Bacteria utilize extracellular factors to interact with neighboring cells. Bacteria may secrete toxins that inhibit the growth of other bacteria in order to compete for nutrient and space availability. These signaling molecules can also modulate gene expression and surrounding cellular behavior. Signaling molecules act in long range signaling via secreted peptides and compounds, as in quorum sensing, or in the short range using extracellular proteins requiring contact to deliver their signal.

Contact dependent growth inhibition (CDI) was originally discovered in an Escherichia coli strain that dominated the rat intestine. This strain, called EC93, was able to inhibit the growth of other laboratory strains of E. coli. CdiB and CdiA are essential for growth inhibition and an immunity protein confers protection to CDI. The CdiA C-terminus (CdiA-CT) encodes a toxin responsible for CDI. The outer membrane protein, BamA, and inner membrane drug pump, AcrB, are needed for target cell inhibition. CDI systems have been discovered in many species of bacteria, including uropathogenic E. coli 536 (UPEC 536).

This dissertation covers several aspects of UPEC 536 CDI. The visualization of CdiA-CT peptide transfer into the cytoplasm of a target cell is shown for the first time. Although both the N-terminus (NT) and CT of CdiA are delivered to the surface of target cells, only CdiA-CT becomes intracellular. It is also demonstrated that the NT and the CT of CdiA are exposed on the surface of inhibitor cells. Mutations in both inhibitor cells and targets that affect inhibitory activity are then described. Attempts to make non-binding inhibitor and target cells are discussed. Unknown point mutants that are partially resistant to EC93 and UPEC 536 CDI are introduced. Point mutations in AcrB that may affect CdiA binding and/or transit of CdiA-CT are also shown. Furthermore, ClpXP and ClpAP protease expression in target cells is shown to be necessary for inhibition by UPEC 536. Finally, UPEC 536 inhibitors containing a CdiA-CT deletion and an inactive CdiA-CT mutant were found to be defective in early biofilm development.