Abstract
Phytohormones mediate most diverse processes in plants, ranging from organ development to immune responses. Receptor protein complexes perceive changes in intracellular phytohormone levels and trigger a signaling cascade to effectuate downstream responses. The in planta analysis of elements involved in phytohormone signaling can be achieved through transient expression in mesophyll protoplasts, which are a fast and versatile alternative to generating plant lines that stably express a transgene. While promoter-reporter constructs have been used successfully to identify internal or external factors that change phytohormone signaling, the range of available marker constructs does not meet the potential of the protoplast technique for large scale approaches. The aim of our study was to provide novel markers for phytohormone signaling in the Arabidopsis mesophyll protoplast system. We validated 18 promoter::luciferase constructs towards their phytohormone responsiveness and specificity and suggest an experimental setup for high-throughput analyses. We recommend novel markers for the analysis of auxin, abscisic acid, cytokinin, salicylic acid and jasmonic acid responses that will facilitate future screens for biological elements and environmental stimuli affecting phytohormone signaling.
Highlights
Elucidating the in planta function of genes or regulatory factors is key in the process to understand how individual signaling components are interconnected and contribute to signaling pathways and networks
We focused on responses of the phytohormones abscisic acid, auxin, cytokinin, salicylic acid and jasmonic acid due to their principal significance in growth, abiotic stress and disease resistance
Some of these genes are characterized towards their function and position within the signaling network but many have been selected for their consistent transcriptional response to the presence of a phytohormone
Summary
Elucidating the in planta function of genes or regulatory factors is key in the process to understand how individual signaling components are interconnected and contribute to signaling pathways and networks. This task often involves generating transgenic plants which is timeconsuming, laborious and cannot be applied in large-scale screening approaches. The use of transient gene expression in protoplasts is an alternative technique that offers many advantages such as a high-throughput, cost effectiveness and great flexibility towards the components (e.g. proteins) to be tested [1]. Phytohormone markers for protoplast-based screening and analysis, decision to publish, or preparation of the manuscript
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