Heat tolerance in Arabidopsis thaliana seedlings requires functional DMS3, a component of de novo methylation

Abstract

The protein Defective in RNA-directed DNA Methylation 3 (DMS3) is part of RNA-directed DNA methylation, an epigenetic mechanism involved in the regulation of plant development and stress response. However, the specific role of the DMS3 protein in thermotolerance remains unclear. To determine how altered DMS3 expression and functionality affects thermotolerance, DMS3-overexpressor (oeDMS3), DMS3-mutant (dms3-1) and wild-type Arabidopsis thaliana seedlings were heat-treated and analyzed, focusing on morphological, physiological, biochemical and molecular changes. The dms3-1 line showed the highest thermosensitivity after short-term exposure to 45 °C for 45 min. However, both dms3-1 and oeDMS3 showed a greater reduction in morphological traits compared to wild type after exposure to 40 °C for 6 h. Hormonal profiling showed that the dms3-1 and oeDMS3 lines had similar hormonal profiles characterized by lower jasmonate levels compared to wild type, both under stress and control conditions. The heat-stressed dms3-1 line contained increased cytokinin levels predominantly in the form of ribosides, and also accumulated inactive auxin metabolites. Exposure to 37 °C for 24 h destabilized and altered the localization of the DMS3 protein in the root tissue. After exposure to 37 °C for 6 h, the dms3-1 line showed a delayed recovery of reduced photosynthetic efficiency, accompanied by a partial activation of the antioxidant system and increased proline content. Under control conditions, dms3-1 plants exhibited reduced growth and lower expression of RuBisCO, HSP90 and HSP70 proteins. Overall, our results suggest a crucial role of DMS3 in thermotolerance, hormone balance, antioxidant defense and photosynthetic efficiency, indicating the importance of a functional and balanced DMS3 protein for thermotolerance and for plant growth and development under control conditions.