Transcriptome analysis and functional validation reveal the novel role of LhCYCL in axillary bud development in hybrid Liriodendron.

Shoot branching significantly influences yield and timber quality in woody plants, with hybrid Liriodendron being particularly valuable due to its rapid growth. However, understanding of the mechanisms governing shoot branching in hybrid Liriodendron remains limited. In this study, we systematically examined axillary bud development using morphological and anatomical approaches and selected four distinct developmental stages for an extensive transcriptome analysis. A total of 9,449 differentially expressed genes have been identified, many of which are involved in plant hormone signal transduction pathways. Additionally, we identified several transcription factors downregulated during early axillary bud development, including a noteworthy gene annotated as CYC-like from the TCP TF family, which emerged as a strong candidate for modulating axillary bud development. Quantitative real-time polymerase chain reaction results confirmed the highest expression levels of LhCYCL in hybrid Liriodendron axillary buds, while histochemical ß-glucuronidase staining suggested its potential role in Arabidopsis thaliana leaf axil development. Ectopic expression of LhCYCL in A. thaliana led to an increase of branches and a decrease of plant height, accompanied by altered expression of genes involved in the plant hormone signaling pathways. This indicates the involvement of LhCYCL in regulating shoot branching through plant hormone signaling pathways. In summary, our results emphasize the pivotal role played by LhCYCL in shoot branching, offering insights into the function of the CYC-like gene and establishing a robust foundation for further investigations into the molecular mechanisms governing axillary bud development in hybrid Liriodendron.

Genome-Wide Identification and Expression Analysis of R2R3-MYB Family Genes Associated with Petal Pigment Synthesis in Liriodendron.

The MYB transcription factor family is one of the largest families in plants, and its members have various biological functions. R2R3-MYB genes are involved in the synthesis of pigments that yield petal colors. Liriodendron plants are widely cultivated as ornamental trees owing to their peculiar leaves, tulip-like flowers, and colorful petals. However, the mechanism underlying petal coloring in this species is unknown, and minimal information about MYB genes in Liriodendron is available. Herein, this study aimed to discern gene(s) involved in petal coloration in Liriodendron via genome-wide identification, HPLC, and RT-qPCR assays. In total, 204 LcMYB superfamily genes were identified in the Liriodendron chinense genome, and 85 R2R3-MYB genes were mapped onto 19 chromosomes. Chromosome 4 contained the most (10) R2R3-MYB genes, and chromosomes 14 and 16 contained the fewest (only one). MEME analysis showed that R2R3-MYB proteins in L. chinense were highly conserved and that their exon-intron structures varied. The HPLC results showed that three major carotenoids were uniformly distributed in the petals of L. chinense, while lycopene and ß-carotene were concentrated in the orange band region in the petals of Liriodendron tulipifera. Furthermore, the expression profiles via RT-qPCR assays revealed that four R2R3-MYB genes were expressed at the highest levels at the S3P/S4P stage in L. tulipifera. This result combined with the HPLC results showed that these four R2R3-MYB genes might participate in carotenoid synthesis in the petals of L. tulipifera. This work laid a cornerstone for further functional characterization of R2R3-MYB genes in Liriodendron plants.

Transcriptomic and microstructural analyses in Liriodendron tulipifera Linn. reveal candidate genes involved in nectary development and nectar secretion.

BACKGROUND: Nectar is a major floral attractant and reward for insects that ensures pollination. Liriodendron, a genus of the Magnoliaceae family, includes only two relict species, L. chinense and L. tulipifera, which are considered "basal angiosperms" according to plant evolutionary history. The flowers of Liriodendron plants are insect pollinated and secrete nectar to attract pollinators. To date, the morphology and anatomy of nectaries, the mechanism of nectar secretion and the molecular mechanism of nectary development in Liriodendron remain poorly understood. METHODS: In this study, we examined the nectary surface cells and change in starch in L. tulipifera by using scanning electron microscopy and periodic acid-Schiff techniques to select appropriate samples for subsequent research. Transcriptome sequencing was of the top and middle parts of immature nectaries and the middle part of mature and postsecretory nectaries in L. tulipifera was performed. We evaluated the expression profiles of 21 DEGs that are closely related to nectary development and nectar secretion for real-time quantitative PCR analysis. RESULTS: L. tulipifera nectaries are starch-storing nectaries and are located in the top and middle parts of L. tulipifera petals. After analyzing the RNA-seq data, we obtained 115.26 Gb of clean data in 12 libraries and mapped the results to the L. chinense reference genome with 71.02-79.77% efficiency. In total, 26,955 DEGs were identified by performing six pairwise comparisons. The flavonoid biosynthesis, phenylpropanoid biosynthesis, anthocyanin biosynthesis and starch and sucrose metabolism pathways were enriched and related to nectar secretion and pigment change. We identified 56 transcription factor families, and members of the TCP, Trihelix, C2H2, ERF, and MADS families changed dynamically during nectary development. Moreover, to further verify the accuracy of the RNA-seq results, we validated the expression profiles of 21 candidate genes. CONCLUSIONS: We evaluated the nectary development and secretion processes comprehensively and identified many related candidate genes in L. tulipifera. These findings suggest that nectaries play important roles in flavonoid synthesis and petal color presentation.

