Choi Man-Yeon, Ahn Seung-Joon, Kim A Young, Koh Youngho
USDA-ARS, Horticultural Crops Research Laboratory, 3420 NW Orchard Avenue, Corvallis, OR 97330, USA.
USDA-ARS, Horticultural Crops Research Laboratory, 3420 NW Orchard Avenue, Corvallis, OR 97330, USA; Department of Crop and Soil Science, Oregon State University, Corvallis, OR 97331, USA.
Gen Comp Endocrinol. 2017 May 15;246:354-362. doi: 10.1016/j.ygcen.2017.01.011. Epub 2017 Jan 7.
The family of FXPRLamide peptides serves as a major insect hormone. It is characterized by a core active amino acid sequence conserved at the C-terminal ends, and provides various physiological roles across the Insecta. In this study we identified and characterized pyrokinin (PK) and CAPA cDNAs encoding two FXPRLamide peptides, pyrokinin and CAPA-DH (diapause hormone), and two corresponding G protein-coupled receptors (GPCRs) from spotted wing drosophila (SWD), Drosophila suzukii. Expressions of PK and CAPA mRNAs were differentially observed during all life stages except the embryo, and the detection of CAPA transcription was relatively strong compared with the PK gene in SWD. Both D. suzukii pyrokinin receptor (DrosuPKr) and CAPA-DH receptor (DrosuCAPA-DHr) were functionally expressed and confirmed through binding to PK and DH peptides. Differential expression of two GPCRs occurred during all life stages; a strong transcription of DrosuPKr was observed in the 3rd instar. DrosuCAPA-DHr was clearly expressed from the embryo to the larva, but not detected in the adult. Gene regulation during the life stages was not synchronized between ligand and receptor. For example, SWD CAPA mRNA has been up-regulated in the adult while CAPA-DHr was down-regulated. The difference could be from the CAPA mRNA translating multiple peptides including CAPA-DH and two CAPA-PVK (periviscerokinin) peptides to act on different receptors. Comparing the genes of SWD PK, CAPA, PKr and CAPA-DHr to four corresponding genes of D. melanogaster, SWD CAPA and the receptor are more similar to D. melanogaster than PK and the receptor. These data suggest that the CAPA gene could be evolutionally more conserved to have a common biological role in insects. In addition, the effect of Kozak sequences was investigated by the expression of the GPCRs with or without Kozak sequences in Sf9 insect cells. The Kozak sequenced PK receptor was significantly less active than the original (= no Kozak sequenced) receptor. Our results provide a knowledge for potential biological function(s) of PK and CAPA-DH peptides in SWD, and possibly offer a novel control method for this pest insect in the future.
FXPRLamide肽家族是一种主要的昆虫激素。其特征是在C末端具有保守的核心活性氨基酸序列，并在昆虫纲中发挥多种生理作用。在本研究中，我们从铃木果蝇（Spotted wing drosophila，SWD）中鉴定并表征了编码两种FXPRLamide肽——速激肽（PK）和CAPA - DH（滞育激素）的速激肽和CAPA cDNA，以及两种相应的G蛋白偶联受体（GPCRs）。除胚胎期外，在所有生命阶段均观察到PK和CAPA mRNA的差异表达，并且与SWD中的PK基因相比，CAPA转录的检测相对较强。铃木果蝇速激肽受体（DrosuPKr）和CAPA - DH受体（DrosuCAPA - DHr）均实现了功能表达，并通过与PK和DH肽结合得到证实。两种GPCRs在所有生命阶段均出现差异表达；在三龄幼虫中观察到DrosuPKr的强烈转录。DrosuCAPA - DHr从胚胎期到幼虫期均有明显表达，但在成虫中未检测到。配体和受体在生命阶段的基因调控并不同步。例如，SWD的CAPA mRNA在成虫中上调，而CAPA - DHr则下调。这种差异可能源于CAPA mRNA翻译出多种肽，包括CAPA - DH和两种CAPA - PVK（围脏速激肽）肽，从而作用于不同的受体。将SWD的PK、CAPA、PKr和CAPA - DHr基因与黑腹果蝇的四个相应基因进行比较，SWD的CAPA及其受体比PK及其受体与黑腹果蝇更相似。这些数据表明，CAPA基因在进化上可能更保守，在昆虫中具有共同的生物学作用。此外，通过在Sf9昆虫细胞中表达有无Kozak序列的GPCRs，研究了Kozak序列的影响。带有Kozak序列的PK受体活性明显低于原始（即无Kozak序列）受体。我们的结果为SWD中PK和CAPA - DH肽的潜在生物学功能提供了知识，并可能在未来为这种害虫提供一种新的控制方法。
