Journal Article

Ribosomal frameshifting and dual-target antiactivation restrict quorum-sensing–activated transfer of a mobile genetic element

Tester, Laura G. L., Kleffmann, Torsten, Edgar, Christina D., Ramsay, Joshua P., Patterson-House, Jackson R., Tate, Warren P., Hynes, Michael F., Major, Anthony S., Ronson, Clive W., Sullivan, John T., Hall, Drew A.

2015

Overview

Symbiosis islands are integrative and conjugative mobile genetic elements that convert nonsymbiotic rhizobia into nitrogen-fixing symbionts of leguminous plants. Excision of the Mesorhizobium loti symbiosis island ICE Ml Sym ᴿ⁷ᴬ is indirectly activated by quorum sensing through TraR-dependent activation of the excisionase gene rdfS . Here we show that a +1 programmed ribosomal frameshift (PRF) fuses the coding sequences of two TraR-activated genes, msi172 and msi171 , producing an activator of rdfS expression named Frameshifted excision activator (FseA). Mass-spectrometry and mutational analyses indicated that the PRF occurred through +1 slippage of the tRNA ᵖʰᵉ from UUU to UUC within a conserved msi172 -encoded motif. FseA activated rdfS expression in the absence of ICE Ml Sym ᴿ⁷ᴬ, suggesting that it directly activated rdfS transcription, despite being unrelated to any characterized DNA-binding proteins. Bacterial two-hybrid and gene-reporter assays demonstrated that FseA was also bound and inhibited by the ICE Ml Sym ᴿ⁷ᴬ-encoded quorum-sensing antiactivator QseM. Thus, activation of ICE Ml Sym ᴿ⁷ᴬ excision is counteracted by TraR antiactivation, ribosomal frameshifting, and FseA antiactivation. This robust suppression likely dampens the inherent biological noise present in the quorum-sensing autoinduction circuit and ensures that ICE Ml Sym ᴿ⁷ᴬ transfer only occurs in a subpopulation of cells in which both qseM expression is repressed and FseA is translated. The architecture of the ICE Ml Sym ᴿ⁷ᴬ transfer regulatory system provides an example of how a set of modular components have assembled through evolution to form a robust genetic toggle that regulates gene transcription and translation at both single-cell and cell-population levels. Significance Integrative and conjugative elements (ICEs) facilitate horizontal transfer of multiple genetic determinants. Here we show that a programmed ribosomal frameshift (PRF) contributes to the regulation of ICE transfer. The low-frequency PRF fuses the coding sequences of two genes, resulting in a single-protein Frameshifted excision activator (FseA) that activates ICE excision. An antiactivator, QseM, known to disrupt the quorum-sensing regulator TraR, also disrupted FseA. The evolved PRF site, together with the dual-target antiactivator, QseM, likely provides robust suppression of ICE transfer in the face of the inherent biological noise of quorum-sensing autoinduction. This work illustrates how a complex multipartite regulatory system has assembled through evolution to form a robust genetic toggle to control gene transcription and translation at both single-cell and cell-population levels.

• Publisher: National Academy of Sciences
• Subject: Bacteria / Base Sequence / beta-Galactosidase - metabolism / Binding Sites / Biological Sciences / Cells / evolution / excision / Frameshifting, Ribosomal / Gene expression / Gene Transfer Techniques / genes / Genetics / Genomic Islands / Interspersed Repetitive Sequences / Mass Spectrometry / Mesorhizobium - metabolism / Mesorhizobium loti / Microbiology / Mutation / Nitrogen fixation / Plants - microbiology / Plasmids - metabolism / Population levels / Promoter Regions, Genetic / Protein Biosynthesis / Proteins / Quorum Sensing / Rhizobium - metabolism / Ribonucleic acid / Ribosomes - chemistry / Ribosomes - ultrastructure / RNA / Spectrometry / Symbiosis / transcription (genetics) / Transcription Factors / Transcription, Genetic / translation (genetics) / Two-Hybrid System Techniques