Effect of 5′-proximal elements on decay of a model mRNA in Bacillus subtilis

Previous work showed that a 42-nucleotide sequence from an SP82 bacteriophage early RNA functions as a 5' mRNA stabilizer in Bacillus subtilis. Real-time reverse transcriptase polymerase chain reaction (RT-PCR) analysis of decay of a model mRNA with alterations at the 5'-end was used to elucidate the mechanism of SP82-mediated stability. A predicted 5'-terminal stem-loop structure was essential for stabilization. Increasing the strength of the 5'-terminal structure above a minimum level did not result in increased stability. A thorough analysis of the context in which the stabilizing structure occurred included the effects of distance from 5'-end, translation of downstream coding sequence, and distance between the secondary structure and the ribosome binding site. Our data are consistent with the dominant mRNA decay pathway in B. subtilis being 5'-end dependent.