Expression Analysis of Ethylene-related Genes in Programmed Cell Death of Apical-bud-senescence Chestnut Buds

Purpose To investigate key candidate genes involved in ethylene (ET) synthesis and signal transduction during programmed cell death (PCD) in buds of apical-bud-senescence chestnut, and to elucidate the regulatory role of ET in bud PCD.

Methods Upper buds (undergoing PCD) and lower latent buds (non-PCD) from one-year-old shoots of apical-bud-senescence chestnut variety X12 were used as materials. ET release amount in both bud types was measured by gas chromatography at three stages (20, 25, and 30 days after flowering, representing non-PCD, PCD initiation, and PCD completion, respectively). Based on transcriptome sequencing and quantitative real-time PCR, the expression patterns of differentially expressed genes (DEGs) involved in ET synthesis and signal transduction were analyzed and validated, and key candidate genes mediating bud PCD were preliminarily screened.

Results During PCD progression in the upper buds from 20 days after flowering, DEGs were significantly enriched in the plant hormone signal transduction pathway. ET release amount in the upper buds increased rapidly, while no significant change was observed in the lower latent buds. Most genes related to ET biosynthesis and positive regulators of ET signal transduction were up-regulated in the upper buds, whereas most of these genes showed no significant differential expression in the lower latent buds. Eight key candidate genes involved in ET-mediated bud PCD were screened, including five ET biosynthesis genes EVM0017576 (ACO1), EVM0001055 (ACS1), EVM0023867 (ACO11), EVM0006661 (ACO4), and EVM0029965 (ACS7); and three ET signal transduction genes EVM0021672 (EIN2), EVM0023854 (EIL3), and EVM0022486 (ERF1).

Conclusion During PCD in the upper buds, ET release amount increases sharply, which is consistent with the up-regulation of most ET biosynthesis-related genes. Moreover, positive regulators of ET signal transduction are up-regulated, promoting ET signal propagation. Therefore, it is preliminarily concluded that ET positively regulates PCD in the upper buds of apical-bud-senescence chestnut shoots. The eight candidate genes related to ET synthesis and signal transduction provide a basis for future cloning and functional validation.