ANALYSIS OF THE SECONDARY STRUCTURE THAT NEGATIVELY REGULATES INDUCIBLE CAT TRANSLATION BY USE OF CHEMICAL PROBING AND MUTAGENESIS

Induction of chloramphenicol (Cm) acetyltransferase-encoding genes (ca t) by Cm has been proposed to result from the destabilization of a ste m-loop that sequesters the ribosome-binding site for the cat coding se quence. Destabilization is caused by the stalling of a ribosome at a s pecific site in the leader of cat transcripts that immediately precede s the stem-loop. By use of in vivo dimethylsulfate probing of cat-86 l eader mRNA, we demonstrate the existence of the stem-loop structure in cat transcripts isolated from uninduced cells and its release during induction. Leader mutations chosen to provide the mRNA with an alterna tive-folding pattern that destabilizes the stem-loop cause constitutiv e cat expression. Our results establish the occurrence in vivo of the stem-loop in cat-86 transcripts and its role as a negative regulator o f cat expression.