Ethylene regulates aerenchyma formation in cotton under hypoxia stress by inducing the accumulation of reactive oxygen species

Aerenchyma formation is one of the most effective mechanisms adapted by plants to cope with waterlogging (WL). It which reduces the negative effects of WL by facilitating gas diffusion within roots. To understand the response of cotton roots to WL, hypoxia-tolerant genotype Cx094 and hypoxia-sensitive genotype Wang18 were used in this study. The seedlings of two varieties were treated by three methods: flooding in pots as Treatment 1 (flooding), low oxygen with agar solution as Treatment 2 (flooding + agar) and hypoxia with agar solution and nitrogen gas as Treatment 3 (flooding + agar + N₂). The results showed that in Cx094, the highest proportion of aerenchyma was found in Treatment 2, with no corresponding aerenchyma formation in Wang18. Further studies were carried out in Treatment 2 to check the aerenchyma formation in six cotton accessions and the results showed that aerenchyma only formed in hypoxia-tolerance genotypes, confirming the important role of aerenchyma formation in WL-tolerant genotypes. The unique characteristic of cotton is that four symmetrical aerenchyma were formed under WL stress, and they appeared one by one. We also showed that hypoxia signal induced expression of ACS and ACO genes, resulting more ethylene synthesis in hypoxia-tolerant genotypes. Moreover, ethylene induced reactive oxygen species (ROS) accumulated by up-regulation of RBOHs expression and down-regulation of MT-3a expression. Finally, ROS caused up-regulation of CEL and XET expressions to induce programmed cell death, and result in lysigenous aerenchyma. Exogenous ethylene inhibitor decreased the formation of aerenchyma in Cx094, while ethephon, an ethylene precursor, promote the formation of aerenchyma in Wang18, signifying the important role of ethylene in the formation of aerenchyma in cotton.