Putrescine-responsive post-transcriptional feedback regulation of Arabidopsis ADC2 mediated by 5'-UTR elements

Abstract

Putrescine (Put) is a bioactive polyamine that contributes to plant growth, development, and stress responses, whereas excessive Put accumulation impairs growth. However, how plants maintain Put levels within an appropriate range remains poorly understood. In Arabidopsis thaliana, expression of the arginine decarboxylase 2 (ADC2) gene is a major determinant of Put abundance. Here, we show that the 5' untranslated region (5'-UTR) of ADC2 mediates Put-responsive feedback regulation and contributes to Put homeostasis in Arabidopsis. In transient expression assays using an emerald luciferase (Eluc) gene placed downstream of the ADC2 5'-UTR (ADC2 5'-UTR:Eluc), Eluc activity was reduced by exogenously applied Put but increased when endogenous Put biosynthesis was impaired by adc2 mutations or treatment with an arginine decarboxylase inhibitor. In transgenic plants carrying ADC2 5'-UTR:Eluc, Put treatment modestly decreased Eluc mRNA abundance but substantially reduced Eluc activity and translation efficiency, suggesting that this Put-responsive repression occurs mainly at the translational level. Among the cis-regulatory elements in the ADC2 5'-UTR, mutations affecting either the conserved AUU/CUG-initiated upstream ORF or the predicted RNA secondary structure in the ADC-box, a conserved GC-rich motif, abolished Put-responsive repression, indicating that these elements are required for this regulation. Two genome-edited lines carrying deletions spanning these elements exhibited delayed germination and impaired growth, and the more severely affected line showed increased Put accumulation. Together, these findings reveal a feedback mechanism that regulates ADC2 expression through its 5'-UTR and support a role for this mechanism in Put homeostasis in Arabidopsis.