Brown planthopper (BPH) Nilaparvata lugens Stål is a serious insect pest of rice in Asian countries. Active compounds have close relationship with rice resistance against BPH. In this study, HPLC, MS/MS, and NMR techniques were used to identify active compounds in total flavonoids of rice. As a result, a BPH resistance-associated compound, Peak 1 in HPLC chromatogram of rice flavonoids, was isolated and identified as schaftoside. Feeding experiment with artificial diet indicated that schaftoside played its role in a dose dependent manner, under the concentration of 0.10 and 0.15 mg mL-1, schaftoside showed a significant inhibitory effect on BPH survival (p < 0.05), in comparison with the control. The fluorescent spectra showed that schaftoside has a strong ability to bind with NlCDK1, a CDK1 kinase of BPH. The apparent association constant KA for NlCDK1 binding with schaftoside is 6.436 × 103 L/mol. Docking model suggested that binding of schaftoside might affect the activation of NlCDK1 as a protein kinase, mainly through interacting with amino acid residues Glu12, Thr14 and Val17 in the ATP binding element GXGXXGXV (Gly11 to Val18). Western blot using anti-phospho-CDK1 (pThr14) antibody confirmed that schaftoside treatment suppressed the phosphorylation on Thr-14 site of NlCDK1, thus inhibited its activation as a kinase. Therefore, this study revealed the schaftoside-NlCDK1 interaction mode, and unraveled a novel mechanism of rice resistance against BPH.
Autophagy plays multiple roles in regulating various physiological processes in cells. However, we currently lack a systematic analysis of autophagy and the autophagyrelated gene 1 ATG1 in the brown planthopper (BPH, Nilaparvata lugens), one of the most destructive of the insect pests of rice. In this study, the full-length cDNA of an autophagy-related gene, NlATG1, was cloned from BPH. Real-time qPCR (RT-qPCR) revealed that this NlATG1 gene was expressed differently across developmental stages, at higher levels in nymphs but lower levels in adults. RNA interference with dsNlATG1 significantly decreased the mRNA level of the target gene to 14.6% at day 4 compared with that of the dsGFP control group. The survival of the dsNlATG1-treated group decreased significantly from day 4 onward, dropping to 48.3% on day 8. Examination using transmission electron microscopy (TEM) showed that epithelial cells of the BPH's midgut in the dsNlATG1-treated group had less autophagic vacuoles than did the dsGFP control, and knockdown of NlATG1 clearly inhibited the starvation-induced autophagy response in this insect. RNA interference of NlATG1 upregulated the NlFis1 gene involved in mitochondrial fission, leading to reductions in mitochondrial width and area. Furthermore, knockdown of NlATG1 also decreased the ATP content and accumulation of glycogen. Together, these results demonstrate that the NlATG1 gene participates in regulating autophagy and fission of mitochondria in the brown planthopper, making it a potentially promising target for pest control given its key role in autophagy, including maintaining the normal structure and function of mitochondria.
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