Ethylene negatively regulates cold tolerance through HbEIN3-HbICE2 regulatory module in Hevea brasiliensis

Cold stress can result in reduced growth rates, decreased latex production, and restricted areas for the Para rubber tree (Hevea brasiliensis). However, the molecular mechanisms governing the response of Hevea brasiliensis to cold stress remain elusive. Here, we found that ethylene plays a negative role in Hevea brasiliensis responses to cold stress. Treatment with the ethylene synthesis precursor 1-aminocyclopropane-1-carboxylic acid (ACC) decreased the cold tolerance of Hevea brasiliensis, while exogenous treatment with Ag⁺ (an ethylene signal inhibitor) had the opposite effect. Additionally, overexpressing HbEIN3 decreased cold stress tolerance in Arabidopsis and Taraxacum koksaghyz plants. Quantitative real-time PCR analysis indicated that HbEIN3-1 and HbEIN3-2 repress the expression of the cold-responsive genes HbCBF1-3 in Hevea brasiliensis. Moreover, HbEIN3-1 and HbEIN3-2 directly bind to the HbCBF1 promoter to suppress its transcription. Further investigation revealed that HbEIN3s interact with and dampen the transcriptional activity of HbICE2, a crucial transcription factor that positively regulates the cold signaling pathway, thereby attenuating the expression of HbICE2-targeted genes. Collectively, these findings indicate that HbEIN3s play a crucial role in ethylene-regulated cold tolerance through the repression of HbCBF1 expression and HbICE2 transcriptional activity.