A HAMMERHEAD RIBOZYME INHIBITS ADE1 GENE-EXPRESSION IN YEAST

To study factors that affect in vivo ribozyme (Rz) activity, a model s ystem has been devised in Saccharomyces cerevisiae based on the inhibi tion of ADE1 gene expression. This gene was chosen because Rz action c an be evaluated visually by the Red phenotype produced when the activi ty of the gene product is inhibited. Different plasmid constructs allo wed the expression of the Rz either in cis or in trans with respect to ADE1. Rz-related inhibition of ADE1 expression was correlated with a Red phenotype and a diminution of ADE1 mRNA levels only when the Rz ge ne was linked 5' to ADE1. The presence of the expected 3' cleavage fra gment was demonstrated using a technique combining RNA ligation and PC R. This yeast system and detection technique are suited to the investi gation of general factors affecting Rz-catalyzed inhibition of gene ex pression under in vivo conditions.