Wang Yang, Jiang Haobo
Department of Entomology and Plant Pathology, Oklahoma State University, Stillwater, OK 74078, USA.
Insect Biochem Mol Biol. 2008 Aug;38(8):763-9. doi: 10.1016/j.ibmb.2008.04.008. Epub 2008 May 20.
In Manduca sexta, pathogen recognition triggers a branched serine proteinase cascade which generates active phenoloxidase (PO) in the presence of a proPO-activating proteinase (PAP) and two noncatalytic serine proteinase homologs (SPHs). PO then catalyzes the production of reactive compounds for microbe killing, wound healing, and melanin formation. In this study, we discovered that a minute amount of PAP1 (a final component of the proteinase pathway) caused a remarkable increase in PO activity in plasma from naïve larvae, which was significantly higher than that from the same amounts of PAP1, proPO and SPHs incubated in vitro. The enhanced proPO activation concurred with the proteolytic activation of HP6, HP8, PAP1, SPH1, SPH2 and PO precursors. PAP1 cleaved proSPH2 to yield bands with mobility identical to SPH2 generated in vivo. PAP1 partially hydrolyzed proHP6 and proHP8 at a bond amino-terminal to the one cut in the PAP1-added plasma. PAP1 did not directly activate proPAP1. These results suggest that a self-reinforcing mechanism is built into the proPO activation system and other plasma proteins are required for cleaving proHP6 and proHP8 at the correct site to strengthen the defense response, perhaps in the early stage of the pathway activation.
在烟草天蛾中，病原体识别会触发一个分支的丝氨酸蛋白酶级联反应，该反应在存在前酚氧化酶激活蛋白酶（PAP）和两种非催化性丝氨酸蛋白酶同源物（SPH）的情况下产生活性酚氧化酶（PO）。然后，PO催化产生用于杀灭微生物、伤口愈合和黑色素形成的反应性化合物。在本研究中，我们发现极少量的PAP1（蛋白酶途径的最终成分）会使未接触过病原体的幼虫血浆中的PO活性显著增加，这明显高于在体外孵育相同量的PAP1、前酚氧化酶和SPH时的PO活性。前酚氧化酶激活的增强与HP6、HP8、PAP1、SPH1、SPH2和PO前体的蛋白水解激活同时发生。PAP1切割前SPH2，产生迁移率与体内产生的SPH2相同的条带。PAP1在添加PAP1的血浆中切割位点的氨基末端的一个键处部分水解前HP6和前HP8。PAP1不会直接激活前PAP1。这些结果表明，前酚氧化酶激活系统中存在一种自我强化机制，并且需要其他血浆蛋白在正确位点切割前HP6和前HP8以增强防御反应，这可能发生在途径激活的早期阶段。
