The czcD (NiCo) Riboswitch Responds to Iron(II)

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• Iron is vital for life, but its dysregulation impacts pathogen virulence.
• Existing bacterial metal-responsive riboswitches do not typically sense iron (FeII).
• The czcD riboswitch was previously thought to respond only to nickel (NiII) and cobalt (CoII).

Purpose of the Study:

• To investigate the czcD riboswitch's response to FeII.
• To develop a novel, genetically encoded fluorescent sensor for FeII.
• To understand the in vivo function of czcD riboswitches in iron homeostasis.

Main Methods:

• Constructed a fluorescent sensor by fusing the czcD riboswitch with the Spinach2 RNA aptamer.
• Performed anaerobic in vitro assays to test metal ion responses.
• Assayed sensor function in Escherichia coli cells and tested downstream metal exporters.

Main Results:

• The engineered sensor responds anaerobically to FeII, MnII, CoII, NiII, and ZnII.
• Apparent Kd values for FeII (0.4 μM) and MnII (11 μM) fall within physiological ranges.
• The sensor successfully detected iron in E. coli, and downstream exporters mitigated iron toxicity.

Conclusions:

• Iron (FeII) is a plausible physiological ligand for czcD riboswitches.
• This work presents the first reversible, genetically encoded fluorescent sensor for FeII.
• The findings suggest czcD riboswitches play a role in bacterial iron homeostasis and metal stress responses.