Nitrogen fertilization and δ18 O of CO2 have no effect on 18 O-enrichment of leaf water and cellulose in Cleistogenes squarrosa (C4 ) - is VPD the sole control?

The oxygen isotope composition of cellulose (δ18 OCel ) archives hydrological and physiological information. Here, we assess previously unexplored direct and interactive effects of the δ18 O of CO2 (δ18 OCO2 ), nitrogen (N) fertilizer supply and vapour pressure deficit (VPD) on δ18 OCel , 18 O-enrichment of leaf water (Δ18 OLW ) and cellulose (Δ18 OCel ) relative to source water, and pex px , the proportion of oxygen in cellulose that exchanged with unenriched water at the site of cellulose synthesis, in a C4 grass (Cleistogenes squarrosa). δ18 OCO2 and N supply, and their interactions with VPD, had no effect on δ18 OCel , Δ18 OLW , Δ18 OCel and pex px . Δ18 OCel and Δ18 OLW increased with VPD, while pex px decreased. That VPD-effect on pex px was supported by sensitivity tests to variation of Δ18 OLW and the equilibrium fractionation factor between carbonyl oxygen and water. N supply altered growth and morphological features, but not 18 O relations; conversely, VPD had no effect on growth or morphology, but controlled 18 O relations. The work implies that reconstructions of VPD from Δ18 OCel would overestimate amplitudes of VPD variation, at least in this species, if the VPD-effect on pex px is ignored. Progress in understanding the relationship between Δ18 OLW and Δ18 OCel will require separate investigations of pex and px and of their responses to environmental conditions.