Clip-SP1 cleavage activates downstream prophenoloxidase activating protease (PAP) in Plutella xylostella

• Published in: Developmental and comparative immunology 2023-09, Vol.146, p.104737-104737, Article 104737
• Main Authors: Dong, Yi, Hou, Qing, Ye, Min, Li, Zeyun, Li, Jingge, You, Minsheng, Yuchi, Zhiguang, Lin, Junhan, You, Shijun
• Format: Article
• Language: English
• Subjects: Animals; Bacillus thuringiensis; Clip-SP; Enzyme Precursors - metabolism; Insect Proteins - metabolism; Larva; Lepidoptera; Midgut; Monophenol Monooxygenase; Phenoloxidase; Serine Endopeptidases - genetics; Serine Endopeptidases - metabolism; Serine Proteases - genetics; Serine Proteases - metabolism
• ISSN: 0145-305X; 1879-0089
• DOI: 10.1016/j.dci.2023.104737

Melanization is a component of the humoral immune defense of insects and is induced by serine protease-mediated phenoloxidase (PO) catalysis. Prophenoloxidase (PPO) in the midgut of Plutella xylostella is activated by the CLIP domain serine protease (clip-SP) in response to Bacillus thuringiensis (Bt) infection, but the detailed signaling cascade following this activation is unknown. Here, we report that activation of clip-SP enhances PO activity in the P. xylostella midgut by cleaving three downstream PPO-activating proteases (PAPs). First, the expression level of clip-SP1 was increased in the midgut after Bt8010 infection of P. xylostella. Then, purified recombinant clip-SP1 was able to activate three PAPs - PAPa, PAPb and PAP3 - which in turn enhanced their PO activity in the hemolymph. Furthermore, clip-SP1 showed a dominant effect on PO activity compared to the individual PAPs. Our results indicate that Bt infection induces the expression of clip-SP1, which is upstream of a signaling cascade, to efficiently activate PO catalysis and mediate melanization in the midgut of P. xylostella. And it provides a basis for studying the complex PPO regulatory system in the midgut during Bt infection. •Clip-SP1 is highly expressed in midgut and significantly upregulated after Bt infection.•Clip-SP1 can cleave to activate the downstream PAPa, PAPb and PAP3.•Clip-SP1, PAPa, PAPb and PAP3 are all able to significantly increase PO activity in the hemolymph.