Our results indicated that in strain RN6911 heterologous ��-toxin expression was sufficient to mediate phagosomal escape in HeLa. However, RN6911 shows drastically reduced ��-toxin production (Novick et?al., 1993). We thus currently assume either that the residual amount of ��-toxin produced by RN6911 is capable of inducing phagosomal escape in presence of ��-toxin or that alternative virulence factors can functionally replace ��-toxin. In L. monocytogenes infections, for instance, a broad range phospholipase C is involved in escape from the endomembrane system (Goldfine et?al., 1995; Marquis et?al., 1995; Sibelius et?al., 1996). With plc S. aureus possesses a homologue enzyme. Also, the Listeria metalloprotease mlp is able to disrupt infected HeLa phagosomes (Marquis et?al., AP24534 molecular weight
1995). Aureolysin is the homologous protease in S. aureus. We therefore hypothesize that enzymes other than ��-toxin can be co-opted for phagosomal escape. The peptide ��-toxin is a PSM. PSMs have been originally identified in Staphylococcus epidermidis (Mehlin et?al., 1999) and were recently discovered in S. aureus (Wang et?al., 2007). PSMs are also regulated by agr and their importance as virulence determinants was described recently (Vuong STI571 ic50
et?al., 2004; Wang et?al., 2007; Kretschmer et?al., 2010). PSMs of type �� are homologues of SLUSH and about 40?AA in length, whereas PSM�� are shorter (<?20?AA) and share structural similarities with ��-toxin (Wang et?al., 2007). Haemolysis of PSM�� was shown to be higher than that of PSM�� (Wang et?al., 2007). Figure?7C corroborates these findings and illustrates production of functional PSM by our transgenic laboratory strains. However, in our analyses PSM�� did not lead to phagosomal escape, whereas the recombinant strain expressing the PSM�� gene cluster efficiently escaped from the phagosome of HeLa (Fig.?7B). Our studies thus suggest that ��-type PSMs might only play a subordinate role in phagosomal escape. In contrast, PSM�� can functionally substitute ��-toxin in phagosomal escape (Fig.?7). The differential behaviour between PSM�� and PSM�� was unexpected, as ��-toxin is structurally more similar to PSM�� than to PSM�� (Wang et?al., 2007). In summary we conclude that S. aureus can employ at least two peptide toxins to avoid degradation within the phagosome, ��-toxin and PSM��. However, the detailed roles of the toxins identified <a href="http://en.wikipedia.org/wiki/MERTK
">UNC2881 in this work and possibly additional factors contributing to evasion of intracellular host cell defences as well as clinical significance of the toxins need further elucidation. Among the recombinant strains used in our study ��-toxin expressing S. aureus led to phagosomal escape; however, host cell numbers were comparable with the vector control (Fig.?6). Phagosomal escape thus might not be directly linked to host cell death. Thus, non-cytotoxic strains are a valuable tool in gain-of-function approaches to observe specific host�Cpathogen interactions.