KK-42对日本沼虾表皮蛋白基因MnCP-1表达的上调效应

为探讨KK-42显著缩短日本沼虾(Macrobrachium nipponense)蜕皮周期的可能机制,本研究采用RACE技术首次从头胸甲中获得了一个含几丁质结合-4(chitin_bind_4)结构域的表皮蛋白(cuticle proteins,CPs)c DNA基因全长,命名为Mn CP-1。该序列全长604 bp,可编码122个氨基酸。氨基酸序列比对显示,Mn CP-1与普通黄道蟹(Cancer pagurus,P81576.1)相似性最高(61%)。系统进化分析发现,Mn CP-1与地中海实蝇(Ceratitis capitata)聚类为一支。Real-time PCR结果显示,头胸甲中Mn CP-1在蜕皮前期晚期(D3-4期)和蜕皮后期(A期)的表达量较高,而在蜕皮间期(C期)和蜕皮前期早期(D0-2期)的表达量较低。KK-42处理能显著上调C期和D0-2期Mn CP-1的表达,在处理后6、24和48 h,其mRNA表达水平较对照组高3倍以上。结果表明,头胸甲上皮细胞Mn CP-1的表达与蜕皮周期有关,KK-42可显著上调C期和D0-2期Mn CP-1的表达。

Effects of KK-42 on up-regulation of cuticle protein gene MnCP-1 of Macrobrachium nipponense

In order to investigate the possible mechanism of KK-42 significant shortening the duration of molting cycle of Macrobrachium nipponense, a gene containing chitin bind 4 conserved domain called MnCP-1 was firstly cloned in car-apace from M.nipponense.The full length cDNA of MnCP-1 was 604 bp and encoded a putative protein of 122 amino acid residues.Amino acid sequence alignment results showed that MnCP-1 had the highest similarity of 61% with a cuticle pro-tein(CP)from Cancer pagurus.Phylogenetic analysis showed that a CP from Ceratitis capitata was the most closely related to MnCP-1.The results of Real-time PCR showed MnCP-1 mRNA expression levels were higher during late premolt stage (stage D3-4) and postmolt stage (stage A)while lower during intermolt stage (stage C)and early premolt stage (stage D0-2) in carapace.In addition, the expression levels during stage C and D0-2were increased by more than 3 times compared with the control at 6, 24 and 48 h after KK-42 treatment, respectively.Consequently, this research suggested that the expression of MnCP-1 in epithelial cell from carapace was relevant to the molting cycle.Furthermore, KK-42 could significantly up-regulate the expression of MnCP-1 during stage C and D0-2.