Over-expression analysis of TaDWF4˗4B in mutant wheat lines identifies a candidate regulator of heat stress tolerance

Wheat production faces great threat due to rising environmental temperatures resulting from climate change. Here we analyzed a bulk of wheat exotic, elite, synthetic and local wheat lines to evaluate the impact of heat stress on the physiological and biochemical parameters of different genotypes. Mutant susceptible genotypes were generated by treating with Ethyl Methane Sulfonate (EMS) and its performance was evaluated under normal and heat stress conditions. Based on the heat stress index and yield measurement, the lines were grouped into four classes, i.e. tolerant (T), Susceptible (S), moderately tolerant (MT), and moderately susceptible (MS). The results showed significant impact of heat stress on all parameters of the tested gene pool; however, the affect was less intense on the tolerant lines compared to the other classes. To validate heat stress tolerance, we conducted RNA sequencing analysis and identified eight genes associated with heat tolerance. Among them, TaDWF4˗4B showed highest expression under heat stress condition and selected for further functional analysis. Overexpression of TaDWF4˗4B in the wheat line ESWYT-4 enhanced heat tolerance. Treatment with 2 μM brassinosteroids (BR) decreased seed germination in the transgenic lines, suggesting that TaDWF4˗4B enhances BR response. Endogenous BR contents increased in overexpression lines, along with increasing the expression of several BR biosynthetic pathway genes in overexpression line under heat stress condition. Moreover, the overexpression of TaDWF4˗4B improved reactive oxygen species (ROS) scavenging by increasing the activities of TaCAT3, TaSOD1, and TaGPx1-D under heat stress condition. These findings indicate that TaDWF4˗4B plays an important role in regulating BR biosynthesis, increasing BR response, and ROS scavenging under heat stress condition. These results present mechanistic insights into the role of TaDWF4˗4B in plant responses under heat stress condition.