Developmental stage specificity and the role of mitochondrial metabolism in the response of Arabidopsis leaves to prolonged mild osmotic stress

Aleksandra Skirycz , Stefanie De Bodt , Toshihiro Obata , Inge De Clercq , Hannes Claeys , Riet De Rycke , Megan Andriankaja , Olivier Van Aken , Frank Van Breusegem , Alisdair R. Fernie , and Dirk Inze ^*

Department of Plant Systems Biology, Flanders Institute for Biotechnology, Ghent University, B–9052 Gent, Belgium; Department of Plant Biotechnology and Genetics, Ghent University, B–9052 Gent, Belgium; Max-Planck Institute for Molecular Plant Physiology, D-14476 Potsdam-Golm, Germany

^* Corresponding author; email: dirk.inze{at}psb.vib-ugent.be.

When subjected to stress, plants reprogram their growth bylargely unknown mechanisms. To provide insights into this process,growth of Arabidopsis (Arabidopsis thaliana) leaves that developunder mild osmotic stress was studied. Early during leaf development,cell number and size were reduced by stress, but growth wasremarkably adaptable, as division and expansion rates were identicalto control levels within a few days of leaf initiation. To investigatethe molecular basis of the observed adaptability leaves withonly proliferating, exclusively expanding, and mature cellswere analyzed by transcriptomics and targeted metabolomics.The stress response measured in growing and mature leaves waslargely distinct; several hundred transcripts and multiple metabolitesresponded exclusively in the proliferating and/or expandingleaves. Only a few genes were differentially expressed acrossthe three stages. Data analysis showed that proliferation andexpansion were regulated by common regulatory circuits, involvingethylene and gibberellins, but not abscisic acid. The role ofethylene was supported by the analysis of ethylene-insensitivemutants. Exclusively in proliferating cells, stress inducedgenes of the so-called "mitochondrial dysfunction regulon",comprising alternative oxidase. Up-regulation for eight of thesegenes was confirmed with promoter: ${beta}$ -glucuronidase reporter lines.Furthermore, mitochondria of stress-treated dividing cells weremorphologically distinct from control ones and growth of plantsoverexpressing the alternative oxidase gene was more tolerantto osmotic and drought stresses. Taken together, our data underlinethe value of analyzing stress responses in function of developmentand demonstrate the importance of mitochondrial respirationfor sustaining cell proliferation under osmotic stress conditions.