Effects of stem size on stem respiration and its flux components in yellow-poplar (Liriodendron tulipifera L.) trees.

Carbon dioxide (CO2) released from respiring cells in the stems of trees (RS) can diffuse radially to the atmosphere (EA) or dissolve in xylem sap and move internally in the tree (FT). Previous studies have observed that EA decreases as stem or branch diameter increases, but the cause of this relationship has not been determined, nor has the relationship been confirmed between stem diameter and RS, which includes both EA and FT. In this study, for the first time the mass balance technique was used to estimate RS of stems of Liriodendron tulipifera L. trees of different diameters, ranging from 16 to 60 cm, growing on the same site. The magnitude of the component fluxes scaled with tree size. Among the five trees, the contribution of EA to RS decreased linearly with increasing stem diameter and sapwood area while the contribution of FT to RS increased linearly with stem diameter and sapwood area. For the smallest tree EA was 86% of RS but it was only 46% of RS in the largest tree. As tree size increased a greater proportion of respired CO2 dissolved in sap and remained within the tree. Due to increase in FT with tree size, we observed that trees of different sizes had the same RS even though they had different EA. This appears to explain why the EA of stems and branches decreases as their size increases.

Fabrication of Sustained-release CA-PU Coaxial Electrospun Fiber Membranes for Plant Grafting Application.

Plant grafting is a well-known activity in orchards, greenhouses and vineyards gardens. However, low survival rate still limits the promotion of grafting and breeding of improved varieties. We report on polymeric fibers, obtained through coaxial electrospun, as carriers for the sustained release of drugs to heal wounds in plants. The CA-PU co-electrospun fibers show a rather uniform diameter, a smooth and hydrophilic surface. As long as 10days of sustained drugs release meets with the physiological requirement of plant grafting. The callus toxicity test shows that the CA-PU fibers are not toxic for plant cells. We show that loading the core of CA-PU fibers with 6-Benzylaminopurine (6-BA), a first-generation synthetic cytokinin that elicits plant growth and development, results in fibers that can efficiently promote the healing of plant wounds, thereby significantly improving the grafting survival rate.

Canopy gradients in leaf functional traits for species that differ in growth strategies and shade tolerance.

In temperate deciduous forests, vertical gradients in leaf mass per area (LMA) and area-based leaf nitrogen (Narea) are strongly controlled by gradients in light availability. While there is evidence that hydrostatic constraints on leaf development may diminish LMA and Narea responses to light, inherent differences among tree species may also influence leaf developmental and morphological response to light. We investigated vertical gradients in LMA, Narea and leaf carbon isotope composition (δ13C) for three temperate deciduous species (Carpinus caroliniana Walter, Fagus grandifolia Ehrh., Liriodendron tulipifera L.) that differed in growth strategy (e.g., indeterminate and determinate growth), shade tolerance and leaf area to sapwood ratio (Al:As). Leaves were sampled across a broad range of light conditions within three vertical layers of tree crowns to maximize variation in light availability at each height and to minimize collinearity between light and height. All species displayed similar responses to light with respect to Narea and δ13C, but not for LMA. Light was more important for gradients in LMA for the shade-tolerant (C. caroliniana) and -intolerant (L. tulipifera) species with indeterminate growth, and height (e.g., hydrostatic gradients) and light were equally important for the shade-tolerant (F. grandifolia) species with determinate growth. Fagus grandifolia had a higher morphological plasticity in response to light, which may offer a competitive advantage in occupying a broader range of light conditions throughout the canopy. Differences in responses to light and height for the taller tree species, L. tulipifera and F. grandifolia, may be attributed to differences in growth strategy or Al:As, which may alter morphological and functional responses to light availability. While height was important in F. grandifolia, height was no more robust in predicting LMA than light in any of the species, confirming the strong role of light availability in determining LMA for temperate deciduous species.

