Impact of Jojoba wax ester synthase overexpression on drought performance and wood properties of Populus × canescens

Populus species, particularly Populus x canescens, are essential biomass trees due to their rapid growth rates. However, their susceptibility to drought due to high water consumption poses a significant challenge, especially in the face of climate change. In this thesis, I explore innovative biotechnological strategies to improve the water-saving abilities and wood properties of Populus species through genetic engineering. This thesis primarily aimed to explore the specific role of wax ester synthase in improving poplar performance and to assess how these modifications influence growth, biomass production, wood properties, and physiological performance. To achieve this, transgenic P. x canescens trees with elevated levels of wax esters in their vegetative tissues were used. For this assessment, we formulated two distinct hypotheses: (a) that increasing wax ester concentrations improves wood hydrophobicity, as evidenced by decreased wood swelling, thereby influencing a major technological feature of the wood; and (b) that wax ester accumulation in the cuticle strengthens the transgenic trees' resistance to drought. Two separate approaches were applied to examine these hypotheses. One approach involved overexpressing the jojoba wax ester synthase (ScWS) gene from Simmondsia chinensis in Populus x canescens to enhance cuticular properties with the ubiquitous 35S promoter. This manipulation led to lipid droplet accumulation within cells and resulted in significant shifts in surface wax composition. The content of the substance classes such as alkane, primary alcohol, and wax ester was reduced. Despite these changes, non-stomatal water loss remained unaffected. However, ScWS lines exhibited decreased stomatal conductance and increased water-use efficiency, delaying leaf shedding. Stomata phenotyping using microscopic techniques revealed shorter stomata lengths in 35S::ScWS lines. This phenotype was ascribed to a notable proportion of wax-occluded or semi-occluded stomata, thereby constraining poplar photosynthesis particularly under conditions of high light intensity. Occluded stomata were observed previously in Arabidopsis thaliana due to suppression of the expression of the Occluded Stomatal Pore1 (OSP1) gene. Here, the ScWS lines also showed suppressed OSP1 transcript bundances. . Increasing the expression of ScWS in P. x canescens triggered the suppression of the endogenous form of wax ester synthase, WSD4, which is the closest homologue to ScWS. However, the expression of WSD1 remained unchanged. Under Introduction 12 field conditions, leaf damage and insect scores were unaffected in the transgenic compared with the wildtype poplars. While biomass production was initially unaffected, long-term field experiments revealed a growth trade-off compared to wild-type poplars. In another approach, Populus x canescens plants overexpressing ScWS under the tissue-specific DX15 promoter from Populus trichocarpa were used. The aim of this transformation, utilizing a wood-specific promoter and the ScWS gene, was to improve wood properties and biomass production. Transgenic lines were cultivated separately in two distinct experiments, one conducted in a greenhouse over a period of 100 days, and the other in a climate chamber for 90 days. During these experiments, the growth and physiological performance of the transgenic lines were evaluated. Morphological features and growth parameters of DX15::ScWS lines were similar to those of wild-type plants. However, in the DX15::ScWS lines increased lipid droplet accumulation and enhanced wood water repellency was observed. Furthermore, the DX15::ScWS demonstrated an increased fiber cell wall thickness, which likely contributed to the higher wood density observed in these lines. The optimized distribution of biomass in DX15::ScWS lines directed more resources towards leaf and stem growth. These findings underscore the potential of combining biotechnological approaches to enhance both water-saving abilities and wood characteristics in Populus species. Manipulating the expression of ScWS gene in conjunction with tissue-specific promoters offers a promising strategy for genetic engineering aimed at improving biomass yield and wood quality in plants. This thesis contributes to the understanding of the intricate relationship between wax biosynthesis, stomatal regulation, and wood properties, paving the way for future advancements in sustainable biomass production and forestry management.

2025-08-18