Ethylene Signaling Negatively Regulates Freezing Tolerance by Repressing Expression of <i>CBF</i> and Type-A <i>ARR</i> Genes in <i>Arabidopsis</i>

TLDR

Ethylene modulates plant growth and stress responses, but its specific role in freezing tolerance is not well defined. This study investigates whether ethylene negatively regulates freezing tolerance in Arabidopsis thaliana. EIN3 binds promoters of CBFs and type‑A ARR genes, repressing their expression and thereby dampening cold signaling. Ethylene overproduction or precursor application reduces freezing tolerance, while inhibition or ethylene‑insensitive mutants enhance it; overexpressing type‑A ARR genes improves tolerance, confirming that ethylene suppresses cold responses through EIN3‑mediated repression of CBFs and type‑A ARR genes and that these ARR genes integrate ethylene and cytokinin signaling.

Abstract

The phytohormone ethylene regulates multiple aspects of plant growth and development and responses to environmental stress. However, the exact role of ethylene in freezing stress remains unclear. Here, we report that ethylene negatively regulates plant responses to freezing stress in Arabidopsis thaliana. Freezing tolerance was decreased in ethylene overproducer1 and by the application of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid but increased by the addition of the ethylene biosynthesis inhibitor aminoethoxyvinyl glycine or the perception antagonist Ag+. Furthermore, ethylene-insensitive mutants, including etr1-1, ein4-1, ein2-5, ein3-1, and ein3 eil1, displayed enhanced freezing tolerance. By contrast, the constitutive ethylene response mutant ctr1-1 and EIN3-overexpressing plants exhibited reduced freezing tolerance. Genetic and biochemical analyses revealed that EIN3 negatively regulates the expression of CBFs and type-A Arabidopsis response regulator5 (ARR5), ARR7, and ARR15 by binding to specific elements in their promoters. Overexpression of these ARR genes enhanced the freezing tolerance of plants. Thus, our study demonstrates that ethylene negatively regulates cold signaling at least partially through the direct transcriptional control of cold-regulated CBFs and type-A ARR genes by EIN3. Our study also provides evidence that type-A ARRs function as key nodes to integrate ethylene and cytokinin signaling in regulation of plant responses to environmental stress.