c-di-GMP modulates ribosome assembly by inhibiting rRNA methylation

Cyclic diguanosine monophosphate (c-di-GMP) is a ubiquitous bacterial secondary messenger, with diverse functions, many of which are yet to be uncovered. Stemming from an Escherichia coli proteome microarray, we found that c-di-GMP bound to 23S rRNA methyltransferases (RlmI and RlmE). rRNA methylation assays showed that c-di-GMP inhibits RlmI activity, thereby modulating ribosome assembly. Based on molecular dynamic simulation and mutagenesis studies, we found that c-di-GMP binds to RlmI at residues R64, R103, G114, and K201. Structural simulation revealed that c-di-GMP quenches RlmI activity by inducing the closure of the catalytic pocket. Furthermore, we revealed that c-di-GMP promotes antibiotic tolerance by regulating RlmI activity, which played a role in antibiotic-resistant strains. Finally, the binding and methylation assays showed that the effect of c-di-GMP on RlmI is conserved, at least in various pathogenic bacteria. This study discovered an unexpected functional role of c-di-GMP in regulating ribosome assembly by inhibiting rRNA methylases. This study identified an unexpected but crucial member among the c-di-GMP effectors. HighlightsO_LIc-di-GMP regulates ribosome assembly in Escherichia coli. C_LIO_LIc-di-GMP inhibits rRNA methylation activity of RlmI by inducing catalytic pocket closure. C_LIO_LIc-di-GMP promotes antibiotic resistance by regulating ribosome assembly. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/597503v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@52baforg.highwire.dtl.DTLVardef@176f86dorg.highwire.dtl.DTLVardef@1afed11org.highwire.dtl.DTLVardef@f53687_HPS_FORMAT_FIGEXP M_FIG C_FIG