Department of Comparative Physiology, Uppsala University, Norbyvägen 18A, SE-752 36 Uppsala, Sweden.
Dev Comp Immunol. 2011 Jan;35(1):51-61. doi: 10.1016/j.dci.2010.08.005. Epub 2010 Aug 22.
Recognition of microbial polysaccharide by pattern recognition receptors triggers the prophenoloxidase (proPO) cascade, resulting in melanin synthesis and its deposition on the surface of invading pathogens. Several masquerade-like proteins and serine proteinase homologues have been shown to be involved in the proPO activation in insects. In this study, a novel serine proteinase homologue, Pl-SPH2, was found and isolated as a 30kDa protein from hemocytes of the freshwater crayfish, Pacifastacus leniusculus, by its binding property to a partially lysozyme digested or TCA-treated insoluble Lysine (Lys)-type peptidoglycan (PGN) and soluble polymeric Lys-type PGN. Two other proteins, the Pl-SPH1 and lipopolysaccharide- and β-1,3-glucan-binding protein (LGBP) were also found in the several different PGN-binding assays. However no PGRP homologue was detected. Neither was any putative PGRP found after searching available crustacean sequence databases. If RNA interference of Pl-SPH2, Pl-SPH1 or LGBP in the crayfish hematopoietic tissue cell culture was performed, it resulted in lower PO activity following activation of the proPO-system by soluble Lys-type PGN. Taken together, we report for the first time that Lys-type PGN is a trigger of proPO-system activation in a crustacean and that two Pl-SPHs are involved in this activation possibly by forming a complex with LGBP and without a PGRP.
模式识别受体识别微生物多糖会触发原酚氧化酶（proPO）级联反应，导致黑色素合成并沉积在入侵病原体的表面。已经有几种伪装样蛋白和丝氨酸蛋白酶同源物被证明参与了昆虫中的 proPO 激活。在这项研究中，一种新型的丝氨酸蛋白酶同源物 Pl-SPH2 被发现并从淡水小龙虾（Pacifastacus leniusculus）的血细胞中分离出来，它通过与部分溶菌酶消化或 TCA 处理的不溶性赖氨酸（Lys）型肽聚糖（PGN）和可溶性聚合 Lys 型 PGN 的结合特性而被鉴定出来。在几种不同的 PGN 结合测定中，还发现了另外两种蛋白质 Pl-SPH1 和脂多糖和β-1,3-葡聚糖结合蛋白（LGBP）。然而，在搜索可用的甲壳动物序列数据库后，没有检测到任何 PGRP 同源物。如果在小龙虾造血组织细胞培养物中对 Pl-SPH2、Pl-SPH1 或 LGBP 进行 RNA 干扰，则在可溶性 Lys 型 PGN 激活 proPO 系统后，PO 活性会降低。总之，我们首次报道 Lys 型 PGN 是甲壳动物 proPO 系统激活的触发因素，两种 Pl-SPH 可能通过与 LGBP 形成复合物而参与这种激活，而无需 PGRP。
