Starch Degradation and Soluble Sugar Accumulation Are Associated with Cold Tolerance in Winter Rapeseed (Brassica...

Winter rapeseed suffers severe damage under cold stress, which seriously restricts its growth and yield. However, differences among cultivars in leaf starch and soluble sugar, as well as their underlying molecular regulatory mechanisms under low-temperature stress, remain poorly understood. We hypothesized that winter rapeseed cultivars with differences in cold tolerance would vary in starch accumulation, soluble sugar contents, and expression of the starch and sugar metabolism-related genes under cold stress. This study aimed to investigate differences in starch accumulation, soluble sugar content, and the gene expression patterns involved in the starch and sugar metabolism pathway in the leaves of winter rapeseed (Brassica napus L.) cultivars with contrasting cold tolerance under cold stress. In this study, two winter rapeseed cultivars, cold-tolerant L88 and cold-sensitive T2288, received 24 h cold treatments at 22 °C (control), 4 °C, 0 °C, and -4 °C. Plant morphology and leaf ultrastructure were observed, iodine-potassium iodide (I₂-KI) staining was performed, and the starch, soluble sugar contents (sucrose, fructose, glucose, and maltose) were analyzed for each temperature regime. Transcriptome sequencing was performed exclusively at -4 °C (0 h versus 24 h). Under -4 °C stress, the two cultivars showed the greatest differences in plant phenotype and leaf ultrastructure, with the cold-sensitive cultivar Tianyou 2288 suffering more severe freezing injury. In Longyou 88, chloroplasts in mesophyll cells were swollen and contained fewer starch granules, whereas in Tianyou 2288 some chloroplasts disintegrated and large numbers of starch granules remained undegraded. As the treatment temperature decreased, leaf starch content declined in both cultivars. At -4 °C, starch content in Longyou 88 was 3.84 mg/g lower than in Tianyou 2288. The contents of soluble sugars, sucrose, glucose, and maltose in leaves increased continuously in both cultivars as stress temperature decreased and reached their maximum values at -4 °C. However, fructose content showed cultivar-specific changes. In Longyou 88, fructose content reached its maximum value of 12.59 mg/g at -4 °C, whereas in Tianyou 2288 it peaked at 7.54 mg/g at 4 °C. At -4 °C, fructose content was 6.69 mg/g higher in Longyou 88 than in Tianyou 2288. Transcriptome analysis revealed that after 24 h of -4 °C stress, Longyou 88 had more upregulated differentially expressed genes (DEGs) than Tianyou 2288, whereas the number of downregulated DEGs in Tianyou 2288 was approximately twice that in Longyou 88. The genes BAM1, BAM3, SUS1, SCRK1, TPS9, and E1314, which were involved in the starch and sucrose metabolism pathway, were upregulated in both cultivars, whereas BGL44 was downregulated. These findings indicate that cold-tolerance winter rapeseed is associated with enhanced starch degradation, maintenance of chloroplast structural integrity in mesophyll cells, and increased soluble sugar accumulation, providing potential physiological indicators for cold-tolerance evaluation.