Serine protease RAYM_01812 (SspA) inhibits complement-mediated killing and monocyte chemotaxis and contributes to virulence of Riemerella anatipestifer in ducks.

IF 5.5 1区 农林科学 Q1 IMMUNOLOGY Virulence Pub Date : 2024-12-01 Epub Date: 2024-11-04 DOI:10.1080/21505594.2024.2421219
Rongkun Yang, Sen Li, Jie Guo, Yanhua Wang, Zeyuan Dong, Qing Wang, Hongying Bai, Congran Ning, Xiaotong Zhu, Jiao Bai, Sishun Hu, Yuncai Xiao, Zili Li, Zutao Zhou
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Abstract

Riemerella anatipestifer (RA) is a significant poultry pathogen causing acute septicemia and inflammation. The function of protease RAYM_01812, responsible for gelatin degradation, is unexplored in RA pathogenesis. To elucidate its role, we generated a deletion mutant ΔRAYM_01812 (ΔRAYM) and complementary CΔRAYM_01812 (CΔRAYM) strain and revealed the protease's role in extracellular gelatinase activity. By expressing full-length 76 kDa RAYM_01812 protein without signal peptide as well as seven partial structural domains fragments, we evidence that the N-terminal propeptide acts as an enzymatic activity inhibitor and it gets cleaved at A112. Also, we show that the β-fold sheet domain is necessary for enhancing the enzymatic protease activity. Sequential auto-proteolysis forms a stable 40 kDa enzyme. Then, testing the strains in duck sera indicated that the absence or presence of RAYM_01812 results in reduced or enhanced bacterial survival, respectively. Furthermore, we found that the protease is able to cleave IgY antibodies as well as the complement factors C3a and C5a, that the protease reduces C3a- or C5a-mediated monocyte chemotaxis, and results in enhanced membrane attack complex (MAC) formation on the surface of ΔRAYM compared to CΔRAYM. This suggests that RAYM_01812 plays a crucial role in protecting against the serum complement-mediated bactericidal effect through inhibiting MAC formation and monocyte chemotaxis. Animal infection assays showed a 1090-fold reduced virulence of ΔRAYM compared to RA-YM, evidenced by decreased tissue loading and weaker histopathological changes. In conclusion, RAYM_01812 acts as a vital virulence factor, enabling host innate immune defence escape through complement killing evasion and monocyte chemotaxis inhibition.

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丝氨酸蛋白酶 RAYM_01812 (SspA) 可抑制补体介导的杀伤作用和单核细胞趋化作用,并有助于提高鸭毒酵母菌的毒力。
锐毛霉菌(RA)是一种重要的家禽病原体,可引起急性败血症和炎症。负责明胶降解的蛋白酶 RAYM_01812 在 RA 致病机理中的功能尚未得到研究。为阐明其作用,我们生成了缺失突变体ΔRAYM_01812(ΔRAYM)和互补的CΔRAYM_01812(CΔRAYM)菌株,并揭示了该蛋白酶在细胞外明胶酶活性中的作用。通过表达不含信号肽的全长 76 kDa RAYM_01812 蛋白以及 7 个部分结构域片段,我们证明 N-末端的前肽是酶活性的抑制剂,它在 A112 处被裂解。此外,我们还发现,β-折叠片状结构域是增强酶蛋白酶活性的必要条件。连续的自动蛋白水解形成了稳定的 40 kDa 酶。然后,在鸭血清中检测菌株,结果表明 RAYM_01812 的缺失或存在分别导致细菌存活率降低或提高。此外,我们还发现该蛋白酶能裂解 IgY 抗体以及补体因子 C3a 和 C5a,该蛋白酶能降低 C3a 或 C5a 介导的单核细胞趋化作用,并且与 CΔRAYM 相比,能增强膜攻击复合物(MAC)在 ΔRAYM 表面的形成。这表明,RAYM_01812 通过抑制 MAC 的形成和单核细胞趋化,在抵御血清补体介导的杀菌作用方面发挥了重要作用。动物感染实验表明,与 RA-YM 相比,ΔRAYM 的致病力降低了 1090 倍,表现为组织负荷减少和组织病理学变化减弱。总之,RAYM_01812 是一种重要的毒力因子,可通过逃避补体杀伤和抑制单核细胞趋化来逃避宿主的先天免疫防御。
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来源期刊
Virulence
Virulence IMMUNOLOGY-MICROBIOLOGY
CiteScore
9.20
自引率
1.90%
发文量
123
审稿时长
6-12 weeks
期刊介绍: Virulence is a fully open access peer-reviewed journal. All articles will (if accepted) be available for anyone to read anywhere, at any time immediately on publication. Virulence is the first international peer-reviewed journal of its kind to focus exclusively on microbial pathogenicity, the infection process and host-pathogen interactions. To address the new infectious challenges, emerging infectious agents and antimicrobial resistance, there is a clear need for interdisciplinary research.
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