Structural and functional insights into the Rcs phosphorelay.

Area of Science:

• Microbiology
• Structural Biology
• Molecular Biology

Background:

• The Rcs phosphorelay controls gene expression during cell envelope stress, impacting virulence in bacteria like *Klebsiella pneumoniae*.
• Key components include RcsC, RcsD, and RcsB, negatively regulated by inner membrane protein IgaA.
• Outer membrane lipoprotein RcsF activates the system via IgaA, but the mechanism is not fully understood.

Purpose of the Study:

• To elucidate the structural mechanisms underlying Rcs phosphorelay activation.
• To determine the structures of IgaA, the IgaA/RcsF complex, RcsC, and RcsD.

Main Methods:

• Cryo-electron microscopy (Cryo-EM) was used to determine the structures of IgaA, the IgaA/RcsF complex, RcsC, and RcsD.
• AlphaFold3 structure predictions were employed for IgaA/RcsD and RcsF/IgaA/RcsD complexes.
• Genetic studies were integrated with structural data.

Main Results:

• The structures of IgaA, IgaA/RcsF complex, RcsC, and RcsD were determined.
• RcsC and RcsD were found to form stable homodimers with ladder-like structures.
• A model was proposed where RcsF binding to IgaA alters the IgaA/RcsD interaction, activating the Rcs phosphorelay.

Conclusions:

• Structural and genetic data provide a high-resolution view of the Rcs stress response system.
• The findings reveal how RcsF binding to IgaA relieves negative regulation and activates the Rcs phosphorelay.
• The study suggests potential targets for developing small molecule inhibitors against this bacterial system.