Poster abstracts
Poster number 77 submitted by Jiansong Xu
The czcD (NiCo) riboswitch responds to iron(II)
Jiansong Xu (Department of Chemistry, Penn State University), Joseph Cotruvo (Department of Chemistry, Penn State University)
Abstract:
Iron is essential for nearly every organism, and mismanagement of its intracellular concentrations (either deficiency or excess) contributes to diminished virulence in human pathogens, necessitating intricate metalloregulatory mechanisms. To date, although several metal-responsive riboswitches have been identified in bacteria, none has been shown to respond to FeII. The czcD riboswitch, present in numerous human gut microbiota and pathogens, was recently shown to respond to NiII and CoII but claimed not to respond to FeII, on the basis of aerobic, in vitro assays. Its function in vivo is not well understood. We constructed a fluorescent sensor using this riboswitch fused to the RNA aptamer, Spinach2. When assayed anaerobically, the resulting sensor responds in vitro to FeII, as well as to MnII, CoII, NiII, and ZnII, but only in the cases of FeII and MnII do the apparent Kds (0.4 µM and 11 µM, respectively) fall within the range of labile metal concentrations maintained by known metalloregulators. We also show that the sensor – which is, to the best of our knowledge, the first reversible genetically encoded fluorescent sensor for FeII – responds to iron in E. coli cells. Finally, we demonstrate that the putative metal exporters directly downstream of two czcD riboswitches efficiently rescue iron toxicity in a heterologous expression system. Together, our results suggest that iron should be considered as a plausible physiological ligand for czcD riboswitches, although response to general metal stress cannot be ruled out at present.
Keywords: riboswitch, iron, NiCo