Most riboswitches are metabolite-binding RNA structures located in bacterial messenger RNAs where they control gene expression. We have discovered a riboswitch class in many bacterial and archaeal species whose members are selectively triggered by fluoride but reject other small anions, including chloride. These fluoride riboswitches activate expression of genes that encode putative fluoride transporters, enzymes that are known to be inhibited by fluoride, and additional proteins of unknown function. Our findings indicate that most organisms are naturally exposed to toxic levels of fluoride and that many species use fluoride-sensing RNAs to control the expression of proteins that alleviate the deleterious effects of this anion.
Group I self-splicing ribozymes commonly function as components of selfish mobile genetic elements. We identified an allosteric group I ribozyme, wherein self-splicing is regulated by a distinct riboswitch class that senses the bacterial second messenger c-di-GMP. The tandem RNA sensory system resides in the 5′ untranslated region of the messenger RNA for a putative virulence gene in the pathogenic bacterium Clostridium difficile. c-di-GMP binding by the riboswitch induces folding changes at atypical splice site junctions to modulate alternative RNA processing. Our findings indicate that some self-splicing ribozymes are not selfish elements, but are harnessed by cells as metabolite sensors and genetic regulators.
Outer membrane vesicles (OMVs) are nanoscale proteoliposomes that are ubiquitously secreted by Gram-negative bacteria. Interest in these bioparticles has escalated over the years, leading to discoveries regarding their composition, production, and vaccine potential. Given that many steps in vesicle biogenesis are ‘engineerable,’ it is now possible to tailor OMVs for specific applications. Such tailoring involves modifying the OMV-producing bacterium through protein, pathway, or genome engineering in a manner that specifically alters the final OMV product. For example, targeted deletion or upregulation of genes associated with the cell envelope can modulate vesicle production or remodel the composition of vesicle components such as lipopolysaccharide. Likewise, bacteria can be reprogrammed to incorporate heterologously expressed proteins into either the membrane or lumenal compartment of OMVs. We anticipate that further research in the field of OMV engineering will enable continued design and biosynthesis of specialized vesicles for numerous biotechnological purposes ranging from the delivery of vaccines to the deconstruction of cellulosic substrates.
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