Controlling Messenger RNA Stability in Bacteria: Strategies for Engineering Gene Expression

Abstract

Recent advances in the understanding of prokaryotic gene expression have led scientists to look beyond traditional promoter control for new methods of regulating gene expression. A promising, new technique centers on controlling the stability of messenger RNA. To exploit the potential of mRNA stability for gene expression control, it is important to understand the mechanisms of prokaryotic mRNA decay as well as the cellular factors that can be used to enhance bacterial gene expression through mRNA stabilization. Factors involved in controlling prokaryotic mRNA stability such as nucleases, secondary structures, translation influences, and transcription effects are discussed and analyzed within the context of three prevailing mRNA decay theories. Several strategies for manipulating mRNA stability in genetically-engineered cells are developed from these discussions and presented as a future direction in gene expression control. In the near future, it should be possible to use these strategies to control mRNA stability in such applications as pharmaceutical protein production and metabolic pathway design.

Rare codons play a positive role in the expression of the stationary phase sigma factor RpoS (σS) inEscherichia coli

“…16,17 With respect to protein product yield and mRNA turnover, the presence of rare codons and pausing ribosomes at these codons can have different and often case-specific consequences. Early on, it was recognized that heterologous protein production in E. coli could often (but not always) be improved by replacing non-optimal codons by E. coli-specific frequent codons, 18,19 because ribosomes stalled at rare codons are thought to allow endonucleolytic attack of the mRNA downstream. 20,21 Yet, ribosome pausing and overall ribosome coverage of transcripts can have very different effects on mRNA turnover.…”

“…Consistent with this notion, the synthesis of heterologous proteins often can be improved when non-optimal codons are replaced by recombinant hostspecific frequent codons. 18,19 Nevertheless, this strategy can also fail completely, with the introduction of frequent codons actually resulting in more rapid mRNA turnover. 25,26 In view of the general notion that rare codons reduce the efficiency of protein synthesis, the question arose why they have been maintained in many genes during evolution.…”

A Secondary Structure in the 5' Untranslated Region ofadhEmRNA Required for RNase G-Dependent Regulation

“…Recently, stabilization of mRNAs has been attempted in order to control gene expression. 42,43) For example, suppression of mRNA degradation has been achieved by ribosome occupation due to enhancement of translation efficiency. 38,44,45) The introduction of hairpin structures into untranslated regions of mRNA, [46][47][48][49] removal of RNase recognition sequences, [50][51][52][53][54] and mutations of RNase 55) have been reported.…”

Transcripts of the genes sacB, amyE, sacC and csn expressed in Bacillus subtilis under the control of the 5′ untranslated sacR region display different stabilities that can be modulated

“…These factors include both gene-specific characteristics and bulk cellular conditions (Carrier & Kiesling, 1997), and can be divided into several categories which we will examine below.…”