trans-Zeatin N-glucosides can delay salt accelerated leaf senescence in Arabidopsis thaliana

While cytokinin (CK) can delay natural leaf senescence, its effects on abiotic stress accelerated leaf senescence are less studied. Here we show N-conjugated trans-zeatin CK forms (tZ7G and tZ9G, or tZNGs) have the ability to delay salt stress senescence. Using a modified dark-induced senescence bioassay with Arabidopsis leaves, exogenous salt treatment accelerated leaf senescence as measured by lower photosystem II efficiency (F_v/F_m) and chlorophyll content. tZNGs were able to delay these parameters at concentrations as low as 10 nM similar to tZ, indicating that tZ7G and tZ9G can function in delaying salt accelerated senescence (SAS). To better understand physiological effects regulating tZNG delay of senescence, transcriptomics, proteomics, as well as CK measurements were examined. Salt treatment has strong transcriptome and proteome effects in accelerating senescence and reducing overall CK levels. Exogenous CK treatments could be quickly detected from changes seen in endogenous CK measurements, where each CK has a distinct profile contributing to transcript/protein alterations. Interestingly, transcriptomics show tZNGs are primarily responsive at later stages of salt senescence, in contrast to an immediate and continual response of tZ treatment. Known CK-regulated genes are induced by tZNGs and tZ, as corroborated by ARR:GUS reporter lines. Differences between tZNGs and tZ DEGs were revealed by WGCNA that included salt and CK specifically gene modules. In contrast, proteomic analysis revealed unique, but similar numbers of tZNG compared to tZ DAPs across senescence. GO term analysis of tZNG DEGs and DAPs showed enrichment of senescence, chloroplast, and CK signaling. Together this indicates tZNGs function as active CK forms in delaying salt accelerated leaf senescence.