Imagine that you are interested in a protein that interacts with proteins of the cytoskeleton in human epithelial cells. Describe an experimental protocol based on the yeast two-hybrid system that would allow you to identify proteins that might interact with your protein of interest.

To identify proteins interacting with the protein of interest through the yeast two-hybrid system, first clone the gene encoding the protein of interest into the DNA-binding domain of a hybrid protein in a yeast two-hybrid vector. Prepare a yeast two-hybrid library of cDNAs from human epithelial cells, clone these into the activation domain of another hybrid protein in another vector. Co-transform these vectors into yeast cells, and select for interactions (which activate a reporter gene) by growing the cells on selective agar plates. The protein interactions can be determined by sequencing the cDNA associated with growing colonies of yeast. Validate these interactions through reciprocal assays or other suitable methods.