The relationship of abscisic acid metabolism to stomatal conductance in Douglas-fir during water stress

Changes in abscisic acid and its metabolites were followed through two drought cycles in Pseudotsuga menziesii (Mirb.) Franco seedlings to determine the metabolic pathway of the hormone and its relationship to branch (stomatal) conductance. Three year-old, intact seedlings were water-stressed, watered, and restressed over a period of 30 days. Water potential was sampled with a pressure chamber and branch conductance with a steady-state porometer. Needle content of abscisic acid and 2- trans -abscisic acid and their saponifiable conjugates were quantified with gas-liquid chromatography. The typical water potential threshold in branch conductance, decreasing abruptly at -2.0 MPa, corresponded to an increase in abscisic acid content of 240 ng g⁻¹. The relationship between abscisic acid and water potential was not definitive, though the general trend was an increase in the hormone with intensifying stress until water potential was -5.0 MPa, when concentration sharply declined. No adjustment to stress was observed in the relationships, but stress during the second cycle progressed more slowly. A linear relationship between abscisic acid and its conjugate indicated the importance of the interconversion of the two compounds for storage and supply of the free acid.     

Uptake and translocation of manganese in seedlings of two varieties of Douglas fir (Pseudotsuga menziesii var. viridis and glauca)

Douglas fir (Pseudotsuga menziesii) variety glauca (DFG) but not the variety viridis (DFV) showed symptoms of manganese (Mn) toxicity in some field sites. We hypothesized that these two varieties differed in Mn metabolism. To test this hypothesis, biomass partitioning, Mn concentrations, subcellular localization and 54Mn-transport were investigated. Total Mn uptake was three-times higher in DFG than in DFV. DFV retained > 90% of 54Mn in roots, whereas > 60% was transported to the shoot in DFG. The epidermis was probably the most efficient Mn barrier since DFV contained lower Mn concentrations in cortical cells and vacuoles of roots than DFG. In both varieties, xylem loading was restricted and phloem transport was low. However, sieve cells still contained high Mn concentrations. DFV displayed higher biomass production and higher shoot : root ratios than DFG. Our results clearly show that both varieties of Douglas fir differ significantly in Mn-uptake and allocation patterns rendering DFG more vulnerable to Mn toxicity.     

Dynamics of water transport and storage in conifers studied with deuterium and heat tracing techniques

The volume and complexity of their vascular systems make the dynamics of long-distance water transport in large trees difficult to study. We used heat and deuterated water (D2)) as tracers to characterize whole-tree water transport and storage properties in individual trees belonging to the coniferous species Pseudotsuga menziesii (Mirb.) Franco and Tsuga heterophylla (Raf.) Sarg. The trees used in this study spanned a broad range of height (13.5-58 m) and diameter (0.14-1.43 m). Sap flow was monitored continuously with heat dissipation probes near the base of the trunk prior to, during and following injection of D2O. The transit time for D2O transport from the base of the trunk to the upper crown and the tracer residence time were determined by measuring hydrogen isotope ratios in water extracted from leaves sampled at regular intervals. Transit times for arrival of D2O in the upper crown ranged from 2.5 to 21 d and residence times ranged from 36 to 79 d. Estimates of maximum sap velocity derived from tracer transit times and path length ranged from 2.4 to 5.4 m d(-1). Tracer residence time and half-life increased as tree diameter increased, independent of species. Species-independent scaling of tracer velocity with sapwood-specific conductivity was also observed. When data from this study were combined with similar data from an earlier study of four tropical angiosperm trees, species-independent scaling of tracer velocity and residence time with sapwood hydraulic capacitance was observed. Sapwood capacitance is an intrinsic tissue-level property that appears to govern whole-tree water transport in a similar manner among both tracheid- and vessel-bearing species.     

In vitro fertilization from co-cultured pollen tubes and female gametophytes of Douglas fir (Pseudotsuga menziesii)

 Our previous attempt on in vitro fertilization (IVF) in conifers resulted in pollen tube penetration of female gametophytes, but because of the rapid decline in egg viability, no further interaction occurred. In this report, we describe for the first time that IVF has been achieved in conifers. Using Douglas fir (Pseudotsuga menziesii), we describe a two-step process which involved induction of pollen tubes in culture followed by introduction of isolated female gametophytes at the tips of growing pollen tubes. Pollen tubes penetrated the introduced isolated female gametophytes at various places, but a number of tubes entered the egg cell through the neck cells similar to the in vivo condition. Under our current culture conditions, longevity of pollen tubes and eggs has been improved resulting in the release of sperms, fusion of gametes, and initial formation of the proembryo. Continued plasmolysis of the egg limited the number of successful gametic interactions. IVF has been accomplished in flowering plants in several ways, but the gametophyte-gametophyte IVF system described in this paper is unique. IVF offers a novel breeding technology that takes advantage of the sexual reproductive route. When coupled with hybridization and genetic transformation, IVF could result in the development of stable novel genotypes of economically superior trees. Received: 28 October 1997 / Accepted: 9 December 1997     

Close relationship between the state of the xanthophyll cycle pigments and photosystem II efficiency during recovery from winter stress

