Mutations affecting mRNA processing and fimbrial biogenesis in the Escherichia coli pap operon

Abstract

The Escherichia coli pap genetic determinant includes 11 genes and encodes expression of Pap pili on the bacterial surface. An RNase E-dependent mRNA-processing event in the intercistronic papB-papA region results in the accumulation of a papA-gene-specific mRNA in considerable excess of the primary papB-papA mRNA transcription product. We have introduced mutations in the intercistronic region and studied the effect in vivo of these mutations on the processing event, PapA protein expression, and the biogenesis of fimbriae on the bacterial surface. Our studies establish that mRNA processing is an important event in the mechanism resulting in differential gene expression of the major pap operon. The deletion of sequences corresponding to the major cleavage site abolished processing, reduced expression of PapA protein, and resulted in "crew-cut" bacteria with short fimbrial structures on the bacterial surface. Only a limited part of the intercistronic region appeared to be required as the recognized target for the processing to occur. Upstream sequences to a position within 10 nucleotides of the major RNase E-dependent cleavage site could be deleted without any detectable effect on papB-papA mRNA processing, PapA protein expression, or fimbria formation. Substitution mutations of specific bases at the cleavage site by site-directed mutagenesis showed that there were alternative positions at which cleavage could be enhanced, and tests with an in vitro processing assay showed that such cleavages were also RNase E dependent. Our findings are discussed in relation to other fimbrial operons and other known targets of the RNase E endoribonuclease.