Role of ethylene and proteolytic N-degron pathway in the regulation of Arabidopsis seed dormancy

Primary dormant seeds of Arabidopsis thaliana did not germinate in darkness at temperature higher than 10–15°C. Ethylene improved the germination of dormant wild-type (Col-0) seeds at 25°C in darkness but seeds of the mutant affected in the proteolytic N-degron pathway, proteolysis6 (prt6), were insensitive to ethylene suggesting that PRT6 was involved in dormancy release by ethylene. The substrates of the N-degron pathway, the Ethylene Response Factors from group VII (HRE1, HRE2, RAP2.2, RAP2.3, and RAP2.12), were identified to be involved in this insensitivity with an increased germination in prt6 rap2.2 rap2.3 rap2.12 rather than in prt6 hre1 hre2, which also indicated that the three RAPs acted downstream of PRT6, while the two HREs acted upstream of PRT6. Ethylene reduced the expression of the three RAPs in Col-0 seeds but they were maintained or induced by ethylene in prt6 seeds. The promoting effect of ethylene was associated with a down-regulation of dormancy-related genes in gibberellins (GAs) and abscisic acid (ABA) signaling, such as RGA, RGL2, and ABI5, and with a strong decrease in ABA/GA₄ ratio in the presence of ethylene. In contrast, we show that the insensitivity of prt6 seeds to ethylene was mainly related to GA signaling disturbance.