Mutational Analysis of a RNase E Dependent Cleavage Site from a Bacteriophage T4 mRNA

Recently E.coli endoribonuclease RNase E has been implicated in the functional and chemical decay of many bacteriophage T4 mRNAs. We have sought to identify the features of the phage RNA sequence that are necessary for RNase E dependent cleavage. A 117 bp synthetic sequence that contains the RNase E dependent processing site from the 5′ leader of T4 gene 32 was assembled and inserted into pUC18. Correct RNase E dependent processing of a hybrid transcript containing this sequence was demonstrated in vivo. Mutations which reduce processing at this site have been obtained by in vitro mutagenesis