Signatures of local adaptation to current and future climate in phenology-related genes in natural populations of Quercus robur.

BACKGROUND: Local adaptation is a key evolutionary process that enhances the growth of plants in their native habitat compared to non-native habitats, resulting in patterns of adaptive genetic variation across the entire geographic range of the species. The study of population adaptation to local environments and predicting their response to future climate change is important because of climate change. RESULTS: Here, we explored the genetic diversity of candidate genes associated with bud burst in pedunculate oak individuals sampled from 6 populations in Poland. Single nucleotide polymorphism (SNP) diversity was assessed in 720 candidate genes using the sequence capture technique, yielding 18,799 SNPs. Using landscape genomic approaches, we identified 8 FST outliers and 781 unique SNPs in 389 genes associated with geography, climate, and phenotypic variables (individual/family spring and autumn phenology, family diameter at breast height (DBH), height, and survival) that are potentially involved in local adaptation. Then, using a nonlinear multivariate model, Gradient Forests, we identified vulnerable areas of the pedunculate oak distribution in Poland that are at risk from climate change. CONCLUSIONS: The model revealed that pedunculate oak populations in the eastern part of the analyzed geographical region are the most sensitive to climate change. Our results might offer an initial evaluation of a potential management strategy for preserving the genetic diversity of pedunculate oak.

Cambium Reactivation Is Closely Related to the Cell-Cycle Gene Configuration in Larix kaempferi.

Dormancy release and reactivation in temperate trees are mainly controlled by temperature and are affected by age, but the underlying molecular mechanisms are still unclear. In this study, we explored the effects of low temperatures in winter and warm temperatures in spring on dormancy release and reactivation in Larix kaempferi. Further, we established the relationships between cell-cycle genes and cambium cell division. The results showed that chilling accelerated L. kaempferi bud break overall, and the longer the duration of chilling is, the shorter the bud break time is. After dormancy release, warm temperatures induced cell-cycle gene expression; when the configuration value of the cell-cycle genes reached 4.97, the cambium cells divided and L. kaempferi reactivated. This study helps to predict the impact of climate change on wood production and provides technical support for seedling cultivation in greenhouses.

High Expression of ALTERNATIVE OXIDASE 2 in Latent Axillary Buds Suggests Its Key Role in Quiescence Maintenance in Rosebush.

Most vegetative axes remain quiescent as dormant axillary buds until metabolic and hormonal signals, driven by environmental changes, trigger bud outgrowth. While the resumption of growth activity is well documented, the establishment and maintenance of quiescence is comparatively poorly understood, despite its major importance in the adaptation of plants to the seasonal cycle or in the establishment of their shape. Here, using the rosebush Rosa hybrida 'Radrazz' as a plant model, we highlighted that the quiescent state was the consequence of an internal and active energy control of buds, under the influence of hormonal factors previously identified in the bud outgrowth process. We found that the quiescent state in the non-growing vegetative axis of dormant axillary buds displayed a low energy state along with a high expression of the ALTERNATIVE OXIDASE 2 (AOX2) and the accumulation of the corresponding protein. Conversely, AOX2 expression and protein amount strongly decreased during bud burst as energy status shifted to a high state, allowing growth. Since AOX2 can deviate electrons from the cytochrome pathway in the mitochondrial respiratory chain, it could drastically reduce the formation of ATP, which would result in a low energy status unfavorable for growth activities. We provide evidence that the presence/absence of AOX2 in quiescent/growing vegetative axes of buds was under hormonal control and thus may constitute the mechanistic basis of both quiescence and sink strength manifestation, two important aspects of budbreak.

Genomic signatures of natural selection at phenology-related genes in a widely distributed tree species Fagus sylvatica L.

