Feedback regulation of the spc operon in Escherichia coli: translational coupling and mRNA processing

Abstract

The spc operon of Escherichia coli encodes 10 ribosomal proteins in the order L14, L24, L5, S14, S8, L6, L18, S5, L30, and L15. This operon is feedback regulated by S8, which binds near the translation start site of L5 and inhibits translation of L5 directly and that of the distal genes indirectly. We constructed plasmids carrying a major portion of the spc operon genes under lac transcriptional control. The plasmids carried a point mutation in the S8 target site which abolished regulation and resulted in overproduction of plasmid-encoded ribosomal proteins upon induction. We showed that alteration of the AUG start codon of L5 to UAG decreased the synthesis rates of plasmid-encoded distal proteins, as well as L5, by approximately 20-fold, with a much smaller (if any) effect on mRNA synthesis rates, indicating coupling of the distal cistrons' translation with the translation of L5. This conclusion was also supported by experiments in which S8 was overproduced in trans. In this case, there was a threefold reduction in the synthesis rates of chromosome-encoded L5 and the distal spc operon proteins, but no decrease in the mRNA synthesis rate. These observations also suggest that transcription from ribosomal protein promoters may be special, perhaps able to overcome transcription termination signals. We also analyzed the state of ribosomal protein mRNA after overproduction of S8 in these experiments and found that repression of ribosomal protein synthesis was accompanied by stimulation of processing (and degradation) of spc operon mRNA. The possible role of mRNA degradation in tightening the regulation is discussed.