Plant U-Box 4 regulates chloroplast stress signaling and programmed cell death via Salicylic acid modulation

Abstract

BackgroundIn response to environmental stresses, chloroplasts generate reactive oxygen species, including singlet oxygen (1O2), an excited state of oxygen that regulates chloroplast-to-nucleus (retrograde) signaling, chloroplast turnover, and programmed cell death (PCD). Yet, the central signaling mechanisms and downstream responses remain poorly understood. The Arabidopsis thaliana plastid ferrochelatase two (fc2) mutant conditionally accumulates 1O2 and Plant U-Box 4 (PUB4), a cytoplasmic E3 ubiquitin ligase, is involved in propagating 1O2 signals for chloroplast turnover and cellular degradation. Thus, the fc2 and fc2 pub4 mutants are useful genetic tools to elucidate these signaling pathways. Previous studies have focused on the role of 1O2 in promoting cellular degradation in fc2 mutants, but its impact on retrograde signaling from mature chloroplasts (the major site of 1O2 production) is poorly understood. ResultsTo gain mechanistic insights into 1O2 signaling pathways, we compared transcriptomes of adult wt, fc2, and fc2 pub4 plants. The accumulation of 1O2 in fc2 plants broadly repressed genes involved in chloroplast function and photosynthesis, while inducing genes and transcription factors involved in abiotic and biotic stress, the biosynthesis of jasmonic acid (JA) and salicylic acid (SA), microautophagy, and senescence. Elevated JA and SA levels were observed in 1O2-stressed fc2 plants. pub4 reversed most of this 1O2-induced gene expression and reduced the JA content in fc2 plants. The pub4 mutation also blocked JA-induced senescence pathways in the dark. However, fc2 pub4 plants maintained constitutively elevated levels of SA even in the absence of bulk 1O2 accumulation. ConclusionsTogether, this work demonstrates that in fc2 plants, 1O2 leads to a robust retrograde signal that may protect cells by downregulating photosynthesis and ROS production while simultaneously mounting a stress response involving SA and JA. The induction of microautophagy and senescence pathways indicate that 1O2-induced cellular degradation is a genetic response to this stress, and the bulk of this transcriptional response is modulated by the PUB4 protein. However, the effect of pub4 on hormone synthesis and signaling is complex and indicates that an intricate interplay of SA and JA are involved in promoting stress responses and programmed cell death during photo-oxidative damage.