Messenger RNA degradation in bacterial cells

Abstract

mRNA degradation is an important mechanism for controlling gene expression in bacterial cells. This process involves the orderly action of a battery of cellular endonucleases and exonucleases, some universal and others present only in certain species. These ribonucleases function with the assistance of ancillary enzymes that covalently modify the 5' or 3' end of RNA or unwind base-paired regions. Triggered by initiating events at either the 5' terminus or an internal site, mRNA decay occurs at diverse rates that are transcript specific and governed by RNA sequence and structure, translating ribosomes, and bound sRNAs or proteins. In response to environmental cues, bacteria are able to orchestrate widespread changes in mRNA lifetimes by modulating the concentration or specific activity of cellular ribonucleases or by unmasking the mRNA-degrading activity of cellular toxins.

Keywords: gene regulation; mRNA stability; ribonuclease; sRNA; translation.

Figure 1. Constituent domains of mRNA-degrading ribonucleases

The domain composition of representative ribonucleases from E. coli (RNase E, RNase III, PNPase, RNase II, and RNase R) and B. subtilis (RNase Y and RNase J) is shown. Structural domains are depicted as colored rectangles: red, catalytic domain; blue, RNA-binding domain; yellow, protein-binding domain; green, membrane-binding domain; gray, miscellaneous domain. The sites where RhlB, enolase (Eno), and PNPase (PNP) bind to RNase E are marked, as are its two arginine-rich RNA-binding domains (AR). Of the two PH domains in PNPase, only the second is catalytically active. The single metallo-β-lactamase domain (MβL) of RNase J comprises two noncontiguous segments of the polypeptide. CC, coiled coil domain; dsRBD, double-stranded RNA-binding domain; CSD, cold shock domain; HTH, helix-turn-helix domain; CTD, carboxy-terminal domain. The catalytic domains of RNase E and RNase III have not been named.

Figure 2. Direct-access pathway for mRNA degradation

An endonuclease (black-handled scissors), usually but not always RNase E or RNase Y, cleaves the primary transcript internally to generate two fragments. Unprotected at its 3’ end, the 5’ fragment is quickly attacked by 3’ exonucleases (blue Pac-Man) in all bacterial species. The fate of the monophosphorylated 3’ fragment depends on the ribonucleases present in the cell. In some species, like E. coli, this fragment undergoes further endonucleolytic cleavage by RNase E (gray scissors), which rapidly degrades such intermediates by selectively binding the monophosphorylated 5’ terminus in a discrete pocket on the surface of the catalytic domain and cutting downstream. In others, like B. subtilis, it undergoes rapid 5’-exonucleolytic digestion by RNase J (red Pac-Man), whose exonuclease activity aggressively degrades RNAs bearing a single phosphate at the 5’ end.

Figure 3. 3’-exonucleolytic degradation of decay intermediates

Endonucleolytic cleavage of mRNA generates a 5’-terminal fragment whose single-stranded 3’ end is trimmed exonucleolytically until a structural barrier is encountered, as well as a 3’-terminal fragment whose 3’ end is protected from exonucleolytic digestion by a terminator stem-loop. Subsequent polyadenylation by poly(A) polymerase or PNPase provides the 3’ exonucleases PNPase (operating with help from the RNA helicase RhlB) and RNase R (operating alone) an opportunity to overcome the barriers by creating a single-stranded binding site from which they can launch an attack. The process of poly(A) addition and removal is repeated until the structural barriers are breached. By contrast, RNase II can degrade poly(A) and other unpaired 3’ ends but not structured 3’ ends. Degradation by these 3’ exonucleases (blue Pac-Man) eventually generates fragments that are too small for them to shorten further and are instead degraded by an oligoribonuclease (yellow Pac-Man).

Figure 4. 5’-end-dependent pathway for initiating mRNA degradation

The RNA pyrophosphohydrolase RppH (hatchet) converts the 5’-terminal triphosphate of the primary transcript to a monophosphate. The resulting full-length decay intermediate is then rapidly degraded by either RNase E (scissors) or RNase J (Pac-Man), depending on which of these enzymes is present in the host species.