BACKGROUND: Diversity among phenology-related genes is predicted to be a contributing factor in local adaptations seen in widely distributed plant species that grow in climatically variable geographic areas, such as forest trees. European beech (Fagus sylvatica L.) is widespread, and is one of the most important broadleaved tree species in Europe; however, its potential for adaptation to climate change is a matter of uncertainty, and little is known about the molecular basis of climate change-relevant traits like bud burst. RESULTS: We explored single nucleotide polymorphisms (SNP) at candidate genes related to bud burst in beech individuals sampled across 47 populations from Europe. SNP diversity was monitored for 380 candidate genes using a sequence capture approach, providing 2909 unlinked SNP loci. We used two complementary analytical methods to find loci significantly associated with geographic variables, climatic variables (expressed as principal components), or phenotypic variables (spring and autumn phenology, height, survival). Redundancy analysis (RDA) was used to detect candidate markers across two spatial scales (entire study area and within subregions). We revealed 201 candidate SNPs at the broadest scale, 53.2% of which were associated with phenotypic variables. Additive polygenic scores, which provide a measure of the cumulative signal across significant candidate SNPs, were correlated with a climate variable (first principal component, PC1) related to temperature and precipitation availability, and spring phenology. However, different genotype-environment associations were identified within Southeastern Europe as compared to the entire geographic range of European beech. CONCLUSIONS: Environmental conditions play important roles as drivers of genetic diversity of phenology-related genes that could influence local adaptation in European beech. Selection in beech favors genotypes with earlier bud burst under warmer and wetter habitats within its range; however, selection pressures may differ across spatial scales.

The genetic basis of adaptation in phenology in an introduced population of Black Cottonwood (Populus trichocarpa, Torr. & Gray).

BACKGROUND: Entering and exiting winter dormancy present important trade-offs between growth and survival at northern latitudes. Many forest trees display local adaptation across latitude in traits associated with these phenology transitions. Transfers of a species outside its native range introduce the species to novel combinations of environmental conditions potentially requiring different combinations of alleles to optimize growth and survival. In this study, we performed genome wide association analyses and a selection scan in a P. trichocarpa mapping population derived from crossings between clones collected across the native range and introduced into Sweden. GWAS analyses were performed using phenotypic data collected across two field seasons and in a controlled phytotron experiment. RESULTS: We uncovered 584 putative candidate genes associated with spring and autumn phenology traits as well as with growth. Many regions harboring variation significantly associated with the initiation of leaf shed and leaf autumn coloring appeared to have been evolving under positive selection in the native environments of P. trichocarpa. A comparison between the candidate genes identified with results from earlier GWAS analyses performed in the native environment found a smaller overlap for spring phenology traits than for autumn phenology traits, aligning well with earlier observations that spring phenology transitions have a more complex genetic basis than autumn phenology transitions. CONCLUSIONS: In a small and structured introduced population of P. trichocarpa, we find complex genetic architectures underlying all phenology and growth traits, and identify multiple putative candidate genes despite the limitations of the study population.

Differential springtime branch warming controls intra-crown nitrogen allocation and leaf photosynthetic traits in understory saplings of a temperate deciduous species.

Between-branch nitrogen competition is expected to be important during spring in temperate deciduous trees as nitrogen allocation would be higher in branches from earlier budburst than in those from later budburst. Such phenology-induced branch interaction would influence plant photosynthesis, but this has not been evaluated. Warming experiments were conducted on whole crowns (warmed trees; trunks and all branches of the same tree were warmed) or parts of the crowns (warmed branches with unwarmed control branches in the same tree), with unwarmed control trees, in saplings of the deciduous species Fraxinus lanuginosa. Spring leaf phenology and leaf photosynthetic traits were investigated to determine how the difference in temperature affects leaf phenology and photosynthetic traits. The timing of budburst was influenced by temperature-budburst was earlier in warmed trees and warmed branches than in control trees and control branches, but budburst timing did not differ between control trees and control branches or between warmed trees and warmed branches. In contrast, leaf traits were affected by the variation in phenology within crowns-nitrogen content and photosynthetic capacity were greater in the leaves of the warmed branches than in the control branches, but they did not differ between the leaves of warmed trees and control trees. Thus, branch warming altered the distribution of nitrogen between warmed and unwarmed branches as warmed branches developed faster, resulting in intracrown variation in leaf photosynthetic traits.

Varying temperature sensitivity of bud-burst date at different temperature conditions.

The relationship between the rate of development (DR) of bud-burst and temperature may be nonlinear, which could lead to varying temperature sensitivity (TS) of budburst date under different climate conditions. In order to determine the functional form between DR/TS and temperature, we gathered twigs with flower buds of five woody plants (Malus halliana, Forsythia suspense, Crataegus pinnatifida, Prunus cerasifera F. atropurpurea, and Berberis thunbergii var. atropurpurea Chenault) in early spring of 2017 at Beijing, and placed them in six growth chambers at same daylength but different temperature conditions (5, 10, 15, 20, 25, and 30 °C). The proportion of bud- burst was recorded every 2 or 3 days for each species at each temperature condition. The results showed that the proportion of bud-burst followed the logistic function over time at a given temperature. Subsequently, we developed a mathematical model to simulate the proportion of bud-burst at any temperature and date. The DR and TS were parameterized using a differential method. The simulation results showed that the DR increased monotonically with the rise in temperature, but only two species could reach the maximum value at 30 °C. The TS decreased with the increase in temperature, but this effect was weak when the temperature was high enough. These findings suggested that the predicted warming in the future may result in a slowdown in the advance of spring phenology of woody plants.

