-based switch polypeptide so that the degron is exposed and can target the signaling protein for degradation, thereby inducing degradation of the signaling protein in the eukaryotic cell.