For a long time, scientists around the world have been trying to continuously improve crop yield and quality through various means. However, with global climate change and environmental degradation, the stress resistance of crops has attracted much attention. How to economically and quickly improve plant stress resistance has become a focus of scientists' attention. Most studies have shown a close relationship between plant stress resistance and epidermal wax, and people have also extracted epidermal wax from plants for processing in industries and cosmetics [1]. Therefore, designing methods to control epidermal wax synthesis is of great significance and a focus of attention in the field of synthetic biology. Our project uses Nicotiana benthamiana as the reactor for wax synthesis, thanks to the fact that Nicotiana benthamiana undergoes Agrobacterium mediated transient transformation, typically achieving the highest yield of target proteins or metabolites within 3-7 days. Moreover, the cells of Nicotiana benthamiana have a powerful endoplasmic reticulum and other inner membrane systems, which can efficiently synthesize, process, and fold exogenous proteins, which are crucial for the synthesis of various enzyme proteins required for wax (belonging to lipid compounds). The most important Nicotiana benthamiana plants have a short growth cycle (about 6-8 weeks), wide and thick leaves, and can provide a large amount of biomass for production, which is very suitable for large-scale promotion and application, providing a theoretical basis for us to cultivate high wax raw materials.
In the early stage, we discovered through extensive literature reading that there may be a key gene CER1 [2] regulating wax synthesis in most plants, and confirmed through experiments that the CER1 gene is also a key regulatory gene for wax synthesis in tobacco. Furthermore, through interaction experiments, we confirmed the existence and interaction of the light regulatory factor HY5 and CER1, which jointly mediate the wax synthesis pathway in tobacco plants. This provides reliable evidence for us to improve the wax content of plant epidermis by modifying light. We design to use different intensities of light to treat tobacco plants, detect the content of wax substances in the plant body, and obtain the most suitable light intensity for plant wax synthesis, in order to develop high wax plant raw materials.
It has been demonstrated through CRISPR-Cas9 technology that the CER1 gene in Nicotiana benthamiana plants can indeed regulate wax synthesis, and the loss of CER1 function leads to a significant decrease in wax content in the plant, indicating that CER1 plays a positive regulatory role in wax synthesis (Figure 1). Afterwards, we utilized the dual luciferase Luc reporter system and demonstrated through Dual Luc experiments that the light regulatory factor HY5 can bind to the promoter region of CER1 and drive its expression, promoting wax synthesis (Figure 2).
[1] Rafferty, D. W., Dupin, L., Zellia, J., & Giovannitti-Jensen, A. (2018). Predicting lipstick sensory properties with laboratory tests. International journal of cosmetic science, 40(5), 451–460.
[2] Zhu, L., Guo, J., Zhu, J., & Zhou, C. (2014). Enhanced expression of EsWAX1 improves drought tolerance with increased accumulation of cuticular wax and ascorbic acid in transgenic Arabidopsis. Plant physiology and biochemistry : PPB, 75, 24–35.