On quantifying the apparent temperature sensitivity of plant phenology.

Many plant phenological events are sensitive to temperature, leading to changes in the seasonal cycle of ecosystem function as the climate warms. To evaluate the current and future implications of temperature changes for plant phenology, researchers commonly use a metric of temperature sensitivity, which quantifies the change in phenology per degree change in temperature. Here, we examine the temperature sensitivity of phenology, and highlight conditions under which the widely used days-per-degree sensitivity approach is subject to methodological issues that can generate misleading results. We identify several factors, in particular the length of the period over which temperature is integrated, and changes in the statistical characteristics of the integrated temperature, that can affect the estimated apparent sensitivity to temperature. We show how the resulting artifacts can lead to spurious differences in apparent temperature sensitivity and artificial spatial gradients. Such issues are rarely considered in analyses of the temperature sensitivity of phenology. Given the issues identified, we advocate for process-oriented modelling approaches, informed by observations and with fully characterised uncertainties, as a more robust alternative to the simple days-per-degree temperature sensitivity metric. We also suggest approaches to minimise and assess spurious influences in the days-per-degree metric.

The initiation of bud burst in grapevine features dynamic regulation of the apoplastic pore size.

The physiological constraints on bud burst in woody perennials, including vascular development and oxygenation, remain unresolved. Both light and tissue oxygen status have emerged as important cues for vascular development in other systems; however, grapevine buds have only a facultative light requirement, and data on the tissue oxygen status have been confounded by the spatial variability within the bud. Here, we analysed apoplastic development at early stages of grapevine bud burst and combined molecular modelling with histochemical techniques to determine the pore size of cell walls in grapevine buds. The data demonstrate that quiescent grapevine buds were impermeable to apoplastic dyes (acid fuchsin and eosin Y) until after bud burst was established. The molecular exclusion size was calculated to be 2.1 nm, which would exclude most macromolecules except simple sugars and phytohormones until after bud burst. We used micro-computed tomography to demonstrate that tissue oxygen partial pressure data correlated well with structural heterogeneity of the bud and differences in tissue density, confirming that the primary bud complex becomes rapidly and preferentially oxygenated during bud burst. Taken together, our results reveal that the apoplastic porosity is highly regulated during the early stages of bud burst, suggesting a role for vascular development in the initial, rapid oxygenation of the primary bud complex.

Latitudinal clines in bud flush phenology reflect genetic variation in chilling requirements in balsam poplar, Populus balsamifera.

PREMISE: Boreal and northern temperate forest trees possess finely tuned mechanisms of dormancy, which match bud phenology with local seasonality. After winter dormancy, the accumulation of chilling degree days (CDD) required for rest completion before the accumulation of growing degree days (GDD) during quiescence is an important step in the transition to spring bud flush. While bud flush timing is known to be genetically variable within species, few studies have investigated variation among genotypes from different climates in response to variable chilling duration. METHODS: We performed a controlled environment study using dormant cuttings from 10 genotypes of Populus balsamifera, representing a broad latitudinal gradient (43-58°N). We exposed cuttings to varying amounts of chilling (0-10 weeks) and monitored subsequent GDD to bud flush at a constant forcing temperature. RESULTS: Chilling duration strongly accelerated bud flush timing, with increasing CDD resulting in fewer GDD to flush. Genotypic variation for bud flush was significant and stratified by latitude, with southern genotypes requiring more GDD to flush than northern genotypes. The latitudinal cline was pronounced under minimal chilling, whereas genotypic variation in GDD to bud flush converged as CDD increased. CONCLUSIONS: We demonstrate that increased chilling lessens GDD to bud flush in a genotype-specific manner. Our results emphasize that latitudinal clines in bud flush reflect a critical genotype-by-environment interaction, whereby differences in bud flush between southern vs. northern genotypes depend on chilling. Our results suggest selection has shaped chilling requirements and depth of rest as an adaptive strategy to avoid precocious flush in climates with midwinter warming.

Integrated analysis of miRNAs and their targets reveals that miR319c/TCP2 regulates apical bud burst in tea plant (Camellia sinensis).

