The importance of the urea cycle and its relationships to polyamine metabolism during ammonium stress in Medicago truncatula

The ornithine–urea cycle (urea cycle) makes a signifcant contribution to the metabolic responses of lower photosynthetic eukaryotes to episodes of high nitrogen availability. In this study, we compared the role of the plant urea cycle and its relationships to polyamine metabolism in ammonium-fed and nitrate-fed Medicago truncatula plants. High ammonium resulted in the accumulation of ammonium and pathway intermediates, particularly glutamine, arginine, ornithine, and putrescine. Arginine decarboxylase activity was decreased in roots, suggesting that the ornithine decarboxylase-dependent production of putrescine was important in situations of ammonium stress. The activity of copper amine oxidase, which releases ammonium from putrescine, was signifcantly decreased in both shoots and roots. In addition, physiological concentrations of ammonium inhibited copper amine oxidase activity in in vitro assays, supporting the conclusion that high ammonium accumulation favors putrescine synthesis. Moreover, early supplementation of plants with putrescine avoided ammonium toxicity. The levels of transcripts encoding urea-cyclerelated proteins were increased and transcripts involved in polyamine catabolism were decreased under high ammonium concentrations. We conclude that the urea cycle and associated polyamine metabolism function as important protective mechanisms limiting ammonium toxicity in M. truncatula. These fndings demonstrate the relevance of the urea cycle to polyamine metabolism in higher plants.This work was supported by the grants from the Spanish Government AGL2014-52396-P (MICINN) and AGL2017-86293-P (MINECO/ FEDER) to JFM, and the Basque Government, Spain, IT-1018-16 (UPV/EHU-GV) to RE. MU is a recipient of a pre-doctoral fellowship from the Government of Navarre, Spain. JB and PLG have received pre-doctoral fellowships from the Public University of Navarre, Spain. PT has received funding from the Italian Ministry of Education, University and Research (Grant to Department of Science, University ‘Roma Tre’-‘Dipartimenti di Eccellenza’, ARTICOLO 1, COMMI 314– 337. LEGGE 423 232/2016; PRIN 2017—CUP F84I19000730005). Partial support was obtained from the Spanish State Research Agency AGL2017-83358-R (AEI/FEDER) and the Government of Aragon, Spain, Group A09-20R to YG. Open Access funding was provided by the Public University of Navarra