School of Basic Medicine and Biological Sciences, Soochow University, Suzhou, Jiangsu, China.
National Engineering Laboratory for Modern Silk, Soochow University, Suzhou, Jiangsu, China.
Arch Insect Biochem Physiol. 2020 May;104(1):e21664. doi: 10.1002/arch.21664. Epub 2020 Feb 24.
The high-temperature stress gene GrpE plays an important role in coping with high-temperature stress. The mutation of key sites of this gene can improve the high-temperature resistance of organisms. In the present study, using complementary DNAs from the silkworm fat body as the template, the open reading frame sequence of the GrpE gene (BmGrpE) was amplified and was found to be 644 bp in length and encode a protein with a predicted molecular weight of 24.1 kDa. The presence of a binding site for the heat shock transcription factor (Hsf1) at -1440 bp upstream of its coding region indicates that BmGrpE may respond to high-temperature stress. BmGrpE was constitutively expressed throughout developmental stages, with the highest level observed in the 5th instar larvae stage. Moreover, in 5th instar larvae (the 3th day), BmGrpE was expressed in all tissues examined, with the highest levels in the fat body, silk gland, and midgut. Interestingly, under high-temperature stress, TiO nanoparticle treatment increased the messenger RNA levels of BmGrpE in the fat body and silk gland. After treatment with dsRNA of BmGrpE, the cell viability of BmN cells was significantly decreased under 34°C and H O stress (p < .05). Mutation of BmGrpE (H163L) enhanced the resistance of BmN cells under high-temperature stress. These results provide new clues for the study of molecular mechanisms of insect resistance to high temperatures.
高温应激基因 GrpE 在应对高温应激中发挥重要作用。该基因关键位点的突变可以提高生物的耐高温能力。本研究以家蚕脂肪体 cDNA 为模板，扩增得到 GrpE 基因（BmGrpE）的开放阅读框序列，长度为 644bp，编码一个预测分子量为 24.1kDa 的蛋白。其编码区上游-1440bp 处存在热休克转录因子（Hsf1）结合位点，表明 BmGrpE 可能对高温胁迫产生响应。BmGrpE 在发育各阶段均持续表达，在 5 龄幼虫期表达量最高。此外，在 5 龄幼虫（第 3 天）中，BmGrpE 在检测到的所有组织中均有表达，在脂肪体、丝腺和中肠中表达量最高。有趣的是，在高温胁迫下，TiO2 纳米颗粒处理增加了脂肪体和丝腺中 BmGrpE 的信使 RNA 水平。用 BmGrpE 的 dsRNA 处理后，BmN 细胞在 34°C 和 H2O2 胁迫下的细胞活力显著降低（p<.05）。BmGrpE（H163L）的突变增强了 BmN 细胞在高温胁迫下的抗性。这些结果为研究昆虫耐高温的分子机制提供了新线索。
