Poster abstracts
Poster number 1 submitted by Divyaa Bhagdikar
Identification and characterization of S-box riboswitches that regulate at the level of translation initiation
Divyaa Bhagdikar (Department of Microbiology and Center for RNA biology, The Ohio State University, Columbus, Ohio ), Frank J. Grundy (Department of Microbiology and Center for RNA biology, The Ohio State University, Columbus, Ohio ), Tina M. Henkin (Department of Microbiology and Center for RNA biology, The Ohio State University, Columbus, Ohio )
Abstract:
The S-box riboswitches regulate the expression of genes involved in methionine and cysteine metabolism. Most S-Box riboswitches operate at the level of transcription attenuation, such that a terminator helix is stabilized when S-adenosylmethionine (SAM) binds the aptamer. In silico analyses have suggested that a rarer class of S-box riboswitches may regulate at the level of translation initiation. We identified a leader RNA, metI from Desulfurispirilum indicum, that has the potential for SAM-dependent sequestration of the Shine-Dalgarno (SD) region which suggests translational regulation. The aptamer domain of the RNA bound SAM with affinity comparable to that of the well-characterized Bacillus subtilis yitJ S-box riboswitch, and with similar selectivity against S-adenosylhomocysteine (SAH). A mutation at a position previously shown to disrupt SAM binding in other S-box RNAs resulted in loss of SAM binding in metI. These results demonstrate that the aptamer domain has SAM binding properties similar to those of previously characterized S-box riboswitches. The SD region of the RNA exhibited SAM-dependent structural rearrangements consistent with the hypothesis that regulation occurs at the level of translation initiation. Additionally, binding of 30S subunits to metI RNA was reduced in the presence of SAM, demonstrating that the riboswitch regulates at the level of translation initiation. Preliminary fluorescence studies with the full-length metI RNA suggest that the riboswitch can perform reversible regulation in response to changing levels of SAM. The metI riboswitches from Desulfurispirilum indicum and Bacillus subtilis will be used as a model to compare the characteristics of transcriptional and translational riboswitches in the same class. We hypothesize that the translational riboswitches can perform multiple reversible regulatory decisions whereas transcriptional riboswitches cannot reverse their regulatory decisions. Such a comparative study will provide insight into the functional role of transcriptional vs. translational regulation of the same gene by the same class of riboswitch in different organisms.
Keywords: S-box, Translation, Riboswitch