Impacts of biogeographic history and marginal population genetics on species range limits: a case study of Liriodendron chinense.

Species ranges are influenced by past climate oscillations, geographical constraints, and adaptive potential to colonize novel habitats at range limits. This study used Liriodendron chinense, an important temperate Asian tree species, as a model system to evaluate the roles of biogeographic history and marginal population genetics in determining range limits. We examined the demographic history and genetic diversity of 29 L. chinense populations using both chloroplast and nuclear microsatellite loci. Significant phylogeographic structure was recovered with haplotype clusters coinciding with major mountain regions. Long-term demographical stability was suggested by mismatch distribution analyses, neutrality tests, and ecological niche models (ENM) and suggested the existence of LGM refuges within mountain regions. Differences in genetic diversity between central and marginal populations were not significant for either genomic region. However, asymmetrical gene flow was inferred from central populations to marginal populations, which could potentially limit range adaptation and expansion of L. chinense.

Morphological and proteomic analysis reveal the role of pistil under pollination in Liriodendron chinense (Hemsl.) Sarg.

Pollination is an important physiological process during which interaction between pollen and pistil occurs. This interaction could determine whether or not fertilization will occur and hence the ratio of plant seed setting. Liriodendron chinense (Hemsl.) Sarg. (L. chinense) exhibits a distinct phenomenon where seed setting ratio is not more than 10% in natural environment. To explore the origin of this phenomenon, we conducted a comparative morphological and proteomic analysis on L. chinense pistils upon pollination. The morphological analysis showed that pollen grows well in vitro, but much slower on pistil or nutrient medium containing pistil extract. Proteomic analysis showed that 493 proteins had changed the expression after pollination. Among them, 468 and 51 proteins were identified by isobaric tags for relative and absolute quantitation and two-dimensional gel electrophoresis respectively, and 26 proteins were common in the two methods. After proteins functional categorization, 66 differentially expressed proteins that are involved in reproduction process were found. Further analysis showed that among the reproductive process related proteins, protein disulfide-isomerase A6 and four embryo-defective proteins showed closer relations with the low seed setting phenomenon. The results indicated that the element from pistil might be the main reason leading to low seed setting in L. chinense, which will provide new insights in the mechanisms underlying L. chinense reproduction process.

Exogenously applied 24-epi brassinolide reduces lignification and alters cell wall carbohydrate biosynthesis in the secondary xylem of Liriodendron tulipifera.

The roles of brassinosteroids (BRs) in vasculature development have been implicated based on an analysis of Arabidopsis BR mutants and suspension cells of Zinnia elegans. However, the effects of BRs in vascular development of a woody species have not been demonstrated. In this study, 24-epi brassinolide (BL) was applied to the vascular cambium of a vertical stem of a 2-year-old Liriodendron, and the resulting chemical and anatomical phenotypes were characterized to uncover the roles of BRs in secondary xylem formation of a woody species. The growth in xylary cells was clearly promoted when treated with BL. Statistical analysis indicated that the length of both types of xylary cells (fiber and vessel elements) increased significantly after BL application. Histochemical analysis demonstrated that BL-induced growth promotion involved the acceleration of cell division and cell elongation. Histochemical and expression analysis of several lignin biosynthetic genes indicated that most genes in the phenylpropanoid pathway were significantly down-regulated in BL-treated stems compared to that in control stems. Chemical analysis of secondary xylem demonstrated that BL treatment induced significant modification in the cell wall carbohydrates, including biosynthesis of hemicellulose and cellulose. Lignocellulose crystallinity decreased significantly, and the hemicellulose composition changed with significant increases in galactan and arabinan. Thus, BL has regulatory roles in the biosynthesis and modification of secondary cell wall components and cell wall assembly during secondary xylem development in woody plants.

Drought enhances symbiotic dinitrogen fixation and competitive ability of a temperate forest tree.

