The roles of serine protease, intracellular and extracellular phenoloxidase in activation of prophenoloxidase system, and characterization of phenoloxidase from shrimp haemocytes induced by lipopolysaccharide or dopamine  

作　　者：谢鹏 潘鲁青 徐武杰 岳峰 

机构地区：[1]Feed Science Institute, Zhejiang University [2]Key Laboratory of Mariculture, Ministry of Education, Ocean University of China

出　　处：《Chinese Journal of Oceanology and Limnology》2013年第5期1018-1027,共10页中国海洋湖沼学报（英文版）

基　　金：Supported by the New Century Excellent Talents in University(No.NCET-06-0597);Introducing Talents of Discipline of Universities(111Project)(No.B08049)

摘　　要：We investigated the effects of lipopolysaccharide （LPS） and dopamine （DA） on the activation of the prophenoloxidase （proPO） system of Litopenaeus vannamei. LPS and DA were shown with a negative dose-dependent effect on hyalne cells （HC）, semi-granular cells （SGC）, large granular cells （LGC）, and total haemocyte count （THC）. When haemocytes were treated with LPS or DA, serine proteinase activity and intracellular phenoloxidase （PO） activity were significantly reduced, but extracellular PO activity increased significantly. These findings indicated that the reduction in haemocyte counts was mainly because of the degranulation and activation of the proPO system from semi-granule and large granule cells. The PKC inhibitor, chelerythrine, and the TPK inhibitor, genistein, had an inhibitory effect on extracellular PO activity, while serine proteinase and intracellular PO activity increased. This suggests that the LPS and DA induce the activation of proPO in haemocytes via PKC and TPK-related signaling pathways, but serine proteinase may be activated only by PKC, as the genistein effects were not statistically significant. Electrophoresis analysis revealed that POs induced by LPS or DA have the same molecular mass and high diphenolase activity. Two PO bands at 526 kDa and 272 kDa were observed in PAGE, while in the haemocyte lysate supematant （HLS）, only a 272-kDa band was observed. This band was resolved after SDS-PAGE under non-reducing and reducing conditions into two groups of POs, 166 kDa and 126 kDa, and 78.1 kDa and 73.6 kDa, respectively, suggesting that PO in L. vannamei is an oligomer, which may have different compositions intra- and extracellularly.We investigated the effects of lipopolysaccharide(LPS) and dopamine(DA) on the activation of the prophenoloxidase(proPO) system of Litopenaeus vannamei.LPS and DA were shown with a negative dose-dependent effect on hyalne cells(HC),semi-granular cells(SGC),large granular cells(LGC),and total haemocyte count(THC).When haemocytes were treated with LPS or DA,serine proteinase activity and intracellular phenoloxidase(PO) activity were significantly reduced,but extracellular PO activity increased significantly.These findings indicated that the reduction in haemocyte counts was mainly because of the degranulation and activation of the proPO system from semi-granule and large granule cells.The PKC inhibitor,chelerythrine,and the TPK inhibitor,genistein,had an inhibitory effect on extracellular PO activity,while serine proteinase and intracellular PO activity increased.This suggests that the LPS and DA induce the activation of proPO in haemocytes via PKC and TPK-related signaling pathways,but serine proteinase may be activated only by PKC,as the genistein effects were not statistically significant.Electrophoresis analysis revealed that POs induced by LPS or DA have the same molecular mass and high diphenolase activity.Two PO bands at 526 kDa and 272 kDa were observed in PAGE,while in the haemocyte lysate supernatant(HLS),only a 272-kDa band was observed.This band was resolved after SDS-PAGE under non-reducing and reducing conditions into two groups of POs,166 kDa and 126 kDa,and 78.1 kDa and 73.6 kDa,respectively,suggesting that PO in L.vannamei is an oligomer,which may have different compositions intra-and extracellularly.

关 键 词：lipopolysaccharide （LPS） dopamine （DA） Litopenaeus vannamei phenoloxidase （PO） signaling pathway 

分 类 号：S917.4[农业科学—水产科学]