[Identification of SPS family genes and regulation of photosynthetic pigment stability by PthSPS1/2/3 in Populus cv.741].

Solanesyl diphosphate synthases (SPSs), are key enzymes in the isoprenoid metabolic pathway, catalyze the biosynthesis of solanesyl diphosphate (SPP). In plants, the SPS family includes geranyl diphosphate synthase (GPS), farnesyl diphosphate synthase (FPS), geranylgeranyl diphosphate synthase (GGPPS), and SPS, catalyzing the stepwise condensation to form SPP. SPP is involved in the biosynthesis of solanesol and plastoquinone (PQ), both of which play important roles in plant growth, development, and the regulation of photosynthesis. Here, to elucidate the functions of the SPS subfamily in woody plants, we took 'Poplar 741' [Populus. alba×(Populus. davidiana+Populus. simonii)×Populus. tomentosa] as the experimental material. A total of 29 family members distributed across four subfamilies-PthSPS/PthFPS/PthGPS/PthGGPPS-related to the isoprenoid pathway were identified at the genome-wide level, located on 8 chromosome groups. Among them, 24 members were localized to chloroplasts and 5 to the cytoplasm. Three highly homologous PthSPSs were predicted as unstable hydrophilic proteins in chloroplasts. Subsequently, PthSPS1/2/3-edited mutant regenerated plants of 'Poplar 741' were generated via the CRISPR/Cas9 system. Phenotypic observation revealed three phenotypes: albino, green, and chimeric. PCR identification and sequencing analysis confirmed insertions and deletions (indels) at the target sites of PthSPS1/2/3. Physiological and biochemical analyses showed that the content of SPP, chlorophyll a, chlorophyll b, and carotenoids were significantly lower in the albino mutants (pthsps1/2/3) than in the wild type, indicating that PthSPS1/2/3 regulated the stability of photosynthetic pigments. These results provide a new direction for further dissecting the roles of the isoprenoid metabolic pathway in photosynthesis, growth, and development of woody plants.