General circulation models project more intense and frequent droughts over the next century, but many questions remain about how terrestrial ecosystems will respond. Of particular importance, is to understand how drought will alter the species composition of regenerating temperate forests wherein symbiotic dinitrogen (N2)-fixing plants play a critical role. In experimental mesocosms we manipulated soil moisture to study the effect of drought on the physiology, growth and competitive interactions of four co-occurring North American tree species, one of which (Robinia pseudoacacia) is a symbiotic N2-fixer. We hypothesized that drought would reduce growth by decreasing stomatal conductance, hydraulic conductance and increasing the water use efficiency of species with larger diameter xylem vessel elements (Quercus rubra, R. pseudoacacia) relative to those with smaller elements (Acer rubrum and Liriodendron tulipifera). We further hypothesized that N2 fixation by R. pseudoacacia would decline with drought, reducing its competitive ability. Under drought, growth declined across all species; but, growth and physiological responses did not correspond to species' hydraulic architecture. Drought triggered an 80% increase in nodule biomass and N accrual for R. pseudoacacia, improving its growth relative to other species. These results suggest that drought intensified soil N deficiency and that R. pseudoacacia's ability to fix N2 facilitated competition with non-fixing species when both water and N were limiting. Under scenarios of moderate drought, N2 fixation may alleviate the N constraints resulting from low soil moisture and improve competitive ability of N2-fixing species, and as a result, supply more new N to the ecosystem.

Survival of a tertiary relict species, Liriodendron chinense (Magnoliaceae), in southern China, with special reference to village fengshui forests.

PREMISE OF THE STUDY: We investigate factors supporting the persistence in southern China of a rare Tertiary relict tree species, Liriodendron chinense, which has been almost eliminated by recent land use conversion. We hypothesize that cultural practices and traditional sustainable forest resource uses provide niches for the species' regeneration that will complement infrequent natural disturbances, while the species' survival on remote mountain slopes where there are no humans depends on natural disturbances alone. • METHODS: We examined and analyzed various landscape contexts, community associations, age distributions, and regeneration patterns of Liriodendron chinense. • KEY RESULTS: Forest communities containing Liriodendron chinense were of three types: (1) village fengshui forests-mature forests dominated by Tertiary relict taxa Liriodendron, Toona, and Emmenopterys, protected for their supposed spiritual value; (2) young secondary forests near villages, dominated solely by Liriodendron; and (3) old secondary forest remnants on mountain slopes far from villages, dominated by Liriodendron with other Tertiary relicts of the genera Davidia and Sassafras. The age structure of Liriodendron indicated ample recruitment in the first two forest types, where the activities of local people have provided regeneration niches for the survival of this shade-intolerant pioneer species. On the remote mountain slopes that have never been converted to agriculture, Liriodendron has survived through regeneration made possible by natural disturbances. • CONCLUSIONS: The traditional human land use, influenced by cultural values, has supplemented infrequent natural disturbances, providing regeneration niches for this and other Tertiary remnant species near villages in mountain valleys, while on uninhabited mountain slopes the species depends on natural disturbances to survive.

Contrasting the morphology, anatomy and fungal colonization of new pioneer and fibrous roots.

Not all roots born as first-order branches are the same and this has important consequences for overall function. We hypothesized that, compared with fibrous roots, pioneer roots are built to live longer at the expense of absorptive capacity. We tested this hypothesis by investigating pioneer and fibrous roots in their first 14 d of life in the arbuscular mycorrhizal tree species: Acer negundo, Acer saccharum, Juglans nigra, Liriodendron tulipifera and Populus tremuloides. Root observations were made with root-access boxes that allowed roots to be sampled at known ages in field-grown trees. Compared to fibrous roots, pioneer roots had larger diameter, lower specific root length, greater average length and a lack of mycorrhizal or nonmycorrhizal fungal colonization. Pioneer roots < 14 d old had more layers of hypodermis with a lower percentage of putative passage cells and more protoxylem groups than similar age fibrous roots. Our results suggest that pioneer roots are constructed for defense against biotic and abiotic challenges, exploration of soil distal to the stem, high fibrous root branching and secondary development with high axial hydraulic conductivity at the expense of mycorrhizal colonization and high absorptive capacity for water and nutrients.

Rapid characterization of biomass using near infrared spectroscopy coupled with multivariate data analysis: Part 1. Yellow-poplar (Liriodendron tulipifera L.).

