The starch excess and key genes underlying citrus leaf chlorosis by rootstock-scion incompatibility

Leaf chlorosis caused by rootstock-scion incompatibility in citrus orchard badly affects fruit yield and quality. Starch excess and its key genes underlying citrus leaf chlorosis in incompatible graft remained unknown. Here, using created model incompatible/ compatible rootstock-scion combinations, we investigated starch content and distribution in 116 various chlorotic leaves of incompatible graft, and characterized the relationship between leaf chlorosis and starch accumulation. Further, we identified starch metabolism-related gene families by genome-wide analysis of pomelo genome, and performed comparative transcriptomic analysis on leaves. A total of nine key differentially expressed genes of starch metabolism were validated. Among them, seven starch synthesis-related genes were significantly upregulated, and two starch degradation-related genes, CgBAM4 and CgBAM6, were significantly downregulated. Meanwhile, the relative expression of synthesis-related genes was positively correlated with starch accumulation and leaf chlorosis. Using transient overexpression and VIGS experiments in pomelo, we confirmed the function of CgGBSS2, which was the only amylose synthesis-related key gene with the most significantly upregulated expression level. We proposed a working model to illustrate the regulatory network of starch excess accumulation involving in citrus leaf chlorosis of incompatible graft in the end. This study provides insights into the molecular mechanism underlying leaf chlorosis process in rootstock-scion incompatibility.