Mechanism of Bacterial Expulsion from Infected Bladder Cells.

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Urinary tract infections (UTIs) account for the second most common bacterial infections in man. Most of these infections are caused by E.coli which have a distinct mechanism for entry into the highly impregnable superficial epithelium of the bladder. While investigating molecular aspects of how uropathogenic E.coli (UPEC) enter bladder epithelial cells (BECs), we observed that infected BECs have a powerful and largely overlooked capacity to exocytose most of the infecting UPEC without loss of cellular viability. These observations point to a powerful capacity of BECs to sense intracellular bacteria and initiate bacterial expulsion activities. Using biochemical and molecular approaches, we have identified a distinct mechanism in BECs that are capable of recognizing intracellular UPEC involving the imunosurveillance molecule, Toll like Receptor (TLR)4. We have also identified several key mediators of bacterial exocytosis which include components of the exocyst complex a Rab GTPase, Rab11 and the SNARE complex. Additionally, we have implicated cellular components in lipid raft compartments not traditionally associated in exocytic processes in the extrusion of bacteria through the plasma membrane. Studies to examine how these various signaling components and pathways integrate and achieve bacterial expulsion could provide valuable clues on how to therapeutically maximize bacterial expulsion mechanisms in the bladder. In this proposal, we plan to confirm and extend these observation in the following specific Aims: (i) Elucidate the mechanism by which TLR4 senses intravesicular E. coli in BECs. (ii) Investigate how exocyst complex and vesicular trafficking elements promote bacterial expulsion. (iii) Determine the contribution of SNARE complex to the membrane fusion and apical bacteria discharge. (iv) Identify components in cellular lipid raft fractions involved in bacterial expulsion.