MAIN CONCLUSION: The roles of microRNA-mediated epigenetic regulation were highlighted in the bud dormancy-activity cycle, implying that certain differentially expressed miRNAs play crucial roles in apical bud burst, such as csn-miR319c/TCP2. microRNAs (miRNAs) are a class of small non-coding RNAs that regulate gene expression by targeting mRNA transcripts for cleavage or directing translational inhibition. To investigate whether miRNAs regulate bud dormancy-activation transition in tea plant, which largely affects the yield and price of tea products and adaptability of tea trees, we constructed small RNA libraries from three different periods of bud dormancy-burst transition. Through sequencing analysis, 262 conserved and 83 novel miRNAs were identified, including 118 differentially expressed miRNAs. Quantitative RT-PCR results for randomly selected miRNAs exhibited that our comprehensive analysis is highly reliable and accurate. The content of caffeine increased continuously from the endodormancy bud to flushing bud, and differentially expressed miRNAs coupling with their targets associated with bud burst were identified. Remarkably, csn-miR319c was downregulated significantly from the quiescent bud to burst bud, while its target gene CsnTCP2 (TEOSINTE BRANCHED/CYCLOIDEA/PROLIFERATING CELL FACTOR 2) displayed opposite expression patterns. Co-transformation experiment in tobacco demonstrated that csn-miR319c can significantly suppress the functions of CsnTCP2. This study on miRNAs and the recognition of target genes could provide new insights into the molecular mechanism of the bud dormancy-activation transition in tea plant.

Responses of bud-break phenology to daily-asymmetric warming: daytime warming intensifies the advancement of bud break.

There is evidence that the ongoing climate change is happening through nighttime rather than daytime warming. How such a daily-asymmetric warming modifies plant phenology is still unclear. We investigated the effects of asymmetric warming on bud break by daily monitoring seedlings belonging to 26 black spruce [Picea mariana (Mill.) BSP.] and 15 balsam fir [Abies balsamea (L.) Mill.] provenances from the native range in Canada. Seedlings were subjected to either daytime or nighttime warming in three growth chambers at temperatures ranging between 10 and 24 °C. On average, a warming of 4 °C advanced the timings of bud break in both species by 2.4 days, with the later phases being more sensitive to the treatment. Bud break of both species responded more strongly to daytime warming, with the bud break occurred 1.2 and 3.2 days earlier under daytime than nighttime warming in black spruce and balsam fir, respectively. A marked ecotypic differentiation was only observed in black spruce that originated from provenances distributed broadly across Canada, with seedlings from the warmest provenance completing bud break 8.3 days later than those from the coldest one. However, no significant effect of provenance was observed for balsam fir, the narrowly distributed species. Overall, the above results suggest that a higher temporal resolution such as temperatures during daytime and nighttime, and higher spatial resolution should be taken into account to improve the accuracy of phenological model predictions under global change scenarios. Phenological models based on daily average temperature should take into account the diverging impacts of asymmetric warming on plant phenology. Our findings may indicate that the influence of warming on plant phenology may be less dramatic than expected.

Gene expression analysis of bud burst process in European hazelnut (Corylus avellana L.) using RNA-Seq.

The control of bud burst process depending on temperature is crucial factor in woody perennial plants to survive in unfavorable ecological conditions. Although it has important economic and agronomic values, little information is available on the molecular mechanism of the bud burst process in Corylus avellana. Here for the first time, we conducted a de novo transcriptome-based experiment using eco-dormant leaf bud tissues. Four transcriptome libraries were constructed from the leaf bud tissues and sequenced via Illumina platform. Transcriptome analysis revealed 86,394 unigenes with a mean length of 1189 nt and an N50 of 1916 nt. Among these unigenes, 63,854 (73.78%) of them were annotated by at least one database. De novo assembled transcripts were enriched in phenylpropanoid metabolism, phytohormone biosynthesis and signal transduction pathways. Analyses of phytohormone-associated genes revealed important changes during bud burst, in response to gibberellic acid, auxin, and brassinosteroids. Approximately 2163 putative transcription factors were predicted, of which the largest number of unique transcripts belonged to the MYB transcription factor family. These results contribute to a better understanding of the regulation of bud burst genes in perennial plants.

Bud break responds more strongly to daytime than night-time temperature under asymmetric experimental warming.