This paper is the first of a two series papers on the use of near infrared (NIR) coupled with multivariate data analysis (MVDA) as a process analytical technology (PAT) tool for the rapid characterization of physical and chemical properties of two common West Virginian hardwood species, northern red oak (Quercus rubra) and yellow-poplar (Liriodendron tulipifera L.). These two wood species are potential feed stock for the bio-refinery industry. In Part 1, we report our results on yellow-poplar. The results of this study demonstrated that some preprocessing operations on the NIR spectra (first derivative) greatly improved all the prediction models developed in the study. Predictive PLS1 models developed using selective spectra regions, 1300-1800 nm and the full NIR region (800-2400 nm), were similar. The selective spectra region, 1300-1800 nm, included the first and second overtone of the NIR spectrum (1300-1800 nm). Measured and predicted physical and chemical properties of yellow-poplar yielded moderate to high correlation (R2).

Hardwood seedling growth on different mine spoil types with and without topsoil amendment.

The goal of many owners of reclaimed mined land in the Appalachian region is to restore the diverse native hardwood forest for environmental, economic, and cultural reasons. However, native hardwoods often grow poorly on mined sites because they are planted in unsuitable spoils devoid of native topsoil. In a greenhouse experiment, we examined the suitability of four growth media available for use on many mined sites in the central Appalachians-forest topsoil (FT), weathered sandstone (WS), unweathered sandstone (US), and unweathered shale (UH)-as well as the effects of topsoil amendment (none vs. amended) on the growth of three native hardwood species: Fraxinus americana, Quercus rubra, and Liriodendron tulipifera. A 4 x 2 x 3 factorial greenhouse experiment was conducted with planted 1-yr-old seedlings. Tree growth, foliar nutrients, and soil properties were measured and characterized. The WS was the spoil most conducive to growth for F. americana and Q. rubra. Liriodendron tulipifera did not respond to any treatments. Tree growth was highly correlated with mineralizable soil nitrogen and extractable soil phosphorus. Topsoil amendment significantly increased growth on the UH but not on the US or WS. Topsoil amendment increased the number of native herbaceous plants growing in the pots and improved foliar nutrient content in F. americana and L. tulipifera. Many properties of the WS, such as pH, microbial activity, and water availability, more closely approximated the control soil than the US or UH. This study showed that trees are sensitive to spoil type and that certain spoil types that are conducive to good growth of native trees should be used during the reclamation process, particularly if forest topsoil is not applied. Forest topsoil amendment improved tree growth on some spoil materials, improved tree nutrition, and helped restore the native soil organisms and plants that were present before mining.

Long-term habitat selection and chronic root herbivory: explaining the relationship between periodical cicada density and tree growth.

Periodical cicadas (Magicicada spp.) are insect herbivores that feed on host tree roots, but their distribution among hosts is determined largely by the oviposition of female cicadas in the previous generation. A pattern of decreasing tree growth rates with increasing cicada densities is predicted when considering the costs of chronic root herbivory, but the opposite pattern is expected when considering adaptive habitat selection. Here, we report observations indicating that the relationship between periodical cicada densities and host tree growth rates is hump shaped. We suggest that both herbivory and habitat selection are likely to be key processes explaining this pattern, resulting in regions of positive and negative correlation. These results suggest that the effects of cicada herbivory are most apparent at relatively high cicada densities, while habitat selection tends to distribute cicada herbivory on host trees that are able to compensate for cicada root herbivory up to threshold cicada densities.

[Foraging behavior of various tree species in heterogeneous nutrient environment related to light condition].

A factorial combination pot experiment was designed to study the growth response and foraging behavior of three South China forestation species Pinus massoniana, Schima superba and Liriodendron chinense under heterogenous nutrient envionment and different light conditions. The results demonstrated that under full light, the plant height and biomass of P. massoniana and S. superba were enhanced in heterogenous nutrient environment, with the root biomass increased by 99.5% and 66.7%, respectively, compared with that in homogenous environment; while L. chinense exhibited singnificant growth benefits in homogenous environment. The root scale, foraging precision, and nutrient uptake of S. superba reduced markedly under shading, which in turn decreased the growth benefits in heterogenous nutrient environment. The influence of shading was weaker on P. massoniana than on S. superba, still with some growth benefits for P. massoniana due to its root proliferation and enhanced nutrient uptake in nutrient rich patch. There was a slight influence of shading on the foraging precision and sensitivity of L. chinense root, with no growth response pattern change to heterogenous nutient environment occurred. It was concluded that three tree species not only differed in the inheritance of root foraging mechanism and behavior, but also varied in the response pattern to light condition. Therefore, different measures should be adopted in forestation with the three tree species to enhance the forest productivity.

