Abstract

To clarify the molecular mechanisms by which transcription factors (TFs) regulate reactive oxygen species (ROS) metabolism during low-temperature alleviation of internal browning (IB) in pineapple, changes in ROS accumulation and TF-mediated regulation of ROS scavenging genes, with a focus on basic helix–loop–helix (bHLH) TF, were investigated in ‘Comte de Paris’ pineapple fruit stored at 10 °C (low temperature) and 25 °C. Fruit stored at 10 °C showed lower levels of malondialdehyde (MDA), hydrogen peroxide (H₂O₂), superoxide anion (O₂ •– ), and IB than fruit stored at 25 °C. Weighted gene co-expression network analysis (WGCNA) combined with K-means clustering of transcriptome sequencing data identified low temperature-inducible AcbHLH82 as a TF negatively correlated with ROS accumulation during IB development. Electrophoretic mobility shift assays and dual-luciferase reporter analyses demonstrated that AcbHLH82 directly activated transcription of ROS scavenging genes such as AcAPX1 , AcPOD1 , and AcPOD30 by specifically recognizing E-box cis-elements (CANNTG), thereby enhancing ROS scavenging capacity. Transient overexpression and silencing of AcbHLH82 in pineapple pulp showed that elevated AcbHLH82 expression effectively reduced IB, accompanied by increased expression of ROS scavenging genes and decreased H₂O₂ and O₂ •– levels. These results provide insight into the molecular basis by which low temperature storage alleviates IB, partly through enhanced antioxidant scavenging capacity mediated by AcbHLH82-dependent activation of ROS scavenging genes.