Full functional activity of SSL7 requires binding of both complement C5 and IgA

Abstract

Staphylococcus aureus is an opportunistic bacterial pathogen responsible for a range of diseases, from local skin infections through to life-threatening illnesses such as toxic shock syndrome. S. aureus produces an assortment of molecules designed to evade or subvert the host immune system. One example is the 23 kDa staphylococcal superantigen-like protein 7 (SSL7) that simultaneously binds immunoglobulin A (IgA) and complement C5 to inhibit complement-mediated hemolytic and bactericidal activity. The avirulent bacterium Lactococcus lactis was engineered to express SSL7 so that its role in bacterial survival could be assessed without interference from other virulence factors. Expression of SSL7 by L. lactis led to significantly enhanced bacterial survival in whole human blood and prevented the membrane attack complex (C5b-9) forming on the cell wall. To further understand the mechanism of action of SSL7, the activity of wild-type SSL7 protein was compared with a panel of mutant proteins lacking the capacity to bind IgA, C5, or both IgA and C5. SSL7 potently inhibited in vitro chemotaxis of inflammatory myeloid cells in response to a pathogenic stimulus and when injected into mice, SSL7 blocked the migration of neutrophils into the peritoneum in response to an inoculum of heat-killed S. aureus. Mutagenesis of the C5-binding site on SSL7 abolished all inhibitory activity, while mutation of the IgA-binding site had only partial effects, indicating that while IgA binding enhances activity it is not essential. SSL7 is an important staphylococcal virulence factor with potent anti-inflammatory properties, which are mediated by targeting complement C5 and IgA.

Streptococcal 5′-Nucleotidase A (S5nA), a Novel Streptococcus pyogenes Virulence Factor That Facilitates Immune Evasion

“…A significantly increased survival rate (63%, p Ͻ 0.001) was also observed with equimolar amounts of the control protein SSL7. SSL7 is an S. aureus immune evasion factor previously shown to increase the survival of L. lactis in human blood (34). The addition of 0.01 and 0.1 M rS5nA also increased bacterial survival in a concentration-dependent manner (8.9 and 18.1%, respectively), but this was not statistically significant (p Ͼ 0.1).…”

“…L. lactis is a non-pathogenic, food grade bacterium that is rapidly killed when incubated with whole blood. Increased survival of the bacteria in blood has been used to analyze potential virulence factors (30,34). The addition of rS5nA to L. lactis in human whole blood significantly increased bacterial survival in a dose-dependent manner (Fig.…”

Bacterial superantigens and superantigen-like toxins

“…Additionally, the binding of SSL7 to already activated C5b results in an inhibition of membrane attack complex (MAC) formation. Consequently, inhibition of C5a-mediated chemotaxis of inflammatory myeloid cells and C5a-dependent microbial killing relies on the presence of IgA, whereas MACmediated killing is solely dependent on SSL7 [374,382]. MAC, though, is not to be effective against Gram-positive bacteria due to their thickened outer wall.…”

“…Recent evidence, however, shows that MAC does form on Gram-positive bacteria and SSL7 can inhibit this [374,388]. In vivo studies have shown that SSL7 can inhibit the migration of neutrophils into the peritoneum in response to S. aureus and that this activity is significantly more effective in the presence of IgA binding [374]. SSL7 has the ability to block the binding of IgA to its Fc receptor so it could potentially prevent FcαRI effector functions that include phagocytosis of microbes [389].…”

“…MAC, though, is not to be effective against Gram-positive bacteria due to their thickened outer wall. Recent evidence, however, shows that MAC does form on Gram-positive bacteria and SSL7 can inhibit this [374,388]. In vivo studies have shown that SSL7 can inhibit the migration of neutrophils into the peritoneum in response to S. aureus and that this activity is significantly more effective in the presence of IgA binding [374].…”