MiR164a-targeted NAM3 inhibits thermotolerance in tomato by regulating HSFA4b-mediated redox homeostasis

Abstract Extreme weather events, including high temperatures, frequently occur and adversely affect crop growth, posing substantial challenges to global agriculture. MicroRNAs (miRNAs) play integral roles in regulating plant growth and responses to various stresses. In this study, we reveal that microRNA164a (miR164a) in tomato (Solanum lycopersicum) is a pivotal element that exhibits a rapid positive response to heat stress (HS) among multiple miRNAs, while its target NO APICAL MERISTEM 3 (NAM3) shows an opposite complementary response. MiR164a/b-5p-deficient mutant and NAM3-overexpressing plants resulted in increased sensitivity to HS, whereas mutants with reduced NAM3 levels exhibited enhanced thermotolerance. Importantly, HS-induced reactive oxygen species (ROS) accumulation and antioxidant enzyme activities were positively regulated by miR164a and negatively by NAM3, respectively. Furthermore, we demonstrated that NAM3 transcriptionally activated the expression of HSFA4b, and silencing HSFA4b improved tomato thermotolerance. HSFA4b repressed the expression of the antioxidant gene APX1 and the heat shock protein (HSP) gene HSP90, disrupting redox homeostasis and exacerbating oxidative stress. Our findings unveil a pivotal regulatory pathway governed by the miR164a-NAM3 module that confers thermotolerance in tomato via its influence on ROS-related and HSP pathways. These findings provide valuable insights into the molecular mechanisms that underpin tomato thermotolerance, which are crucial for advancing sustainable agricultural practices, particularly in the face of the challenges presented by global climate change.