EHEC EspF is expressed and secreted in a CesF-dependent process. (A) Proteins from whole bacterial pellets (P) or precipitated supernatants (S) from various strains were separated by polyacrylamide gel electrophoresis and immunoblotted by using antibody raised against EPEC EspF. The lanes are as follows: whole bacterial proteins from EPEC grown in DMEM (lane 1); EHEC whole bacterial proteins (lane 2) and secreted proteins (lane 3) grown in LB; and EHEC whole bacterial protein (lane 4), EHEC secreted proteins (lane 5), and EHEC ΔEspF whole bacterial proteins (lane 6) grown in DMEM. (B) The role of CesF in the expression and secretion of EspF was determined by immunoblotting proteins from bacterial pellets (P) and secreted (S) proteins from wild-type (lanes 1, 3, 5, and 7) and ΔcesF derivatives (lanes 2, 4, 6, and 8) of both EPEC and EHEC. Deletion of cesF decreased the amount of EspF expressed and secreted in both EPEC and EHEC.