Crystal structure of a c‐di‐AMP riboswitch reveals an internally pseudo‐dimeric RNA

Abstract

Cyclic diadenosine monophosphate (c-di-AMP) is a second messenger that is essential for growth and homeostasis in bacteria. A recently discovered c-di-AMP-responsive riboswitch controls the expression of genes in a variety of bacteria, including important pathogens. To elucidate the molecular basis for specific binding of c-di-AMP by a gene-regulatory mRNA domain, we have determined the co-crystal structure of this riboswitch. Unexpectedly, the structure reveals an internally pseudo-symmetric RNA in which two similar three-helix-junction elements associate head-to-tail, creating a trough that cradles two c-di-AMP molecules making quasiequivalent contacts with the riboswitch. The riboswitch selectively binds c-di-AMP and discriminates exquisitely against other cyclic dinucleotides, such as c-di-GMP and cyclic-AMP-GMP, via interactions with both the backbone and bases of its cognate second messenger. Small-angle X-ray scattering experiments indicate that global folding of the riboswitch is induced by the two bound cyclic dinucleotides, which bridge the two symmetric three-helix domains. This structural reorganization likely couples c-di-AMP binding to gene expression.

RNA-Puzzles Round III: 3D RNA structure prediction of five riboswitches and one ribozyme

“…The bubble between P2 and P3 helix was mostly unresolved in the X-ray map, implying that this region could be less stable in structure. Nevertheless, independent crystallographic solutions of homologous c-di-AMP structures (Gao and Serganov 2014;Jones and Ferré-D'Amaré 2014) (also released after RNA-Puzzle 12 modeling) showed strong agreement with all resolved parts of the crystal structure considered herein, suggesting that the overall fold is well defined and a valid target for prediction. Many of the predicted structures do not fully consider the binding of the two c-di-AMP ligands, but the global topologies of the top ranking models are still visually similar to the X-ray structure (Ding group model in Fig.…”

Riboswitch diversity and distribution

“…Therefore, cells that carry this riboswitch class should be able to regulate gene expression more "digitally" (all on or all off) in response to very small changes in ligand concentration. Metabolite-binding riboswitches that exhibit similar cooperative ligand-binding characteristics have been reported for the ligands glycine Butler et al 2011), THF (Trausch et al 2011), c-di-AMP (Gao and Serganov 2014;Jones and Ferré-D'Amaré 2014;Ren and Patel 2014), and guanidine . For cells that carry Mg 2+ -I riboswitches, a more digital genetic switch might be worth the extra cost of conserving and producing a more complex RNA architecture.…”

Identification, Characterization, and Structure Analysis of the Cyclic di-AMP-binding PII-like Signal Transduction Protein DarA

“…This observation was recently confirmed for the corresponding protein CabP from Streptococcus pneumoniae (23). Further studies aimed at the identification of the controlling metabolite of the B. subtilis ydaO riboswitch identified the binding of c-di-AMP to this RNA structure (24,25). It is interesting to note that the ydaO riboswitch controls two transcription units in B. subtilis: the ydaO gene and the ktrAB operon that encodes the c-di-AMP-responsive potassium transporter (26).…”