Poster abstracts

Poster number 57 submitted by Alex Runyon

Importance of conserved elements in the Bacillus subtilis thrS canonical T-box riboswitch

Alexander Runyon (Department of Microbiology and Center for RNA biology, The Ohio State University), Tina M. Henkin (Department of Microbiology, Ohio State Biochemistry Program, Molecular Cellular and Developmental Biology, Center for RNA Biology), Frank J. Grundy (Department of Microbiology, Center for RNA Biology)

Abstract:
T-box riboswitches are cis-acting RNA structures that modulate expression of downstream amino acid-related genes in Gram-positive bacteria. They are induced in response to a specific uncharged tRNA that corresponds to the amino acid specificity of the downstream gene, which is regulated through transcription antitermination or sequestration of the Shine-Dalgarno sequence. T-box riboswitches with all of the conserved elements found in the majority of T-box genes are known as canonical T-box riboswitches. Current biochemical understanding of T-box riboswitch function comes from models that are missing key conserved elements. The Bacillus subtilis thrS riboswitch is located upstream of the threonyl-tRNA synthetase coding sequence and contains all of the key conserved elements, making it a canonical riboswitch. tRNAThr-dependent antitermination in vitro has been demonstrated, which allows comparisons of biochemical data obtained from non-canonical riboswitches to that derived from the more common canonical riboswitch organization. Mutations have been generated in conserved elements in thrS, which are known to disrupt these elements in other T-box RNAs. Initial results from in vitro transcription and tRNA binding assays suggest that the conserved elements in thrS are critical for tRNA-dependent antitermination activity. In addition, some elements of thrS have been modified to resemble the corresponding elements of the non-canonical ileS US riboswitch found in Actinobacteria. Our results have shown that modification of elements in thrS results in lower binding affinity for tRNAThr. This system will allow us to gain a better understanding of the function of canonical T-box RNAs.

Keywords: Riboswitch, T-box, thrS