Nonadditive effects of leaf litter species diversity on breakdown dynamics in a detritus-based stream.

Since species loss is predicted to be nonrandom, it is important to understand the manner in which those species that we anticipate losing interact with other species to affect ecosystem function. We tested whether litter species diversity, measured as richness and composition, affects breakdown dynamics in a detritus-based stream. Using full-factorial analyses of single- and mixed-species leaf packs (15 possible combinations of four dominant litter species; red maple [Acer rubrum], tulip poplar [Liriodendron tulipifera], chestnut oak [Quercus prinus], and rhododendron [Rhododendron maximum]), we tested for single-species presence/absence (additive) or species interaction (nonadditive) effects on leaf pack breakdown rates, changes in litter chemistry, and microbial and macroinvertebrate biomass. Overall, we found significant nonadditive effects of litter species diversity on leaf pack breakdown rates, which were explained both by richness and composition. Leaf packs containing higher litter species richness had faster breakdown rates, and antagonistic effects of litter species composition were observed when any two or three of the four litter species were mixed. Less-consistent results were obtained with respect to changes in litter chemistry and microbial and macroinvertebrate biomass. Our results suggest that loss of litter species diversity will decrease species interactions involved in regulating ecosystem function. To that end, loss of species such as eastern hemlock (Tsuga canadensis) accompanied by predicted changes in riparian tree species composition in the southeastern United States could have nonadditive effects on litter breakdown at the landscape scale.

Variations in organic acid exudation and aluminum resistance among arbuscular mycorrhizal species colonizing Liriodendron tulipifera.

Aluminum (Al) in acidic soils is toxic to plants, affecting growth, water uptake and nutrient assimilation. Aluminum resistance in some plant species and genotypes has been ascribed to organic acid exudation from roots and arbuscular mycorrhizal (AM) fungal symbiosis. We investigated variation among several AM species in altering Al resistance of Liriodendron tulipifera L. and evaluated AM influence on organic acid production as a potential Al resistance mechanism. Growth, nutritional responses and rhizosphere organic acid profiles were assessed for seedlings in association with Acaulospora morrowiae Spain & Schenck, Glomus claroideum Schenck & Smith, G. clarum Nicol. & Schenck or Paraglomus brasilianum (Spain & Miranda) Morton & Redecker and non-mycorrhizal seedlings exposed to 0, 50 or 200 microM Al. Plants colonized by G. clarum had the greatest biomass, least Al and most phosphorus (P) in leaf tissues and exuded malate and citrate into the rhizosphere at rates that complexed 99% of delivered Al in all treatments. Other AM fungi did not confer significant Al resistance on L. tulipifera and did not maintain citrate and malate exudation in response to Al exposure. This study illustrates functional diversity among AM fungal species in conferred Al resistance to plants and highlights the potential importance of fungal diversity in ecosystem responses to environmental stresses.

Diurnal and seasonal changes in stem increment and water use by yellow poplar trees in response to environmental stress.

To evaluate indicators of whole-tree physiological responses to climate stress, we determined seasonal, daily and diurnal patterns of growth and water use in 10 yellow poplar (Liriodendron tulipifera L.) trees in a stand recently released from competition. Precise measurements of stem increment and sap flow made with automated electronic dendrometers and thermal dissipation probes, respectively, indicated close temporal linkages between water use and patterns of stem shrinkage and swelling during daily cycles of water depletion and recharge of extensible outer-stem tissues. These cycles also determined net daily basal area increment. Multivariate regression models based on a 123-day data series showed that daily diameter increments were related negatively to vapor pressure deficit (VPD), but positively to precipitation and temperature. The same model form with slight changes in coefficients yielded coefficients of determination of about 0.62 (0.57-0.66) across data subsets that included widely variable growth rates and VPDs. Model R2 was improved to 0.75 by using 3-day running mean daily growth data. Rapid recovery of stem diameter growth following short-term, diurnal reductions in VPD indicated that water stored in extensible stem tissues was part of a fast recharge system that limited hydration changes in the cambial zone during periods of water stress. There were substantial differences in the seasonal dynamics of growth among individual trees, and analyses indicated that faster-growing trees were more positively affected by precipitation, solar irradiance and temperature and more negatively affected by high VPD than slower-growing trees. There were no negative effects of ozone on daily growth rates in a year of low ozone concentrations.