Division of Plant Sciences, University of Missouri, Columbia, MO, USA.
Division of Animal Sciences, University of Missouri, Columbia, MO, USA.
Insect Mol Biol. 2021 Aug;30(4):436-445. doi: 10.1111/imb.12709. Epub 2021 May 30.
Alternative splicing is a common feature in eukaryotes that not only increases the transcript diversity, but also has functional consequences. In insects, alternative splicing has been found associated with resistance to pesticides and Bt toxins. Up to date, the alternative splicing in western corn rootworm (Diabrotica virgifera virgifera LeConte) has not been studied. To investigate its alternative splicing pattern and relation to Bt resistance, we carried out single-molecule real-time (SMRT) transcript sequencing and Iso-seq analysis on resistant, eCry3.1Ab-selected and susceptible, unselected, western corn rootworm neonate midguts which fed on seedling maize with and without eCry3.1Ab for 12 and 24 h. We present transcriptome-wide alternative splicing patterns of western corn rootworm midgut in response to feeding on eCry3.1Ab-expressing corn using a comprehensive approach that combines both RNA-seq and SMRT transcript sequencing techniques. The results showed genes in western corn rootworm are highly alternatively spliced, which happens on 67.73% of multi-exon genes. One of the alternative splicing events we identified was a novel peritrophic matrix protein with two alternative splicing isoforms. Analysis of differential exon usage between resistant and susceptible colonies showed that in eCry3.1Ab-resistant western corn rootworm, expression of one isoform was significantly higher than in the susceptible colony, while no significant differences between colonies were observed with the other isoform. Our results provide the first survey of alternative splicing in western corn rootworm and suggest that the observed alternatively spliced isoforms of peritrophic matrix protein may be associated with eCry3.1Ab resistance in western corn rootworm.
可变剪接是真核生物中的一种常见特征，它不仅增加了转录本的多样性，而且具有功能后果。在昆虫中，可变剪接与对杀虫剂和 Bt 毒素的抗性有关。迄今为止，尚未对西部玉米根虫（Diabrotica virgifera virgifera LeConte）的可变剪接进行研究。为了研究其可变剪接模式及其与 Bt 抗性的关系，我们对具有抗性、eCry3.1Ab 选择和敏感、未选择的西部玉米根虫幼虫的肠道进行了单分子实时（SMRT）转录测序和 Iso-seq 分析，这些幼虫在食用含和不含 eCry3.1Ab 的幼苗玉米 12 和 24 小时后。我们使用结合 RNA-seq 和 SMRT 转录组测序技术的综合方法，展示了西部玉米根虫肠道对 eCry3.1Ab 表达玉米的转录组范围内的可变剪接模式。结果表明，西部玉米根虫中的基因高度可变剪接，这种情况发生在 67.73%的多外显子基因上。我们鉴定的可变剪接事件之一是一种新型的围食膜蛋白，具有两种可变剪接异构体。对抗性和敏感品系之间差异外显子使用的分析表明，在 eCry3.1Ab 抗性西部玉米根虫中，一种异构体的表达明显高于敏感品系，而另一种异构体在品系之间没有观察到显著差异。我们的研究结果首次提供了西部玉米根虫中可变剪接的调查结果，并表明观察到的围食膜蛋白的可变剪接异构体可能与西部玉米根虫中的 eCry3.1Ab 抗性有关。
