RNA processing and degradation in Bacillus subtilis

Abstract

This review focuses on the enzymes and pathways of RNA processing and degradation in Bacillus subtilis, and compares them to those of its gram-negative counterpart, Escherichia coli. A comparison of the genomes from the two organisms reveals that B. subtilis has a very different selection of RNases available for RNA maturation. Of 17 characterized ribonuclease activities thus far identified in E. coli and B. subtilis, only 6 are shared, 3 exoribonucleases and 3 endoribonucleases. Some enzymes essential for cell viability in E. coli, such as RNase E and oligoribonuclease, do not have homologs in B. subtilis, and of those enzymes in common, some combinations are essential in one organism but not in the other. The degradation pathways and transcript half-lives have been examined to various degrees for a dozen or so B. subtilis mRNAs. The determinants of mRNA stability have been characterized for a number of these and point to a fundamentally different process in the initiation of mRNA decay. While RNase E binds to the 5' end and catalyzes the rate-limiting cleavage of the majority of E. coli RNAs by looping to internal sites, the equivalent nuclease in B. subtilis, although not yet identified, is predicted to scan or track from the 5' end. RNase E can also access cleavage sites directly, albeit less efficiently, while the enzyme responsible for initiating the decay of B. subtilis mRNAs appears incapable of direct entry. Thus, unlike E. coli, RNAs possessing stable secondary structures or sites for protein or ribosome binding near the 5' end can have very long half-lives even if the RNA is not protected by translation.

FIG. 1.

Domain structure of potential B. subtilis RNases. aa, amino acids. Domain abbreviations: KH, ribonucleoprotein K homology domain; HD, His-Asp-containing domain of phosphoesterases; N-OB, oligonucleotide binding fold; Thermo, thermonuclease domain; Phospho-C, phosphatase C domain; RBN, RNase BN; Bla, β-lactamase domain.

FIG. 2.

Model of ermC regulation by translational attenuation and mRNA stabilization. The proposed structure of the ermC leader and the position of the leader peptide and ribosome- stalling site are shown to the left. SD1 and SD2 refer to the SD sequences of the leader peptide and ermC methylase, respectively. In the presence of erythromycin, ribosomes stalled on the leader peptide open up the RNA structure and allow ribosome access to SD2. At the same time, the stalled ribosomes protect the transcript from decay.

FIG. 3.

Sequence and proposed secondary structure of the A-region at the 5′ end of phage SP82. The sites of RNase III cleavage and PNPase stalling (see the text) are indicated. Also shown is the polypurine sequence that functions as a 5′ stabilizer even in the absence of translation initiation at the AUG codon. Bs-RNase III, B. subtilis RNase III.

FIG. 4.

Sequence and proposed secondary structure of the aprE 5′ stabilizer. The SD sequence and initiation codon are in bold capital letters.

FIG. 5.

Model for regulation of thrS gene expression. Uncharged tRNA^Thr interacts with the thrS leader to pull the leader into the antitermination conformation. Specificity for tRNA^Thr is afforded by the ACC codon bulged out of the specifier domain. The antiterminator is stabilized by Watson-Crick base pairing between the CCA end of the tRNA and the complementary sequence bulged out of the antiterminator. Cleavage (scissors) in the loop of the antiterminator structure results in a shorter RNA flanked by factor-independent transcription terminators, which is much more stable than the full-length transcript.

FIG. 6.

Structure and transcripts of the gapA operon. The primary (1°) and secondary (2°) transcripts (wavy lines) are indicated. Cleavage (scissors) of either the bicistronic or hexacistronic transcript just upstream of the secondary structure indicated results in shorter, stable 3′ fragments and an unstable cggR transcript. The relative abundance of the different transcripts is reflected in the relative thickness of the wavy lines.

FIG. 7.

Comparison of the modes of action of E. coli RNase E and the hypothetical 5′-end-dependent RNase of B. subtilis. RNase E can bind to the 5′ end of transcripts (wavy lines) and loop to internal sites (thick portions) or can access cleavage sites directly. Accumulated evidence suggests that the B. subtilis enzyme is strictly 5′ end dependent, i.e., cannot access sites directly, and tracks along the RNA in the 5′-to-3′ direction to find cleavage sites. Thus, while active translation is required to inhibit cleavage by RNase E, static ribosomes can inhibit cleavage by the B. subtilis equivalent.