CRK5 preserves antioxidant homeostasis and prevents cell death during dark-induced senescence through inhibiting the salicylic acid signaling pathway

Background Dark-induced senescence (DIS) is a widely used model for dissecting the regulatory mechanisms that manage leaf aging, redox imbalance, and cell death (CD) in plants. Salicylic acid (SA) is a central hormonal regulator of these processes. However, the mechanism involving upstream components, which integrate SA-dependent pathways with antioxidant homeostasis during DIS, remains unresolved. CYSTEINE-RICH RECEPTOR-LIKE KINASE 5 (CRK5) is a membrane-localized protein that plays a role in developmental and stress-responsive pathways. Its promoter contains multiple W-box cis -elements, indicating regulation by WRKY factors in SA-mediated pathways. This study investigates how CRK5 modulates SA-dependent CD and antioxidant dynamics during DIS. Results In this study, SA-accumulating mutant crk5 exhibited accelerated senescence, elevated electrolyte leakage, enhanced micro-lesion formation, and markedly increased reactive oxygen species (ROS) accumulation under both control and dark conditions. These phenotypes were accompanied by a substantial reduction in carotenoid and xanthophyll pools, enhanced accumulation of phenolic compounds, and increased free radical scavenging capacity, including ascorbate peroxidase, catalase, and superoxide dismutase activities. Importantly, crk5 phenotype was fully reverted in crk5sid2 and crk5 NahG double mutants, confirming that crk5 DIS phenotype is induced by activation of the SA-signaling pathway. Transcriptome profiling revealed extensive deregulation of senescence-, CD-, and redox-associated genes in crk5 during darkness, including strong induction of SAGs, metacaspases, autophagy, and antioxidant-related transcripts. The line with constitutively enhanced SA level ( cpr1 ), used as a control, showed similar phenotypes to crk5 , although transcriptional reprogramming was largely absent in cpr1 after darkness, highlighting CRK5 as a key upstream negative regulator of SA-mediated CD and positive regulator of antioxidant homeostasis. Conclusion Our work presents CRK5 as a central regulatory hub that inhibits the SA-signaling, ROS burst, and CD activation during DIS. Loss of CRK5 function is associated with the activation of SA-signaling, altered antioxidant systems, increased ROS burden, and ROS-driven CD acceleration, resulting in accelerated senescence. Conversely, suppression of SA-biosynthesis or -catabolism in a crk5 background restores the wild-type phenotype. These findings position this receptor kinase as a key mediator that coordinates hormonal, metabolic, and oxidative pathways to maintain leaf viability, providing mechanistic insight into the control of stress-induced senescence and CD in Arabidopsis.