Morine Lempereur
email: morine.lempereur@cefe.cnrs.fr

Jean-Marc Limousin

Frédéric Guibal

Jean-Marc Ourcival

Serge Rambal

Julien Ruffault

Florent Mouillot

Recent climate hiatus revealed dual control by temperature and drought on the stem growth of Mediterranean Quercus ilex

Keywords: Basal area increment, climate change, climate-growth response, climate hiatus, drought, growth duration, growth phenology, Quercus ilex

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INTRODUCTION

Stem growth, recorded in annual tree rings, is a synthetic surrogate of carbon input in standing biomass [START_REF] Babst | Above-ground woody carbon sequestration measured from tree rings is coherent with net ecosystem productivity at five eddycovariance sites[END_REF] and an index for tree vitality and fitness [START_REF] Bigler | Growth-dependent tree mortality models based on tree rings[END_REF]Benito-Garzon et al., 2013). In return, it is the record of the yearly climatic conditions that have enabled stem growth over a period lasting from decades to centuries [START_REF] Fritts | Tree rings and climate[END_REF][START_REF] Briffa | Treering width and density data around the northern hemisphere: Part 1, local and regional climate signals[END_REF]. Understanding the factors that control stem growth is therefore crucial to assess the impact of past and future climate change on forests, but remains a major scientific issue due to a series of currently unresolved uncertainties.

First, the climate sensitivity of tree growth is generally considered to depend on photosynthesis and respiration fluxes because most process-based models assume that tree growth is carbon limited [START_REF] Davi | Sensitivity of water and carbon fluxes to climate changes from 1960 to 2100 in European forest ecosystems[END_REF][START_REF] Gaucherel | Changes of the potential distribution area of French Mediterranean forests under global warming[END_REF][START_REF] Keenan | Predicting the future of forests in the Mediterranean under climate change, with niche-and process-based models: CO2 matters![END_REF]. Carbon allocation to sapwood is therefore the key process driving the carbon sink, and is at the heart of tree growth simulations in most terrestrial biosphere models (e.g. [START_REF] Sitch | Evaluation of ecosystem dynamics, plant geography and terrestrial carbon cycling in the LPJ dynamic global vegetation model[END_REF][START_REF] Friedlingstein | Climate-Carbon Cycle Feedback Analysis: Results from the C4MIP Model Intercomparison[END_REF][START_REF] Fisher | Assessing uncertainties in a second-generation dynamic vegetation model caused by ecological scale limitations[END_REF]. Several allocation schemes exist (e.g. fixed allocation, the pipe model, and hierarchical allocation between plant organs, [START_REF] Schippers | Tree growth variation in the tropical forest: understanding effects of temperature, rainfall and CO2[END_REF], but there is accumulating experimental evidence that cambial activity (sink limitation) is more sensitive to environmental stressors than carbon assimilation (source limitation ;[START_REF] Körner | Carbon limitation in trees[END_REF][START_REF] Fatichi | Moving beyond photosynthesis: from carbon source to sink-driven vegetation modeling[END_REF][START_REF] Guillemot | The dynamic of annual carbon allocation to wood in European forests is consistent with a combined source-sink limitation of growth: implications for modelling[END_REF][START_REF] Körner | Paradigm shift in plant growth control[END_REF]Delpierre et al., 2016a). Taking sink limitation into account in terrestrial biosphere models is thus likely to improve their ability to simulate past and future forest productivity [START_REF] Leuzinger | Beyond global change: lessons from 25 years of CO2 research[END_REF][START_REF] Fatichi | Moving beyond photosynthesis: from carbon source to sink-driven vegetation modeling[END_REF], but a better characterization of the phenology and climate sensitivity of wood formation is a prerequisite [START_REF] Rossi | Predicting xylem phenology in black spruce under climate warming[END_REF][START_REF] Rossi | Lengthening of the duration of xylogenesis engenders disproportionate increases in xylem production[END_REF][START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF]Delpierre et al., 2016a;Delpierre et al., 2016b).

Second, global climate change may simultaneously exert opposing influences on forest functioning, and these impacts are difficult to disentangle. For example, increased water use efficiency due to atmospheric CO2 enrichment is sometime reported to increase tree growth in spite of concurrently increasing aridity (Martinez-Vilalta et al., 2008;Koutouvas 2013), but the opposite finding is far more common [START_REF] Peñuelas | Increased water-use efficiency during the 20th century did not translate into enhanced tree growth[END_REF]. Warming also exerts opposing impacts on tree growth: on one hand it increases the length of the growing season [START_REF] Keeling | Increased activity of northern vegetation inferred from atmospheric CO2 measurements[END_REF][START_REF] Dragoni | Evidence of increased net ecosystem productivity associated with a longer vegetated season in a deciduous forest in south-central Indiana, USA[END_REF] while on the other hand it exacerbates atmospheric evaporative demand and hence water limitation [START_REF] Angert | Drier summers cancel out the CO2 uptake enhancement induced by warmer springs[END_REF][START_REF] Zhao | Drought-induced reduction in global terrestrial net primary production from 2000 through[END_REF]. Warming may therefore affect tree growth positively (e.g. [START_REF] Rossi | Lengthening of the duration of xylogenesis engenders disproportionate increases in xylem production[END_REF] or negatively (e.g. [START_REF] Brzostek | Chronic water stress reduces tree growth and the carbon sink of deciduous hardwood forests[END_REF] depending on the ecosystem under consideration.

