Stapels D A C, Kuipers A, von Köckritz-Blickwede M, Ruyken M, Tromp A T, Horsburgh M J, de Haas C J C, van Strijp J A G, van Kessel K P M, Rooijakkers S H M
Medical Microbiology, University Medical Center Utrecht, Utrecht, the Netherlands.
Department of Physiological Chemistry, University of Veterinary Medicine Hannover, Hannover, Germany.
Cell Microbiol. 2016 Apr;18(4):536-45. doi: 10.1111/cmi.12528. Epub 2015 Oct 23.
Neutrophils store large quantities of neutrophil serine proteases (NSPs) that contribute, via multiple mechanisms, to antibacterial immune defences. Even though neutrophils are indispensable in fighting Staphylococcus aureus infections, the importance of NSPs in anti-staphylococcal defence is yet unknown. However, the fact that S. aureus produces three highly specific inhibitors for NSPs [the extracellular adherence proteins (EAPs) Eap, EapH1 and EapH2], suggests that these proteases are important for host defences against this bacterium. In this study we demonstrate that NSPs can inactivate secreted virulence factors of S. aureus and that EAP proteins function to prevent this degradation. Specifically, we find that a large group of S. aureus immune-evasion proteins is vulnerable to proteolytic inactivation by NSPs. In most cases, NSP cleavage leads to functional inactivation of virulence proteins. Interestingly, proteins with similar immune-escape functions appeared to have differential cleavage sensitivity towards NSPs. Using targeted mutagenesis and complementation analyses in S. aureus, we demonstrate that all EAP proteins can protect other virulence factors from NSP degradation in complex bacterial supernatants. These findings show that NSPs inactivate S. aureus virulence factors. Moreover, the protection by EAP proteins can explain why this antibacterial function of NSPs was masked in previous studies. Furthermore, our results indicate that therapeutic inactivation of EAP proteins can help to restore the natural host immune defences against S. aureus.
中性粒细胞储存大量中性粒细胞丝氨酸蛋白酶（NSPs），这些蛋白酶通过多种机制有助于抗菌免疫防御。尽管中性粒细胞在对抗金黄色葡萄球菌感染中不可或缺，但NSPs在抗葡萄球菌防御中的重要性尚不清楚。然而，金黄色葡萄球菌能产生三种针对NSPs的高度特异性抑制剂[细胞外黏附蛋白（EAPs）Eap、EapH1和EapH2]，这一事实表明这些蛋白酶对宿主抵御这种细菌很重要。在本研究中，我们证明NSPs可使金黄色葡萄球菌分泌的毒力因子失活，而EAP蛋白的作用是防止这种降解。具体而言，我们发现一大类金黄色葡萄球菌免疫逃避蛋白易受NSPs的蛋白水解失活作用影响。在大多数情况下，NSPs的切割会导致毒力蛋白功能失活。有趣的是，具有相似免疫逃避功能的蛋白对NSPs的切割敏感性似乎存在差异。通过在金黄色葡萄球菌中进行靶向诱变和互补分析，我们证明所有EAP蛋白都能保护其他毒力因子免受复杂细菌上清液中NSPs的降解。这些发现表明NSPs可使金黄色葡萄球菌毒力因子失活。此外，EAP蛋白的保护作用可以解释为什么NSPs的这种抗菌功能在以前的研究中被掩盖了。此外，我们的结果表明，EAP蛋白的治疗性失活有助于恢复宿主对金黄色葡萄球菌的天然免疫防御。