Global warming is diurnally asymmetric, leading to a less cold, rather than warmer, climate. We investigated the effects of asymmetric experimental warming on plant phenology by testing the hypothesis that daytime warming is more effective in advancing bud break than night-time warming. Bud break was monitored daily in Picea mariana seedlings belonging to 20 provenances from Eastern Canada and subjected to daytime and night-time warming in growth chambers at temperatures varying between 8 and 16 °C. The higher advancements of bud break and shorter times required to complete the phenological phases occurred with daytime warming. Seedlings responded to night-time warming, but still with less advancement of bud break than under daytime warming. No advancement was observed when night-time warming was associated with a daytime cooling. The effect of the treatments was uniform across provenances. Our observations realized under controlled conditions allowed to experimentally demonstrate that bud break can advance under night-time warming, but to a lesser extent than under daytime warming. Prediction models using daily timescales could neglect the diverging influence of asymmetric warming and should be recalibrated for higher temporal resolutions.

The timing of bud break in warming conditions: variation among seven sympatric conifer species from Eastern Canada.

Phenological changes are expected with the ongoing global warming, which could create mismatches in the growth patterns among sympatric species or create synchrony with insect herbivores. In this study, we performed a comparative assessment of the timings of bud break among seven conifer species of Eastern Canada by evaluating seedling development in growth chambers under different temperatures (16, 20 and 24 °C). Bud break occurred earliest in Larix laricina, while Pinus strobus and Pinus resinosa had the latest. Warmer conditions advanced bud break, with the greatest effects being observed at the lower temperatures. Mixed models estimated that one additional degree of temperature produced advancements of 5.3 and 2.1 days at 16 and 20 °C, respectively. The hypothesis of an asynchronous change between species under warming was demonstrated only for the last phenological phases (split buds and exposed shoots), and principally in pines. Abies balsamea showed changes in bud break comparable with the other species analysed, rejecting the hypothesis of mismatches under warmer conditions. The observed non-linear responses of the timings of bud break to warming suggest that the major changes in bud phenology should be expected at the lowest temperatures.

Simulation of forest tree species' bud burst dates for different climate scenarios: chilling requirements and photo-period may limit bud burst advancement.

This study investigates whether the assumed increase of winter and spring temperatures is depicted by phenological models in correspondingly earlier bud burst (BB) dates. Some studies assume that rising temperatures lead to an earlier BB, but even later BB has been detected. The phenological model PIM (promoter-inhibitor-model) fitted to the extensive phenological database of the German Weather Service was driven by several climate scenarios. This model accounts for the complicated mechanistic interactions between chilling requirements, temperature and photo-period. It predicts BB with a r 2 between 0.41 and 0.62 and a RMSE of around 1 week, depending on species. Parameter sensitivities depict species dependent interactions between growth and chilling requirements as well as photo-period. A mean trend to earlier BB was revealed for the period 2002- 2100, varying between -0.05 and -0.11 days per year, depending on species. These trends are lower than for the period 1951- 2009. Within climate scenario period, trends are decreasing for beech and chestnut, stagnating for birch and increasing for oak. Results suggest that not fulfilled chilling requirements accompanied by an increasing dependency on photo-period potentially limit future BB advancement. The combination of a powerful phenological model, a large scale phenological database and several climate scenarios, offers new insights into the mechanistic comprehension of spring phenology.

Seventeen-year trends in spring and autumn phenophases of Betula pubescens in a boreal environment.

Trends in the timing of spring and autumn phenophases of Betula pubescens were investigated in the southern, middle, and northern boreal zones in Finland. The field observations were carried out at 21 sites in the Finnish National Phenological Network in 1997-2013. The effective temperature sum of the thermal growth period, i.e. the sum of the positive differences between diurnal mean temperatures and 5 °C (ETS1), increased annually on average by 6-7 degree day units. Timing of bud burst remained constant in the southern and middle boreal zones but advanced annually by 0.5 day in the northern boreal zone. The effective temperature sum at bud burst (ETS2) showed no trend in the southern and middle boreal zones, whereas ETS2 increased on average from 20-30 to 50 degree day units in the northern boreal zone, almost to the same level as in the other zones. Increase in ETS2 indicates that the trees did not start their growth in very early spring despite warmer spring temperatures. The timing of leaf colouring and leaf fall remained almost constant in the southern boreal zones, whereas these advanced annually by 0.3 and 0.6 day in the middle boreal zone and by 0.6 and 0.4 day in the northern boreal zone, respectively. The duration of the growth period remained constant in all boreal zones. The results indicate high buffering capacity of B. pubescens against temperature changes. The study also shows the importance of the duration of phenological studies: some trends in spring phenophases had levelled out, while new trends in autumn phases had emerged after earlier studies in the same network for a shorter observation period.

