reporter gene usually a selectable marker or Lacz, or both This bait lies in wait for an interaction with another protein. The gAL4 AD, is fused to either protein Y or protein Z. Should either one of these proteins be able to interact with protein X then the Gal4 AD region will become tethered to the UASGAL region and will recruit and activate the rna polymerase Gal4 DNA Binding Domain Protein x GAL4-binding site Reporter gene Protein y Gal4 Activation Positive interaction (Gal4-AD) Increased transcripti Protein z Gal4 Activation Domain GAL4-binding site Gal4 Chimeric Proteins can Interrogate Yeast Two-Hybrid Assay for Protein Protein-Protein interactions Protein Interactions Figure by MIT OCW Note that the protein x,y and z do not have to be yeast proteins the only requirement is that the dna coding sequence for the protein is available(which is now true for all of the genes from a wide variety of organisms); these sequences are then cloned such that they produce the appropriate Gal4 chimeric proteins In the previous two three lectures we have looked at one particular regulatory network in S. cerevisiae, and have employed a wide range of tools to understand this network. In the next lecture I will be telling you how these and other tools have evolved into technologies that allow us to look globally at gene regulation in eukaryotic cellsreporter gene, usually a selectable marker or LacZ, or both. This bait lies in wait for an interaction with another protein. The GAL4 AD, is fused to either protein Y or protein Z. Should either one of these proteins be able to interact with protein X then the Gal4 AD region will become tethered to the UASGAL region and will recruit and activate the RNA polymerase. Note that the protein X, Y and Z do not have to be yeast proteins; the only requirement is that the DNA coding sequence for the protein is available (which is now true for all of the genes from a wide variety of organisms); these sequences are then cloned such that they produce the appropriate Gal4 chimeric proteins. In the previous two three lectures we have looked at one particular regulatory network in S. cerevisiae, and have employed a wide range of tools to understand this network. In the next lecture I will be telling you how these and other tools have evolved into technologies that allow us to look globally at gene regulation in eukaryotic cells. No interaction Gal4-BD x Y Gal4-AD Gal4 Chimeric Proteins can Interrogate Protein-Protein Interactions Yeast Two-Hybrid Assay for Protein￾Protein Interactions Protein Y Gal4 Activation Domain Gal4 DNA Binding Domain Protein X Protein Z Gal4 Activation Domain A GAL4-binding site Positive interaction Increased transcription Gal4-BD Gal4-AD GAL4-binding site x z Reporter gene Reporter gene B Figure by MIT OCW