Fruit Important Quality Formation And The Molecular Mechanisms In Response To Elevated CO₂ In Lycium Barbarum

Goji berry（Lycium barbarum L.）,is an economically important forest tree species of medicinal and food origin,it has a long cultivation history,excellent adaptability,and rich various nutritional composition,with high economic and medicinal values.As a characteristic economic forest tree species,after decades of large-scale development,the cultivation area continues to expand,the deep processing technology has become increasingly mature,which forms a unique market economic model,and it has obvious economic,ecological,and social benefits in promoting regional economic development,enhancing farmers’livelihoods,and pharmaceutical research and development application.The fruits of goji berries contain a variety of bioactive and pharmacological components,among which sugars are the essential determinants of nutritional quality.At present,with the continuous expansion of the cultivation scale and the in-depth development and utilization of effective substances of goji berry,the production of high-quality fruits and the cultivation of excellent cultivars have become the challenge facing the current industrial development.Especially,elevated CO₂ has significantly affected the accumulation of nutrients in goji berries.Based on the industrial development of goji berry and future climate change scenarios,this study comprehensively investigates the formation features of goji berry fruit quality and its response mechanisms to climate change,particularly for elevated CO₂.These findings provide a basis and reference for cultivating high-quality goji berries and adapting economic forests to climate change.The main research conclusions are as follows.（1）The model for the goji berry fruit growth patterns was established for five cultivars,based on the logistic growth model,the fruit growth period was divided into three phases（Ⅰ,Ⅱ,andⅢ）using two key time points.The fruit’s important nutritional constituents,galactose,fructose,and total sugar,increased until the RF stage（p<0.05）,whereas the polysaccharide content was the highest in the YF stage（p<0.05）,followed by a decrease up to the CF stage and an increase in the RF stage.The sucrose,betaine,and flavonoid contents were found to be the highest in the GF stage and decreased until the RF stage（p<0.05）.Based on the fruit growth curve model and the measurements of nutritional constituents,the polysaccharides accumulated mainly in phases I and III;sucrose,betaine,and flavonoids accumulated in phase II,and galactose,fructose,and total sugar accumulated in phase III.Hence,it can be considered that the two key time points for the goji berry fruit development and accumulation of nutrients are 10–13 days and 21–26 days after full bloom,during which enhanced water and fertilizer management should be provided.The total sugar and polysaccharide were considered the two main evaluation factors.Among the five cultivars,‘Ningqi 7’is the most advantageous with respect to morphological and nutritional quality characteristics.（2）Integrated nutritional components index and multi-omics data of three growth stages to analyze the metabolic characteristics of goji berry（‘Ningqi 1）development.Two co-expression network modules were identified that were significantly associated with galactose,total sugars and polysaccharide components（r>0.95,p<0.01）.These modules analysis identified the Lb SPS and Lb EMP24 are hub gene and protein strongly correlated with DEMs,DEPs,and DEGs.Also,the expression patterns analysis revealed a strongly correlation between the gene Lb SPS and nine transcription factors（r>0.96,p<0.01）.The identified 42 DEGs,43 DEPs,and 5 DEMs are jointly involved in regulating five metabolism pathways,and D-glucose acts as a substrate in glycolysis to promote the biosynthesis of various sugars under the co-regulation of the genes and their corresponding encoded enzymes.Additionally,there were strong protein-protein interactions（Score>0.95）among 19 DEPs.These DEMs,DEPs,and DEGs involved in this process serve as key regulators in carbohydrate conversion biosynthesis.（3）The effects of elevated CO₂ concentration and elevated temperature on fruit morphological traits and nutrient accumulation of three goji berry cultivars were investigated separately.The results showed that elevated temperature significantly increased the fruit weight and promoted the growth of the fruit’longitudinal and transverse diameter of‘Ningqi1’and‘Ningqi7’（p<0.05）.Elevated CO₂ concentration significantly increased the fruit diameter and fruit weight of‘Ningqi1’.Overall,elevated temperature and CO₂ concentration promoted the accumulation of fruit’s major nutrient components,the elevated CO₂ concentration variation had a greater effect on the accumulation of glucose and polysaccharide contents for‘Ningqi 1’.（4）Based on the effect of elevated CO₂ concentration on the accumulation of sugar compounds in goji berry,the gene Lb GALA,which is involved in sugar metabolism in response to elevated CO₂,its gene family was identified and analyzed the characterization,and further the full-length sequences of key members Lb GALA1-Lb GALA4 were cloned.Subcellular localization and bimolecular fluorescence analysis revealed that Lb GALA1-Lb GALA4 are found in the plasma membrane and cytoplasm,and Lb GALA1 and Lb GALA3 protein interactions;overexpression of Lb GALA1-Lb GALA4 in tomato was found to be significantly increased and significantly correlated with sugar fraction.A significant correlation between Lb GALA expression and galactose in goji berries and an increase in fruit sugars in T1 transgenic tomato by overexpressing Lb GALA1-Lb GALA4 under elevated CO₂ levels confirm their primary role and response to elevated CO₂ in sugar metabolism.Chromosomal localization analysis showed that these genes were mainly located on four chromosomes,and the genes Lb GALA4 and Lb GALA5 were a pair of tandem repeats.Analysis of the promoter cis-acting elements identified key cis-acting regulatory elements associated with environmental stress.The key transcription factors binding to the promoter region of gene Lb GALA3 were identified,including Lb NAC,Lb TFIID,and Lb B3,and verified with Y1H.