The potential involvement of the xanthophyll cycle in photoprotection of overwintering evergreen plants was investigated. Leaves from five evergreen species. Pseudotsuga menziesii, Pinus panderosa, Euonyums kiautschovicus. Mahonia repens and Malva neglecta, were collected from the field predawn during winter and transferred to the laboratory where chlorophyll fluorescence emission as well as the chlorophyll and carotenoid composition were ascertained periodically for 4.5 days. Leaves and needles from all species were found to have retained large amounts of the xanthophyll cycle pigments zeaxanthin and antheraxanthin, and they exhibited sustained low values of the intrinsic efficiency of photosystem II (PSII; measured as the ratio of variable to maximal fluorescence, F_v/F_m) upon collection. The increase in PSII efficiency was biphasic, with a rapid phase (requiring several hours) and a slow phase (requiring several days). Changes in the conversion state of the xanthophyll cycle were found to correlate with increases in PSII efficiency in both phases, with the latter phase involving large increases in both F_m (maximal fluorescence) and F_o (minimal fluorescence) throughout the period of recovery. The relationship between F_m quenching (expressed as nonphotochemical or Stern-Volmer quenching [NPQ] of F_m, i.e. F_m/ F_m–1) and F_o quenching (F_o/F_o–1) was linear, as expected for changes in xanthophyll cycle-dependent energy dissipation in the antenna complexes. Furthermore, the relationship between F_v/F_m and NPQ during recovery followed the theoretical relationship predicted for changes in the rate constant for energy dissipation in the antenna complexes. This fit between the theoretical relationship and the actual data indicates that all changes in NPQ or F_v/F_m can be accounted for by changes in this rate constant. The results suggest a role for the photoprotective xanthophyll cycle-dependent dissipation process in the lowered efficiency of PSII observed in coldstressed evergreen plants in the field.     

Metabolite profiling of Douglas-fir (Pseudotsuga menziesii) field trials reveals strong environmental and weak genetic variation

The primary objective of this study was to assess metabolomics for its capacity to discern biological variation among 10 full-sib families of a Douglas-fir tree breeding population, replicated on two sites. The differential accumulation of small metabolites in developing xylem was examined through metabolite profiles (139 metabolites common to 181 individual trees) generated by gas chromatography mass spectrometry and a series of statistical analyses that incorporated family, site, and tree growth and quantitative phenotypic wood traits (wood density, microfibril angle, wood chemistry and fiber morphology). Multivariate discriminant, canonical discriminant and factor analyses and broad-sense heritabilities revealed that metabolic and phenotypic traits alike were strongly related to site, while similar associations relating to genetic (family) structure were weak in comparison. Canonical correlation analysis subsequently identified correlations between specific phenotypic traits (i.e. tree growth, fibre morphology and wood chemistry) and metabolic traits (i.e. carbohydrate and lignin biosynthetic metabolites), demonstrating a coherent relationship between genetics, metabolism, environmental and phenotypic expression in wood-forming tissue. The association between cambial metabolites and tree phenotype, as revealed by metabolite profiling, demonstrates the value of metabolomics for systems biology approaches to understanding tree growth and secondary cell wall biosynthesis in plants.     

Hydraulic redistribution of soil water in two old-growth coniferous forests: quantifying patterns and controls

Although hydraulic redistribution of soil water (HR) by roots is a widespread phenomenon, the processes governing spatial and temporal patterns of HR are not well understood. We incorporated soil/plant biophysical properties into a simple model based on Darcy's law to predict seasonal trajectories of HR. We investigated the spatial and temporal variability of HR across multiple years in two old-growth coniferous forest ecosystems with contrasting species and moisture regimes by measurement of soil water content (theta) and water potential (Psi) throughout the upper soil profile, root distribution and conductivity, and relevant climate variables. Large HR variability within sites (0-0.5 mm d(-1)) was attributed to spatial patterns of roots, soil moisture and depletion. HR accounted for 3-9% of estimated total site water depletion seasonally, peaking at 0.16 mm d(-1) (ponderosa pine; Pinus ponderosa) or 0.30 mm d(-1) (Douglas-fir; Pseudotsuga menziesii), then declining as modeled pathway conductance dropped with increasing root cavitation. While HR can vary tremendously within a site, among years and among ecosystems, this variability can be explained by natural variability in Psi gradients and seasonal courses of root conductivity.     

Comparison between two methods of generating pressure—volume curves

Abstract. Pressure—volume (P—V) curves were generated on roots and shoots of coastal Douglas fir [ Pseudotsuga menziesii (Mirb.) Franco] seedlings using two procedures. In the first (Method A), samples were dehydrated inside a pressure chamber. Exuded stem sap was collected and weighed at successive pressure increases to derive the P—V curve. In the second method (Method B). excised samples were allowed to dry outside the pressure chamber by evapotranspiration. They were weighed periodically to determine sap loss and their corresponding balance pressures were determined in a pressure chamber in order to derive the P—V curve.
Estimates of volume averaged osmotic potential at full turgor and water potential at zero turgor which were derived graphically from the P—V curves, were different for each method. In general, estimates were more negative in Method A, by as much as 1.5 MPa in one case. Also, Method B did not record an osmotic adjustment in seedlings which were subjected to severe water stress while Method A did.