Poster abstracts
Poster number 12 submitted by Janie Frandsen
New tRNA interaction sites in the US T-box riboswitch
Jane K Frandsen (Ohio State Biochemistry Program, Center for RNA Biology), Anna V. Sherwood (Molecular Cellular and Developmental Biology Graduate Program, Center for RNA Biology), Frank J. Grundy (Department of Microbiology, Center for RNA Biology), Tina M. Henkin (Department of Microbiology, Ohio State Biochemistry Program, Molecular Cellular and Developmental Biology Graduate Program, Center for RNA Biology)
Abstract:
T-box riboswitches regulate the expression of essential amino acid-related genes in Firmicutes and Actinobacteria by monitoring the aminoacylation status of a cognate tRNA (1). In T-box RNAs with a canonical Stem I, the Specifier Loop interacts with the anticodon of the cognate tRNA and the terminal loop and AG bulge form a loop-loop structure that interacts with the tRNA elbow. The Ultrashort (US) Stem I class of ileS T-box riboswitches, which regulate isoleucyl-tRNA synthetase genes, have an alternate Specifier Loop structure and lack the elements necessary for interaction with the tRNA elbow, but contain the highly conserved Stem II and Stem IIA/B pseudoknot, whose role in tRNA recognition remained unknown (2). We have now demonstrated that both Stem II and Stem IIA/B of the US ileS RNA contribute to tRNAIle affinity (3). Additionally, using selective 2’-hydroxyl acylation analyzed by primer extension (SHAPE), crosslinking, and mutational studies, we identified two new interaction sites, one between the S-turn element in Stem II and the tRNA T arm and the other between the pseudoknot and the tRNA D loop. This is the first example of tRNA recognition by an S-turn motif or a pseudoknot element, revealing new types of RNA-RNA interactions. These data provide the first biochemical evidence for the functional role of Stem II and the pseudoknot, which are present in the majority of T-box RNAs but absent in the glyQS RNAs that have been used for most of the current biochemical and structural analyses. We hypothesize that these interactions are important for the recognition and binding of the cognate tRNA and discrimination against non-cognate tRNA in those T-box RNAs in which they are present, and propose that structural variability in T-box riboswitches indicates alternate solutions to the tRNA recognition problem.
References:
1. Henkin TM. Biochim Biophys Acta 1839:959-63 (2014).
2. Sherwood AV, Grundy FJ, Henkin TM. Proc Natl Acad Sci USA 112:1113-18 (2015).
3. Sherwood AV, Frandsen JK, Grundy FJ, Henkin TM. Proc Natl Acad Sci USA 115:3894-99 (2018).
Keywords: Riboswitch, tRNA, S-turn, Pseudoknot, SHAPE