Expression of spider silk in tobacco improves drought tolerance with minimal effects on its mechanotype

Spider silk, especially dragline silk from Trichonephila clavipes, is an excellent natural material with remarkable mechanical properties. In this study, we generated nucleus- and plastid-encoded expression of a partial sequence of one of the components of dragline silk, major ampullate spidroin-1 (MaSp1), in tobacco (MaSp1-tobacco), MaSp1nuc and MaSp1pla, respectively. The partial sequence of MaSp1 encodes six repetitive glycine-rich and polyalanine tandem domains, and we evaluated the effect of its expression on plant mechanical properties and physiology. While MaSp1 expression showed no effect on leaf mechanical properties, it conferred drought tolerance. MaSp1-tobacco demonstrated an increased growth on PEG-infused growth medium and higher drought recovery in soil compared to the non-transgenic control (WT). Transcriptome analyses of drought-stressed MaSp1-tobacco revealed an upregulation of genes involved in stress response, antioxidant activity, and phenylpropanoid biosynthesis, with a higher effect of drought on the MaSp1pla transcriptome. Genes encoding peroxidase, stress and channel proteins, and cellular metabolism and homeostasis were consistently upregulated in these lines, which also displayed a higher total antioxidant status (TAS) compared to WT, and a higher TAS in MaSp1pla compared to MaSp1nuc. Under normal watering conditions, the ABA content and the TAS in MaSp1-tobacco were significantly higher than in the WT. Our results show that nucleus- and plastid-encoded expression of MaSp1 in tobacco increased the endogenous ABA levels, which led to the formation of plant stress memory that caused overall adjustments in multiple drought tolerance mechanisms and hence, conferred drought tolerance.

蜘蛛丝，尤其是来自棒络新妇蛛(Trichonephila clavipes)的拖丝，是一类具备优异力学性能的天然优质材料。本研究分别在烟草中实现了拖丝核心组分——大壶状腺丝蛋白-1(major ampullate spidroin-1, MaSp1)部分序列的核编码与质体编码表达，对应转基因株系分别命名为MaSp1-tobacco、MaSp1nuc与MaSp1pla。MaSp1的该部分序列共编码6个富含甘氨酸与聚丙氨酸的串联重复结构域，本研究评估了该序列的表达对植株力学特性与生理状态的影响。研究发现，尽管MaSp1的表达对叶片力学性能无显著影响，但却赋予了烟草耐旱性。与非转基因野生型(WT)对照相比，MaSp1-tobacco株系在聚乙二醇(PEG)浸润的培养基中生长量更高，且土壤干旱后的恢复能力更强。对干旱胁迫下的MaSp1-tobacco进行转录组分析后发现，参与胁迫响应、抗氧化活性与苯丙烷生物合成的基因均出现上调，且干旱对MaSp1pla株系的转录组调控效应更为显著。编码过氧化物酶、胁迫相关蛋白与通道蛋白，以及参与细胞代谢与稳态维持的基因在两类转基因株系中均持续上调；同时，转基因株系的总抗氧化态(total antioxidant status, TAS)显著高于野生型，且MaSp1pla株系的TAS水平高于MaSp1nuc株系。在正常浇水条件下，MaSp1-tobacco株系的脱落酸(abscisic acid, ABA)含量与总抗氧化态均显著高于野生型。本研究结果表明，在烟草中通过核编码与质体编码途径表达MaSp1，可提升植株内源脱落酸水平，进而形成植物胁迫记忆，对多种耐旱机制进行整体调控，最终赋予植株耐旱性。