A M35 family metalloprotease is required for fungal virulence against insects by inactivating host prophenoloxidases and beyond
A M35 family metalloprotease is required for fungal virulence against insects by inactivating host prophenoloxidases and beyond
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DOI：
10.1080/21505594.2020.1731126
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发表时间：
2020-01-01
期刊：
影响因子：
5.2
通讯作者：
Wang, Chengshu
中科院分区：
文献类型：
--
作者：
Huang, Antian;Lu, Mengting;Wang, Chengshu
真核生物中分布着一个不同的金属蛋白酶家族。然而，对MPS的功能研究还不够深入。我们报道了昆虫致病真菌绿僵菌的基因组中编码了7个属于M35家族的MPS。通过基因缺失和昆虫生物测定，我们发现M35家族中的一个MPS，即Mr M35-4，是真菌对昆虫寄主的毒力所必需的。MRM35-4是一种可分泌的酶，具有促进真菌穿透昆虫角质层的蛋白分解活性。经过基因挽救和过表达后，昆虫生物检测表明，Mr M35-4有助于抑制昆虫角质层和血细胞的黑化活性。酶切实验表明，重组果蝇PPO1和PPO2能被Mr M35-4以不同于激活PPO活性的丝氨酸蛋白酶的方式被切割。此外，还发现Mr M35-4参与了抑制昆虫中抗真菌基因的表达。与Mr M35-4对宿主细胞明显的促凋亡作用相一致，我们发现PPO突变果蝇对罗伯茨分枝杆菌野生型和突变株的感染有不同程度的死亡。因此，除了在真菌-昆虫相互作用中靶向PPO之外，Mr M35-4还发挥着多方面的作用，这代表了绿僵菌以前不曾被怀疑的策略，以避开昆虫的免疫防御。
A diverse family of metalloproteases (MPs) is distributed in eukaryotes. However, the functions of MPs are still understudied. We report that seven MPs belonging to the M35 family are encoded in the genome of the insect pathogenic fungus Metarhizium robertsii. By gene deletions and insect bioassays, we found that one of the M35-family MPs, i.e. MrM35-4, is required for fungal virulence against insect hosts. MrM35-4 is a secretable enzyme and shows a proteolytic activity implicated in facilitating fungal penetration of insect cuticles. After gene rescue and overexpression, insect bioassays indicated that MrM35-4 contributes to inhibiting insect cuticular and hemocyte melanization activities. Enzymatic cleavage assays revealed that the recombinant prophenoloxidases PPO1 and PPO2 of Drosophila melanogaster could be clipped by MrM35-4 in a manner differing from a serine protease that can activate PPO activities. In addition, it was found that MrM35-4 is involved in suppressing antifungal gene expression in insects. Consistent with the evident apoptogenic effect of MrM35-4 on host cells, we found that the PPO mutant flies differentially succumbed to the infections of the wild-type and mutant strains of M. robertsii. Thus, MrM35-4 plays a multifaceted role beyond targeting PPOs during fungus-insect interactions, which represents a previously unsuspected strategy employed by Metarhizium to outmaneuver insect immune defenses.