A single-base mutation on the 5' UTR of BrATG5 confers the premature leaf senescence phenotype in Chinese cabbage.

Abstract

Key message: BrATG5 encoding autophagy protein was fine-mapped through MutMap and KASP analysis, and its function in regulating leaf senescence was verified using virus-induced gene silencing and functional complementation assays in Chinese cabbage. Leaf senescence is the final stage of leaf development, and is accompanied by the breakdown of organelle and catabolism of chlorophyll and macromolecules. The generated nutrients are supplied to developing seeds or other growing organs. However, premature leaf senescence will cause a decrease in the yield and quality of leafy head in Chinese cabbage. In this study, a premature leaf senescence mutant M1684 was screened from an ethyl methane sulfonate (EMS) mutagenized population of Chinese cabbage 'FT'. The outer whorl leaves of M1684 showed a premature senescence phenotype from seedling stage, and the yield of leafy head and seed was reduced in M1684. The chloroplast structure and photosynthetic activity of the leaves of M1684 were impaired and programmed cell death (PCD) occurred earlier in M1684 than in 'FT'. Genetic analysis, MutMap and kompetitive allele-specific PCR (KASP) genotyping showed that BraA10g022760.3.5C, which encodes an autophagy protein, was the candidate gene, named BrATG5. The function of BrATG5 in regulating leaf senescence was verified by virus-induced gene silencing (VIGS) and functional complementation assays. 5' rapid amplification of complementary DNA ends (5' RACE) clone, and quantitative real-time PCR (qRT-PCR) assays showed that the single nucleotide polymorphism on the 5' untranslated region (5' UTR) decreased the expression level of BrATG5. BrATG5 interacted with BrATG12 to form a complex. These results suggest that BrATG5 is involved in leaf senescence in Chinese cabbage, and provide a better understanding of the molecular mechanisms of leaf senescence and a clearer theoretical basis for anti-senescence research in Chinese cabbage.