Cytokinin Response Factor 6 Represses Cytokinin-Associated Genes during Oxidative Stress

Abstract

Cytokinin is a phytohormone that is well known for its roles in numerous plant growth and developmental processes, yet it has also been linked to abiotic stress response in a less defined manner. Arabidopsis (Arabidopsis thaliana) Cytokinin Response Factor 6 (CRF6) is a cytokinin-responsive AP2/ERF-family transcription factor that, through the cytokinin signaling pathway, plays a key role in the inhibition of dark-induced senescence. CRF6 expression is also induced by oxidative stress, and here we show a novel function for CRF6 in relation to oxidative stress and identify downstream transcriptional targets of CRF6 that are repressed in response to oxidative stress. Analysis of transcriptomic changes in wild-type and crf6 mutant plants treated with H 2 O 2 identified CRF6-dependent differentially expressed transcripts, many of which were repressed rather than induced. Moreover, many repressed genes also show decreased expression in 35S:CRF6 overexpressing plants. Together, these findings suggest that CRF6 functions largely as a transcriptional repressor. Interestingly, among the H 2 O 2 repressed CRF6-dependent transcripts was a set of five genes associated with cytokinin processes: (signaling) ARR6, ARR9, ARR11, (biosynthesis) LOG7, and (transport) ABCG14. We have examined mutants of these cytokinin-associated target genes to reveal novel connections to oxidative stress. Further examination of CRF6-DNA interactions indicated that CRF6 may regulate its targets both directly and indirectly. Together, this shows that CRF6 functions during oxidative stress as a negative regulator to control this cytokinin-associated module of CRF6-dependent genes and establishes a novel connection between cytokinin and oxidative stress response.The frequent environmental changes to which a plant is subject can lead to physiological alterations and disruption of normal metabolism. In particular, the energetic reactions that take place in chloroplasts, peroxisomes, and mitochondria are susceptible to dysfunction, which results in production of excessive levels of reactive oxygen species (ROS). In fact, many common abiotic stress conditions encountered in agriculture, including temperature extremes, drought, soil salinity, and air pollution, are known to include an oxidative stress component (Gill and Tuteja, 2010). Cellular levels of ROS are carefully maintained at relatively low levels through a wide range of scavenging and detoxification mechanisms. However, if the balance between ROS production and removal is shifted too far toward production (e.g. under stress conditions), cellular damage can occur as a result of oxidation of macromolecules such as lipids, proteins, and nucleic acids (Mittler, 2002;Gill and Tuteja, 2010). Accumulation of ROS beyond some threshold triggers cell death as a response. Therefore, ROS are thought to serve as indicators of oxidative stress within a cell but may also

Salt and oxidative stresses uniquely regulate tomato cytokinin levels and transcriptomic response

“…The classic cytokinin senescence bioassay, normally used to examine developmental senescence, was modified to examine abiotic stress responses of tomato leaf disks (Mok & Mok, ; Zwack, Robinson, Risley, & Rashotte, ). Previous work, again verified here, shows that cytokinin treatment of leaf disks reduces leaf senescence as measured using a chlorophyll fluorimeter to determine the efficiency of photosystem II or F v / F m (Zwack et al., ; Figures and ). Oxidative stress (20 mM H 2 O 2 ) reduces F v / F m by 10%–30% over simple buffer treatment.…”

“…One simple, nondestructive examination of photosystem II efficiency is the measurement of chlorophyll fluorescence or F v / F m using a fluorimeter. The classic cytokinin senescence bioassay that was modified to study abiotic stress responses was used in this study (Letham, ; Zwack et al., , ). Tomato leaf disks were floated in buffer in the presence or the absence of cytokinin, either before (pre) or after (post) salt (NaCl), or oxidative (H 2 O 2 ) stress treatment.…”

Redox regulation at the site of primary growth: auxin, cytokinin and ROS crosstalk

“…Constitutive overexpression of CRF6 leads to improved photosynthesis and increased root growth when exposed to oxidative stress‐inducing conditions. Protective mechanisms depend on the attenuation of cytokinin signalling via CRF6‐dependent repression of AHP1 , ARR1 , ARR6 , ARR9 , LOG7 and cytokinin transport ABCG14 genes (Zwack et al ). In response to low temperature, roots adapt by reducing RAM's division potential.…”

Photoperiod stress alters the cellular redox status and is associated with an increased peroxidase and decreased catalase activity

“…One direct link to regulate ROS formation by CK is through ARR2, 379 a CK-regulated transcription factor known to bind directly to the promoters of the PRX33 and 380 PRX34 genes (Arnaud et al, 2017). Another CK-regulated transcription factor, CYTOKININ 381 RESPONSE FACTOR6, is responsive to oxidative stress (Zwack et al, 2013) and regulates 382 crosstalk between H 2 O 2 and the CK system (Zwack et al, 2016). Notably, CK has anti-oxidative 383 stress activity even in bacterial and human (Othman et al, 2016) cells which 384 suggests that the hormone might have acquired this function very early during evolution and 385 retained it ever since in different organisms (Kabbara et al, 2018).…”