The mRNA for an inducible chloramphenicol acetyltransferase gene is cleaved into discrete fragments in Bacillus subtilis

Abstract

cat-86 is a promoter-deficient plasmid gene that encodes chloramphenicol acetyltransferase (CAT). Insertion of a promoter at a site 144 base pairs 5' to the cat-86 coding sequence activates transcription of the gene and allows cat-86 to specify chloramphenicol-inducible CAT activity in Bacillus subtilis. Induction of cat-86 by chloramphenicol has been shown to result from a regulatory event that activates translation of cat-86 mRNA that is present in cells before the addition of inducer (E. J. Duvall and P. S. Lovett, Proc. Natl. Acad. Sci. USA 83:3939-3943, 1986). In the present study we show an unusual property of cat-86 mRNA. Full-length cat-86 transcripts, consisting of 920 nucleotides (nt), are cleaved in B. subtilis to yield two predominant fragmentation products: an 810-nt species that lacks sequences present at the 5' end of the 920-nt species and a 720-nt species that lacks sequences present at the 3' end of the 920-nt species. A third fragmentation product consisting of 620 nt may result from the cleavage of a single 920-nt transcript at both the 5' and 3' ends. The sequences which are missing from the 720- and 620-nt species suggest that these transcripts cannot be translated into functional CAT. The 810-nt species lacks sequences from the 5' regulatory region, and it is not yet certain whether or not translation of this species can be induced by chloramphenicol. The ratio of 920-nt molecules/720-nt molecules in rifampin-treated cells is increased when the cells are grown in chloramphenicol. Therefore, induction may partially stabilize full-length cat-86 transcripts against inactivation by a novel processing-like system.