N uptake, assimilation and isotopic fractioning control δ (15)N dynamics in plant DNA: A heavy labelling experiment on Brassica napus L.

In last decades, a large body of evidence clarified nitrogen isotope composition (δ(15)N) patterns in plant leaves, roots and metabolites, showing isotopic fractionation along N uptake and assimilation pathways, in relation to N source and use efficiency, also suggesting (15)N depletion in plant DNA. Here we present a manipulative experiment on Brassica napus var. oleracea, where we monitored δ (15)N of purified, lyophilized DNA and source leaf and root materials, over a 60-days growth period starting at d 60 after germination, in plants initially supplied with a heavy labelled (δ (15)N(Air-N2) = 2100 mUr) ammonium nitrate solution covering nutrient requirements for the whole observation period (470 mg N per plant) and controlling for the labelled N species (ṄH(4), ṄO(3) and both). Dynamics of Isotopic Ratio Mass Spectrometry (IRMS) data for the three treatments showed that: (1) leaf and root δ (15)N dynamics strictly depend on the labelled chemical species, with ṄH(4), ṄO(3) and ṄH(4)ṄO(3) plants initially showing higher, lower and intermediate values, respectively, then converging due to the progressive NH(4)(+) depletion from the nutrient solution; (2) in ṄH(4)ṄO(3), where δ(15)N was not affected by the labelled chemical species, we did not observe isotopic fractionation associated to inorganic N uptake; (3) δ(15)N values in roots compared to leaves did not fully support patterns predicted by differences in assimilation rates of NH(4)(+) and NO(3)(-); (4) DNA is depleted in (15)N compared to the total N pools of roots and leaves, likely due to enzymatic discrimination during purine biosynthesis. In conclusion, while our experimental setup did not allow to assess the fractionation coefficient (ε) associated to DNA bases biosynthesis, this is the first study specifically reporting on dynamics of specific plant molecular pools such as nucleic acids over a long observation period with a heavy labelling technique.