Fig 8.

Reverse two-hybrid systems. (A) Repression of toxic genes (228, 373, 667). Interaction between bait X and prey Y stimulates transcription of URA3, which results in cell toxicity in the presence of 5-fluoroorotic acid; CYH2, which makes the cells sensitive to cycloheximide; or GAL1, which converts galactose into galactose-1-phosphate, a toxic compound that accumulates in the absence of Gal7. Compounds or mutations (in X or Y) that interfere with the association of the bait with the prey are identified by growth on selective medium. (B) Activation of positive selection markers (589). Interaction between bait X and prey Y leads to expression of tetR, encoding the Tet repressor. Next, TetR represses activation of HIS3, hindering cell growth on medium without histidine. Disruption of the interaction between bait and prey is analyzed by the appearance of histidine prototrophy. DBD, DNA-binding domain; AD, activaion domain; UAS, upstream activating sequence; Pol, polymerase; Op, operator.