Plasticity in leaf‐level water relations of tropical rainforest trees in response to experimental drought

Summary The tropics are predicted to become warmer and drier, and understanding the sensitivity of tree species to drought is important for characterizing the risk to forests of climate change. This study makes use of a long‐term drought experiment in the Amazon rainforest to evaluate the role of leaf‐level water relations, leaf anatomy and their plasticity in response to drought in six tree genera. The variables (osmotic potential at full turgor, turgor loss point, capacitance, elastic modulus, relative water content and saturated water content) were compared between seasons and between plots (control and through‐fall exclusion) enabling a comparison between short‐ and long‐term plasticity in traits. Leaf anatomical traits were correlated with water relation parameters to determine whether water relations differed among tissues. The key findings were: osmotic adjustment occurred in response to the long‐term drought treatment; species resistant to drought stress showed less osmotic adjustment than drought‐sensitive species; and water relation traits were correlated with tissue properties, especially the thickness of the abaxial epidermis and the spongy mesophyll. These findings demonstrate that cell‐level water relation traits can acclimate to long‐term water stress, and highlight the limitations of extrapolating the results of short‐term studies to temporal scales associated with climate change.


Fig. S1
Relationships between (a) the symplastic fraction of the spongy mesophyll, and (b) the spongy mesophyll symplast volume per area with spongy mesophyll thickness. Eschweilera coriacea and (c) Swartzia racemosa.

Table S1
Slope coefficients for correlations of PV parameters against the symplastic fraction of tissue thickness.
Methods S1 Correlation analysis of symplastic tissue volume vs leaf water relations. Osmotic potential at full turgor and turgor loss point pertain only to the symplast volume of cells. If differences exist in these parameters between tissue types within the leaf one might expect a correlation to exist between the thickness of the particular tissue with the osmotic parameter.
However, such relationship, if one exists, could become decoupled from tissue thickness due to changes in cell size and apoplastic fraction. Therefore, a separate analysis was performed to examine the correlations without the volumetric apoplastic fraction. Because this was just an analysis of symplast volume, the cavity volume was not analysed in these models. Otherwise, the analysis was conducted in exactly the same way as the analysis of the tissue thickness. Therefore, the starting structure of the models, using SM as an example, was: Y ~ Ad symp + Pal symp + SM symp + Ab symp To make this analysis correspond to the analysis of tissue thickness, the symplastic thickness of each tissue was also found as a proportion of total leaf thickness. Proportional measurements were not found by summing the fractional symplastic contribution because this resulted in a high degree of interdependence between values. Thus, SM prop_symp = SM symp / leaf thickness, not SM symp / (Ad symp +Pal symp +SM symp +Ad symp ).

Results and Discussion
Mean cell volume of all tissue layers increased significantly with tissue thickness. This was expected in the case of the epidermal layers which are one cell thick, and for the palisade which was often one cell thick (Fig. S2), but not expected for the SM (R 2 = 0.16, P = 0.002). Because SM cells were assumed to be spherical, the surface area to volume ratio was expected to decrease non-linearly with volume and, therefore, the relationship between symplastic fraction and SM thickness was also predicted to be non-linear (Fig. S1).
The analysis of symplastic volumes with PV parameters provided results similar to those of the tissue thickness (compare Table S1 with Table 4). The absolute values of spongy mesophyll correlate with Ψ π tlp , Ψ π o , SWC and RWC tlp in both analyses, although in the symplastic analysis the proportional measurements also correlate with Ψ π tlp , SWC and RWC tlp . Perhaps the biggest difference between analyses is that neither the absolute or proportional measurements of the palisade correlate with anything. As the analysis of tissue thickness combines the apoplast and symplast, it is possible that disparity between the two analyses (symplast vs thickness) indicates a functional role for the apoplast. However, the results of the symplast analysis must be interpreted with caution due to the assumptions (listed below) made to derive symplast volume and to the reduced degrees of freedom of the analysis.
Assumptions required to determine symplast volume: -Cells accurately represented by designated shape e.g. SM cell is spherical, palisade is cylindrical, epidermal cells are cuboid.
-Cell walls are a comparable thickness to the 14 species measured by Buckley (2015) -Cuticle accounts for negligible proportion of leaf thickness.