Recognition of the bacterial second messenger cyclic diguanylate by its cognate riboswitch

Abstract

C-di-GMP is a bacterial second messenger implicated in processes such as biofilm formation and switches between motile and sedentary lifestyles. The structure of the c-di-GMP–binding GEMM riboswitch is now presented with ligand and the large conformational changes between ligand-bound and unbound forms analyzed by small-angle X-ray scattering. The cyclic diguanylate (bis-(3′-5′)-cyclic dimeric guanosine monophosphate, c-di-GMP) riboswitch is the first known example of a gene-regulatory RNA that binds a second messenger. c-di-GMP is widely used by bacteria to regulate processes ranging from biofilm formation to the expression of virulence genes. The cocrystal structure of the c-di-GMP responsive GEMM riboswitch upstream of the tfoX gene of Vibrio cholerae reveals the second messenger binding the RNA at a three-helix junction. The two-fold symmetric second messenger is recognized asymmetrically by the monomeric riboswitch using canonical and noncanonical base-pairing as well as intercalation. These interactions explain how the RNA discriminates against cyclic diadenylate (c-di-AMP), a putative bacterial second messenger. Small-angle X-ray scattering and biochemical analyses indicate that the RNA undergoes compaction and large-scale structural rearrangement in response to ligand binding, consistent with organization of the core three-helix junction of the riboswitch concomitant with binding of c-di-GMP.