Photoperiod constraints on tree phenology, performance and migration in a warming world.

Increasing temperatures should facilitate the poleward movement of species distributions through a variety of processes, including increasing the growing season length. However, in temperate and boreal latitudes, temperature is not the only cue used by trees to determine seasonality, as changes in photoperiod provide a more consistent, reliable annual signal of seasonality than temperature. Here, we discuss how day length may limit the ability of tree species to respond to climate warming in situ, focusing on the implications of photoperiodic sensing for extending the growing season and affecting plant phenology and growth, as well as the potential role of photoperiod in controlling carbon uptake and water fluxes in forests. We also review whether there are patterns across plant functional types (based on successional strategy, xylem anatomy and leaf morphology) in their sensitivity to photoperiod that we can use to predict which species or groups might be more successful in migrating as the climate warms, or may be more successfully used for forestry and agriculture through assisted migration schemes.

Sucrose is an early modulator of the key hormonal mechanisms controlling bud outgrowth in Rosa hybrida.

Sugar has only recently been identified as a key player in triggering bud outgrowth, while hormonal control of bud outgrowth is already well established. To get a better understanding of sugar control, the present study investigated how sugar availability modulates the hormonal network during bud outgrowth in Rosa hybrida. Other plant models, for which mutants are available, were used when necessary. Buds were grown in vitro to manipulate available sugars. The temporal patterns of the hormonal regulatory network were assessed in parallel with bud outgrowth dynamics. Sucrose determined bud entrance into sustained growth in a concentration-dependent manner. Sustained growth was accompanied by sustained auxin production in buds, and sustained auxin export in a DR5::GUS-expressing pea line. Several events occurred ahead of sucrose-stimulated bud outgrowth. Sucrose upregulated early auxin synthesis genes (RhTAR1, RhYUC1) and the auxin efflux carrier gene RhPIN1, and promoted PIN1 abundance at the plasma membrane in a pPIN1::PIN1-GFP-expressing tomato line. Sucrose downregulated both RwMAX2, involved in the strigolactone-transduction pathway, and RhBRC1, a repressor of branching, at an early stage. The presence of sucrose also increased stem cytokinin content, but sucrose-promoted bud outgrowth was not related to that pathway. In these processes, several non-metabolizable sucrose analogues induced sustained bud outgrowth in R. hybrida, Pisum sativum, and Arabidopsis thaliana, suggesting that sucrose was involved in a signalling pathway. In conclusion, we identified potential hormonal candidates for bud outgrowth control by sugar. They are central to future investigations aimed at disentangling the processes that underlie regulation of bud outgrowth by sugar.

Spatio-temporal relief from hypoxia and production of reactive oxygen species during bud burst in grapevine (Vitis vinifera).

BACKGROUND AND AIMS: Plants regulate cellular oxygen partial pressures (pO2), together with reduction/oxidation (redox) state in order to manage rapid developmental transitions such as bud burst after a period of quiescence. However, our understanding of pO2 regulation in complex meristematic organs such as buds is incomplete and, in particular, lacks spatial resolution. METHODS: The gradients in pO2 from the outer scales to the primary meristem complex were measured in grapevine (Vitis vinifera) buds, together with respiratory CO2 production rates and the accumulation of superoxide and hydrogen peroxide, from ecodormancy through the first 72 h preceding bud burst, triggered by the transition from low to ambient temperatures. KEY RESULTS: Steep internal pO2 gradients were measured in dormant buds with values as low as 2·5 kPa found in the core of the bud prior to bud burst. Respiratory CO2 production rates increased soon after the transition from low to ambient temperatures and the bud tissues gradually became oxygenated in a patterned process. Within 3 h of the transition to ambient temperatures, superoxide accumulation was observed in the cambial meristem, co-localizing with lignified cellulose associated with pro-vascular tissues. Thereafter, superoxide accumulated in other areas subtending the apical meristem complex, in the absence of significant hydrogen peroxide accumulation, except in the cambial meristem. By 72 h, the internal pO2 gradient showed a biphasic profile, where the minimum pO2 was external to the core of the bud complex. CONCLUSIONS: Spatial and temporal control of the tissue oxygen environment occurs within quiescent buds, and the transition from quiescence to bud burst is accompanied by a regulated relaxation of the hypoxic state and accumulation of reactive oxygen species within the developing cambium and vascular tissues of the heterotrophic grapevine buds.