AU-Rich Sequences within 5′ Untranslated Leaders Enhance Translation and Stabilize mRNA in Escherichia coli

Abstract

We have shown previously that when the Escherichia coli chromosomal lacZ gene is put under the control of an extended Shine-Dalgarno (SD) sequence (10 or 6 nucleotides in length), the translation efficiency can be highly variable, depending on the presence of AU-rich targets for ribosomal protein S1 in the mRNA leader. Here, the same strains have been used to examine the question of how strong ribosome binding to extended SD sequences affects the stability of lacZ mRNAs translated with different efficiencies. The steady-state concentration of the lacZ transcripts has been found to vary over a broad range, directly correlating with translation efficiency but not with the SD duplex stability. The observed strain-to-strain variations in lacZ mRNA level became far less marked in the presence of the rne-1 mutation, which partially inactivates RNase E. Together, the results show that (i) an SD sequence, even one that is very long, cannot stabilize the lacZ mRNA in E. coli if translation is inefficient; (ii) inefficiently translated lacZ transcripts are sensitive to RNase E; and (iii) AU-rich elements inserted upstream of a long SD sequence enhance translation and stabilize mRNA, despite the fact that they constitute potential RNase E sites. These data strongly support the idea that the lacZ mRNA in E. coli can be stabilized only by translating, and not by stalling, ribosomes.As a rule, prokaryotic mRNAs are unstable, with half-lives in the minute range. In Escherichia coli, pathways of mRNA decay are complex and include an initial rate-limiting endonucleolytic cleavage (generally mediated by RNase E) followed by 3Ј35Ј exonucleolytic degradation of the cleavage products (reviewed in references 6, 10, 19, and 34). For several mRNAs, decay pathways have been studied in detail, allowing molecular models to be formulated (4-6, 11, 13, 28). Several lines of evidence suggest that translating ribosomes usually protect the mRNA from endonucleolytic attacks (10). Much of the available information has been derived from experiments with lacZ translational fusions. In this case, mRNA translation and degradation are intimately correlated. In particular, mutations in the ribosome binding site (RBS) which reduce translation initiation efficiency, and hence efficiency of the overall translation, accelerate mRNA degradation (14,22,38). However, despite all efforts, the mechanism of the lacZ mRNA decay still remains poorly characterized.Although in many cases translation efficiency positively correlates with stability of the lacZ mRNA in E. coli, the question of whether merely the ribosome binding to the RBS is able to stabilize the whole transcript remains equivocal. On one hand, it has been suggested that a ribosome bound to a strong ShineDalgarno (SD) sequence, but not translation per se, could be the main stability factor for the lacZ mRNA (37). This would imply that E. coli mRNAs can be protected from degradation by the same mechanism as in bacilli, where ribosome stalling within 5Ј untranslated regions can protect kilobases of the...

Noncanonical repression of translation initiation through small RNA recruitment of the RNA chaperone Hfq

“…However, to our knowledge, it is the first time that Hfq has been shown to be directly involved in 30S ribosomes competition in the context of sRNA-mediated gene regulation. Given the marked preference of Hfq for AU-rich regions and the fact that these regions are known to act as translational enhancers (Zhang and Deutscher 1992;Komarova et al 2005;Hook-Barnard et al 2007), we believe that Hfq may play a similar role in the action of many other sRNAs that are not pairing directly to the TIR and are therefore not able to directly compete with initiat- Figure 8. Spot42 binds to sdhCDAB in vivo and recruits Hfq to sdhC TIR.…”

Dynamic evolution of translation initiation mechanisms in prokaryotes

“…One is translation initiation mediated by a ribosomal protein S1 (RPS1), which is a component of the 30S ribosomal subunit. In Escherichia coli, RPS1 interacts with a 5′ UTR of an mRNA, initiating translation efficiently, regardless of the presence of the SD sequence (14,15). RPS1 of E. coli contains six S1 domains that are essential for RNA binding, although the number of domains is different among species (16).…”

Characterization of Riemerella anatipestifer CH-1 gldJ gene and GldJ protein

“…The AU-rich sequence can enhance and stabilize mRNA (Komarova et al, 2005). Moreover, the sequence can be bound by ribosomal protein S1 to instigate translation initiation in the case of mRNA (Komarova et al, 2005;Nakagawa et al, 2010).…”

“…The TATA promoter region is the binding site of general transcription factors, and it is critical for transcription initiation and veracity (Yang et al, 2007). The AU-rich sequence can enhance and stabilize mRNA (Komarova et al, 2005). Moreover, the sequence can be bound by ribosomal protein S1 to instigate translation initiation in the case of mRNA (Komarova et al, 2005;Nakagawa et al, 2010).…”