The glycine riboswitch is a known regulatory element that is unique in having two aptamers that are joined by a linker region. In this study, we investigated a glycine riboswitch located in the 5' untranslated region of a glycine cleavage system homolog (gcvTHP) in Burkholderia spp. Structure prediction using the sequence generated a model with a glycine binding pocket composed of base-triple interactions (G62-A64-A86 and G65-U84-C85) that are supported by A/G minor interactions (A17-C60-G88 and G16-C61-G87, respectively) and two ribose-zipper motifs (C11-G12 interacting with A248-A247 and C153-U154 interacting with A79-A78) which had not been previously reported. The capacity of the riboswitch to bind to glycine was experimentally validated by native gel assays and the crucial role of interactions that make up the glycine binding pocket were proven by mutations of A17U and G16C which resulted in conformational differences that may lead to dysfunction. Using glycine supplemented minimal media, we were able to prove that the expression of the gcvTHP genes found downstream of the riboswitch responded to the glycine concentrations introduced thus confirming the role of this highly conserved Burkholderia riboswitch and its associated genes as a putative glycine detoxification system in Burkholderia spp.
The Burkholderia genus includes many species that are known to survive in diverse environmental conditions including low nutrient environments. One species, Burkholderia pseudomallei is a versatile pathogen that can survive in a wide range of hosts and environmental conditions. In this study, we investigated how a nutrient depleted growth environment evokes sRNA mediated responses by B. pseudomallei. Computationally predicted B. pseudomallei D286 sRNAs were mapped to RNA-sequencing data for cultures grown under two conditions: (1) BHIB as a nutrient rich media reference environment and (2) M9 media as a nutrient depleted stress environment. The sRNAs were further selected to identify potentially cis-encoded systems by investigating their possible interactions with their flanking genes. The mappings of predicted sRNA genes and interactions analysis to their flanking genes identified 12 sRNA candidates that may possibly have cis-acting regulatory roles that are associated to a nutrient depleted growth environment. Our approach can be used for identifying novel sRNA genes and their possible role as cis-mediated regulatory systems.