Leaf cell wall properties and stomatal density influence oxygen isotope enrichment of leaf water.

Measurements of oxygen isotope enrichment of leaf water above source water (Δ18 OLW ) can improve our understanding of the interaction between leaf anatomy and physiology on leaf water transport. Models have been developed to predict Δ18 OLW such as the string-of-lakes model, which describes the mixing of leaf water pools, and the Péclet effect model, which incorporates transpiration rate and the mixing length between unenriched xylem and enriched mesophyll water in the mesophyll (Lm ) or veins (Lv ). Here we compare measurements and models of Δ18 OLW on two cell wall composition mutants grown under two light intensities and relative humidities to evaluate cell wall properties on leaf water transport. In maize (Zea mays), the compromised ultrastructure of the suberin lamellae in the bundle sheath of the ALIPHATIC SUBERIN FERULOYL TRANSFERASE mutant (Zmasft) reduced barriers to apoplastic water movement, resulting in higher E and, potentially, Lv and, consequently, lower Δ18 OLW . The difference in Δ18 OLW in cellulose synthase-like F6 (CslF6) mutants and wild-type of rice (Oryza sativa) grown under two light intensities co-varied with stomatal density. These results show that cell wall composition and stomatal density influence Δ18 OLW and that stable isotopes can facilitate the development of a physiologically and anatomically explicit water transport model.

Cell wall properties in Oryza sativa influence mesophyll CO2 conductance.

Diffusion of CO2 from the leaf intercellular air space to the site of carboxylation (gm ) is a potential trait for increasing net rates of CO2 assimilation (Anet ), photosynthetic efficiency, and crop productivity. Leaf anatomy plays a key role in this process; however, there are few investigations into how cell wall properties impact gm and Anet . Online carbon isotope discrimination was used to determine gm and Anet in Oryza sativa wild-type (WT) plants and mutants with disruptions in cell wall mixed-linkage glucan (MLG) production (CslF6 knockouts) under high- and low-light growth conditions. Cell wall thickness (Tcw ), surface area of chloroplast exposed to intercellular air spaces (Sc ), leaf dry mass per area (LMA), effective porosity, and other leaf anatomical traits were also analyzed. The gm of CslF6 mutants decreased by 83% relative to the WT, with c. 28% of the reduction in gm explained by Sc . Although Anet /LMA and Anet /Chl partially explained differences in Anet between genotypes, the change in cell wall properties influenced the diffusivity and availability of CO2 . The data presented here indicate that the loss of MLG in CslF6 plants had an impact on gm and demonstrate the importance of cell wall effective porosity and liquid path length on gm .

Relationship of leaf oxygen and carbon isotopic composition with transpiration efficiency in the C4 grasses Setaria viridis and Setaria italica.

Leaf carbon and oxygen isotope ratios can potentially provide a time-integrated proxy for stomatal conductance (gs) and transpiration rate (E), and can be used to estimate transpiration efficiency (TE). In this study, we found significant relationships of bulk leaf carbon isotopic signature (δ13CBL) and bulk leaf oxygen enrichment above source water (Δ18OBL) with gas exchange and TE in the model C4 grasses Setaria viridis and S. italica. Leaf δ13C had strong relationships with E, gs, water use, biomass, and TE. Additionally, the consistent difference in δ13CBL between well-watered and water-limited plants suggests that δ13CBL is effective in separating C4 plants with different availability of water. Alternatively, the use of Δ18OBL as a proxy for E and TE in S. viridis and S. italica was problematic. First, the oxygen isotopic composition of source water, used to calculate leaf water enrichment (Δ18OLW), was variable with time and differed across water treatments. Second, water limitations changed leaf size and masked the relationship of Δ18OLW and Δ18OBL with E. Therefore, the data collected here suggest that δ13CBL but not Δ18OBL may be an effective proxy for TE in C4 grasses.

Biochemical effects of salinity on oxygen isotope fractionation during cellulose synthesis.

The current isotope tree ring model assumes that 42% of the sucrose oxygen exchanges with stem water during cellulose synthesis and that the oxygen isotope biochemical fractionation is c. 27‰. However, previous studies have indicated that this model can overestimate the cellulose oxygen isotope ratio of plants under salinity or water stress. Saline stress increases soluble carbohydrates and osmolytes, which can alter exchange and biochemical fractionation during cellulose synthesis. To test the effect of salinity as well as the synthesis of osmolytes on exchange and biochemical fractionation, we grew wild-type and a transgenic mannitol synthesizer Arabidopsis thaliana hydroponically with fresh and saline water. We then measured the oxygen isotope ratios of leaf water, stem water and stem cellulose to determine the effects on exchange and biochemical fractionation. Biochemical fractionation did not change, but oxygen isotope exchange was twice as high for plants grown in saline water relative to freshwater-treated plants (0.64 and 0.3, respectively). Mannitol (osmolyte) synthesis did not affect exchange or biochemical fractionation regardless of salinity. Increases in salinity increased oxygen isotope exchange during cellulose synthesis, which may explain the overestimation of cellulose δ(18) O values under saline conditions.

The role of effective leaf mixing length in the relationship between the δ18 O of stem cellulose and source water across a salinity gradient.

Previous mangrove tree ring studies attempted, unsuccessfully, to relate the δ(18) O of trunk cellulose (δ(18) O(CELL) ) to the δ(18) O of source water (δ(18) O(SW) ). Here, we tested whether biochemical fractionation associated with one of the oxygen in the cellulose glucose moiety or variation in leaf water oxygen isotope fractionation (Δ(LW) ) can interfere with the δ(18) O(SW) signal as it is recorded in the δ(18) O(CELL) of mangrove (saltwater) and hammock (freshwater) plants. We selected two transects experiencing a salinity gradient, located in the Florida Keys, USA. The δ(18) O(CELL) throughout both transects did not show the pattern expected based on that of the δ(18) O(SW) . We found that in one of the transects, biochemical fractionation interfered with the δ(18) O(SW) signal, while in the other transect Δ(LW) differed between mangrove and hammock plants. Observed differences in Δ(LW) between mangroves and hammocks were caused by a longer effective leaf mixing length (L) of the water pathway in mangrove leaves compared to those of hammock leaves. Changes in L could have caused the δ(18) O(CELL) to record not only variations in the δ(18) O(SW) but also in Δ(LW) making it impossible to isolate the δ(18) O(SW) signal.