Fig. 4.

Effect of the sequence of the first three RNA nucleotides on RppH-dependent mRNA degradation in B. subtilis. The half-lives of mini yhxA-glpP mRNA and derivatives thereof with a substitution at the first, second, or third position were compared in isogenic B. subtilis cells containing or lacking an rppH gene. The numeral in the name of each variant indicates the nucleotide position (1, 2, or 3), and the letter indicates the nucleotide identity (A, G, C, or U). In every case, the identity of the unspecified nucleotides was the same as in the 5′ UTR of the wild-type transcript: A at position 1, G at position 2, and G at position 3; therefore, 1A, 2G, and 3G are all equivalent to mini yhxA-glpP with a wild-type 5′ UTR. (A) Representative time courses. The decay of mini yhxA-glpP mRNA with G or U at position 2 (2G or 2U) was monitored by Northern blot analysis of RNA extracted from wild-type or ∆rppH cells at time intervals after arresting transcription with rifampicin. Band intensities were plotted semilogarithmically as a function of time, and best-fit lines were calculated by linear regression. (B) Half-lives. Each mRNA half-life is the average of two or more independent measurements. Error bars correspond to SDs.