Molecular characteristics, mRNA expression, and alternative splicing of a ryanodine receptor gene in the oriental fruit fly, Bactrocera dorsalis (Hendel)

• Published in: PloS one 2014-04, Vol.9 (4), p.e95199
• Main Authors: Yuan, Guo-Rui, Shi, Wen-Zhi, Yang, Wen-Jia, Jiang, Xuan-Zhao, Dou, Wei, Wang, Jin-Jun
• Format: Article
• Language: English
• Subjects: Agrochemicals; Alternative Splicing; Amino Acid Sequence; Amino acids; Analysis; Animals; Bactrocera dorsalis; Binding sites; Biology and Life Sciences; Body parts; Calcium; Calcium (intracellular); Calcium channels (ligand-gated); Chemicals; Conserved Sequence; Diagnostic systems; DNA, Complementary - genetics; DNA, Complementary - metabolism; Drosophila melanogaster; Engineering; Fruit-flies; Fruits; Gene expression; Gene Expression Regulation; Genetic aspects; Genetic engineering; Insect Proteins - genetics; Insect Proteins - metabolism; Insecticide resistance; Insecticides; Insects; Isoelectric Point; Isoforms; Laboratories; Larvae; Lysis; Mammals; Messenger RNA; Molecular modelling; Molecular Sequence Data; Molecular Weight; Musculoskeletal system; Open Reading Frames; Pest control; Phylogeny; Physiological aspects; Protein Structure, Tertiary; Pupa; Pupae; Receptors; RNA, Messenger - genetics; RNA, Messenger - metabolism; Rodents; Ryanodine Receptor Calcium Release Channel - genetics; Ryanodine Receptor Calcium Release Channel - metabolism; Ryanodine receptors; Sequence Alignment; Sequence Homology, Amino Acid; Structure-function relationships; Studies; Tephritidae - classification; Tephritidae - genetics; Tephritidae - metabolism; Thorax; Transmembrane domains; Trends; Vegetables
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0095199

Ryanodine receptors (RyRs) are a distinct class of ligand-gated channels controlling the release of calcium from intracellular stores. The emergence of diamide insecticides, which selectively target insect RyRs, has promoted the study of insect RyRs. In the present study, the full-length RyR cDNA (BdRyR) was cloned and characterized from the oriental fruit fly, Bactrocera dorsalis (Hendel), a serious pest of fruits and vegetables throughout East Asia and the Pacific Rim. The cDNA of BdRyR contains a 15,420-bp open reading frame encoding 5,140 amino acids with a predicted molecular weight of 582.4 kDa and an isoelectric point of 5.38. BdRyR shows a high level of amino acid sequence identity (78 to 97%) to other insect RyR isoforms. All common structural features of the RyRs are present in the BdRyR, including a well-conserved C-terminal domain containing consensus calcium-binding EF-hands and six transmembrane domains, and a large N-terminal domain. Quantitative real-time PCR analyses revealed that BdRyR was expressed at the lowest and highest levels in egg and adult, respectively, and that the BdRyR expression levels in the third instar larva, pupa and adult were 166.99-, 157.56- and 808.56-fold higher, respectively, than that in the egg. Among different adult body parts, the highest expression level was observed in the thorax compared with the head and abdomen. In addition, four alternative splice sites were identified in the BdRyR gene, with the first, ASI, being located in the central part of the predicted second spore lysis A/RyR domain. Diagnostic PCR analyses revealed that alternative splice variants were generated not only in a tissue-specific manner but also in a developmentally regulated manner. These results lay the foundation for further understanding the structural and functional properties of BdRyR, and the molecular mechanisms for target site resistance in B. dorsalis.