Integrated physiological, transcriptomic and metabolomic analyses reveal ROS regulatory mechanisms in two castor bean varieties under alkaline stress.

Saline-alkaline stress has caused severe ecological and environmental problems. Castor bean is a potential alkali-tolerant plant, however, its reactive oxygen species (ROS) regulatory mechanisms under alkaline stress remain unclear. This study investigated the physiological, transcriptomic, and metabolomic characteristics of two varieties (ZB8, alkaline-sensitive; JX22, alkaline-resistant) under alkaline stress. Results showed that under alkaline stress, JX22's root length was 1.66-fold greater than ZB8's, while its superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities were 1.25-, 1.41-, and 1.29-fold higher than ZB8's, respectively. The levels of superoxide anion (O) and malondialdehyde (MDA) in JX22 were 0.2- and 0.68-fold of those in ZB8, respectively. Integrated transcriptomic and metabolomic analyses revealed that regarding ROS generation, alkaline stress promoted the upregulation of ACX1 and RBOHD genes in JX22, enabling more efficient ROS signal transduction and subsequent stress response regulation. In terms of ROS signal transduction, alkaline stress induced significant upregulation of protein kinase-encoding genes including CPK4, CPK9, and CPK10 in JX22, which cooperated with RBOHD to regulate ROS production. Concerning ROS scavenging, significant upregulation of SODA, CAT2, and PRXⅡB genes ensured a more efficient enzymatic ROS scavenging system in JX22 under alkaline stress. In contrast, ZB8 could only rely on less efficient non-enzymatic systems, such as carotenoid antioxidants, to mitigate oxidative damage, where genes like CCD7, CYP897B and metabolites including lutein and zeaxanthin played crucial roles. These findings elucidate the ROS response mechanisms of castor bean under alkaline stress, paving new ways for breeding alkaline-resistant varieties.