Fera Science Ltd., Sand Hutton, York, YO41 1LZ, United Kingdom.
School of Biosciences, Durham University, South Road, Durham, DH1 3LE, United Kingdom.
Sci Rep. 2017 Nov 9;7(1):15147. doi: 10.1038/s41598-017-15534-y.
The Drosophila melanogaster (fruit fly) gene Diap1 encodes a protein referred to as DIAP1 (D rosophila Inhibitor of Apoptosis Protein 1) that acts to supress apoptosis in "normal" cells in the fly. In this study we investigate the use of RNA interference (RNAi) to control two dipteran pests, Musca domestica and Delia radicum, by disrupting the control of apoptosis. Larval injections of 125-500 ng of Diap1 dsRNA resulted in dose-dependent mortality which was shown to be attributable to down-regulation of target mRNA. Insects injected with Diap1 dsRNA have approx. 1.5-2-fold higher levels of caspase activity than controls 24 hours post injection, providing biochemical evidence that inhibition of apoptotic activity by the Diap1 gene product has been decreased. By contrast adults were insensitive to injected dsRNA. Oral delivery failed to induce RNAi effects and we suggest this is attributable to degradation of ingested dsRNA by intra and extracellular RNAses. Non-target effects were demonstrated via mortality and down-regulation of Diap1 mRNA levels in M. domestica larvae injected with D. radicum Diap1 dsRNA, despite the absence of 21 bp identical sequence regions in the dsRNA. Here we show that identical 15 bp regions in dsRNA are sufficient to trigger non-target RNAi effects.
黑腹果蝇（果蝇）基因 Diap1 编码一种称为 DIAP1（果蝇凋亡抑制蛋白 1）的蛋白质，该蛋白可抑制果蝇中“正常”细胞的凋亡。在这项研究中，我们通过破坏凋亡的控制来研究使用 RNA 干扰（RNAi）来控制两种双翅目害虫，家蝇和地种蝇，幼虫注射 125-500ng 的 Diap1 dsRNA 会导致剂量依赖性死亡，这归因于靶 mRNA 的下调。与对照相比，注射 Diap1 dsRNA 的昆虫在注射后 24 小时内具有约 1.5-2 倍更高的半胱天冬酶活性，提供了生化证据表明 Diap1 基因产物抑制凋亡活性的作用降低了。相比之下，成虫对注射的 dsRNA 不敏感。口服递送未能诱导 RNAi 效应，我们认为这归因于细胞内和细胞外 RNAse 对摄入的 dsRNA 的降解。尽管 dsRNA 中缺乏 21bp 相同的序列区域，但在家蝇幼虫中注射地种蝇 Diap1 dsRNA 会导致死亡率和 Diap1 mRNA 水平下调，从而证明了非靶效应。在这里，我们表明 dsRNA 中相同的 15bp 区域足以引发非靶 RNAi 效应。
