Sun L N, Zhang H J, Quan L F, Yan W T, Yue Q, Li Y Y, Qiu G S
Research Institute of Pomology, Chinese Academy of Agricultural Sciences, Xingcheng, Liaoning 125100, People's Republic of China (
J Insect Sci. 2016 Feb 5;16(1). doi: 10.1093/jisesa/iev148. Print 2016.
The ryanodine receptor (RyR), the largest calcium channel protein, has been studied because of its key roles in calcium signaling in cells. Insect RyRs are molecular targets for novel diamide insecticides. The target has been focused widely because of the diamides with high activity against lepidopterous pests and safety for nontarget organisms. To study our understanding of effects of diamides on RyR, we cloned the RyR gene from the oriental fruit moth, Grapholita molesta, which is the most serious pest of stone and pome tree fruits throughout the world, to investigate the modulation of diamide insecticides on RyR mRNA expression in G. molesta (GmRyR). The full-length cDNAs of GmRyR contain a unique 3'-UTR with 625 bp and an open reading frame of 15,402 bp with a predicted protein consisting of 5,133 amino acids. GmRyR possessed a high level of overall amino acid homology with insect and vertebrate isoforms, with 77-92% and 45-47% identity, respectively. Furthermore, five alternative splice sites were identified in GmRyR. Diagnostic PCR showed that the inclusion frequency of one optional exon (f) differed between developmental stages, a finding only found in GmRyR. The lowest expression level of GmRyR mRNA was in larvae, the highest was in male pupae, and the relative expression level in male pupae was 25.67 times higher than that of in larvae. The expression level of GmRyR in the male pupae was 8.70 times higher than in female pupae, and that in male adults was 5.70 times higher than female adults.
兰尼碱受体（RyR）是最大的钙通道蛋白，因其在细胞钙信号传导中的关键作用而受到研究。昆虫RyR是新型双酰胺类杀虫剂的分子靶标。由于双酰胺类化合物对鳞翅目害虫具有高活性且对非靶标生物安全，该靶标受到了广泛关注。为了研究我们对双酰胺类化合物对RyR影响的理解，我们从东方果实蛾（Grapholita molesta）中克隆了RyR基因，东方果实蛾是全世界核果类和仁果类果树最严重的害虫，以研究双酰胺类杀虫剂对东方果实蛾（GmRyR）中RyR mRNA表达的调节作用。GmRyR的全长cDNA包含一个独特的625 bp的3'-UTR和一个15,402 bp的开放阅读框，预测的蛋白质由5,133个氨基酸组成。GmRyR与昆虫和脊椎动物异构体具有高度的整体氨基酸同源性，分别具有77 - 92%和45 - 47%的同一性。此外，在GmRyR中鉴定出五个可变剪接位点。诊断性PCR表明，一个可选外显子（f）的包含频率在不同发育阶段有所不同，这一发现仅在GmRyR中发现。GmRyR mRNA的最低表达水平在幼虫期，最高在雄性蛹期，雄性蛹期的相对表达水平比幼虫期高25.67倍。GmRyR在雄性蛹中的表达水平比雌性蛹高8.70倍，在雄性成虫中的表达水平比雌性成虫高5.70倍。
