Salmonella is able to invade the host cells and survive inside the host cells. In contrary, EHEC exerts its virulence by adhering to the surface of the host cells via actin-rich pedestals. It is not clear whether the different biochemical E3 ligase activities contribute to the different life-style of the two intestinal pathogens. We speculate that NleL may exploit the free ubiquitin pools in the host cell to exert its function without the canonical substrates. Further studies are needed to examine this possibility. Our data demonstrate that loss of the NleL E3 ligase activity leads to increased pedestal formation. Preliminary work showed that the loss of the NleL E3 ligase activity did not alter the expression of Tir or EspFu, suggesting that NleL may exert its function after their translocation. EPEC infection has been shown to lead to pedestal formation much more efficiently than that of EHEC in vitro. A significant difference between the two pathogenic types lies in the nature of the involvement of Tir during pedestal formation. While TirEPEC is tyrosine phosphorylated upon entry into the host cell and interacts with host adaptor protein Nck, which stimulates Arp2/3- mediated actin polymerization, TirEHEC lacks the homologous tyr residue, Y474, and depends on another effector EspFu/TccP to induce actin polymerization. Such studies highlight the differences in the mechanism of pedestal formation between the two organisms. Our data indicate that NleL probably functions at steps common to EPEC and EHECmediated pedestal formation. It is reasonable to speculate that NleL ubiquitinates an unknown factor of bacterial or host origin involved in pedestal formation. One such scenario would be that host cell proteins involved in pedestal formation are ubiquitinated by NleL which, in turn, corresponds to a decrease in the level of host proteins or alters their localization, resulting in a decrease in pedestal formation. Alternatively, NleL may ubiquitinate Tir and promote its endocytosis, thus decreasing the surface availability of Tir. Our preliminary studies have shown that Tir is not a substrate for the E3 ligase activity of NleL in an in vitro ubiquitination assay. However, we cannot rule out that a third factor is required, but absent, in our in vitro reaction. A recent structural study showed that another non-LEE-encoded effector, NleG, is a RING finger ubiquitin E3 ligase. It is tempting to speculate that NleL, NleG and Tir may work together to modulate the actin pedestal formation.