Third, relationships between tree growth and climate may change under the influence of climate change (e.g. [START_REF] Briffa | Reduced sensitivity of recent tree-growth to temperature at high northern latitudes[END_REF][START_REF] Büntgen | Summer temperature variations in the European Alps, a.d. 755-2004[END_REF][START_REF] Carrer | Long-term change in the sensitivity of tree-ring growth to climate forcing in Larix decidua[END_REF][START_REF] D'arrigo | On the 'divergence problem' in northern forests: A review of the tree-ring evidence and possible causes[END_REF]. A classic example of this phenomenon is the so-called 'divergence problem' in northern forests, which is a weakening of the positive temperature response of tree growth in strongly temperature limited ecosystems since the middle of the 20 th century [START_REF] D'arrigo | On the 'divergence problem' in northern forests: A review of the tree-ring evidence and possible causes[END_REF]. The consequence of such temporal variations in tree growth sensitivity to climate is that empirical dendrochronological models based on statistical links between tree growth and climate cannot be extrapolated to long term historical and future climate conditions with confidence (e.g. [START_REF] Gea-Izquierdo | Growth projections reveal local vulnerability of Mediterranean oaks with rising temperatures[END_REF][START_REF] Subedi | Climate-diameter growth relationships of black spruce and jack pine trees in boreal Ontario, Canada[END_REF].

Fourth, climate change itself is a complex phenomenon that may result in changes in the mean climate (IPCC 2014), in the frequency of extreme climatic events [START_REF] Meehl | More intense, more frequent, and longer lasting heat waves in the 21st century[END_REF] and in seasonality [START_REF] Giorgi | Higher Hydroclimatic Intensity with Global Warming[END_REF][START_REF] Ruffault | Differential regional responses in drought length, intensity and timing to recent climate changes in a Mediterranean forested ecosystem[END_REF], all of which may have contrasted impacts on tree growth and function. In addition, climate change is not monotonous and the recent slowing down of the warming trend, early identified by [START_REF] Easterling | Is the climate warming or cooling?[END_REF] and later called the 'climate hiatus' [START_REF] Trenberth | An apparent hiatus in global warming?[END_REF], is currently a topic of increasing scientific interest. But if most scientific studies have focused on explaining the causes and processes behind this 'climate hiatus' or pause in atmospheric warming [START_REF] Meehl | Model-based evidence of deep-ocean heat uptake during surface-temperature hiatus periods[END_REF][START_REF] Balmaseda | Distinctive climate signals in reanalysis of global ocean heat content[END_REF][START_REF] England | Recent intensification of wind-driven circulation in the Pacific and the ongoing warming hiatus[END_REF][START_REF] Steinman | Atlantic and Pacific multidecadal oscillations and Northern hemisphere temperatures[END_REF], few have attempted to assess its ecological consequences, especially for processes that are highly sensitive to climate, such as tree growth.

In water limited ecosystems, where forest productivity is primarily limited by drought [START_REF] Churkina | Contrasting climatic controls on the estimated productivity of global terrestrial biomes[END_REF][START_REF] Zhao | Drought-induced reduction in global terrestrial net primary production from 2000 through[END_REF], the supply of water from rainfall appears to be the key variable to explain tree growth [START_REF] Babst | Site-and species-specific responses of forest growth to climate across the European continent[END_REF]. In the case of the Mediterranean species Quercus ilex, the conclusions of dendrochronological studies converge more specifically to identify winter and spring rainfall amounts as the key drivers of annual stem growth [START_REF] Gutiérrez | Climate controls act at different scales on the seasonal pattern of Quercus ilex L. stem radial increments in NE Spain[END_REF]. In contrast, Q. ilex stem growth is generally poorly correlated with winter and spring temperatures. Daily measurements of basal area increment over an eight year period (2004)(2005)(2006)(2007)(2008)(2009)(2010)(2011) with automatic dendrometers in a Q. ilex forest, demonstrated that the duration of spring growth was an accurate predictor of annual stem growth [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF]. After observing that the duration of spring growth was determined by winter temperature for growth start and a lower threshold of predawn plant water potential of -1.1 MPa for growth cessation, [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] proposed a new hypothesis wherein the annual growth of Q. ilex is under the dual control of winter temperature and spring water limitation.

In this contextual framework, we used both tree ring records over a 40-year period and an annual diameter inventory over a 30-year period to (i) test whether the stem growth phenology approach proposed by [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] can explain Q. ilex growth over a long retrospective period with different growth estimation methods, (ii) assess the relative importance of drought and temperature in limiting Q. ilex growth under a Mediterranean climate, and (iii) evaluate the stationarity of these limitations in recent decades. In particular, we aimed to identify whether any temporal shift in the climate controls of Q. ilex growth (drought or temperature plus drought) are concomitant to, and the functional consequence of, the recent pause in atmospheric warming.

MATERIALS AND METHODS

Site description

The study site is located 35 km north-west of Montpellier (southern France), on a flat plateau, in the Puéchabon state forest (43°44'29"N, 3°35'45"E, 270 m a.s.l.). This forest has been managed as a coppice for centuries and the last clearcut took place in 1942. Vegetation is largely dominated by a dense overstory of the evergreen oak Quercus ilex. In 2014, the average top canopy height was 5.5 m and the stem density was 4700 (±700) stems ha -1 . Understory evergreen species, Buxus sempervirens, Phyllirea latifolia, Pistacia lentiscus and Juniperus oxycedrus, compose a sparse shrubby layer (height < 2 m) with less than 25% cover. The climate is Mediterranean, with 80% of the rainfall occurring between September and April.

Average annual precipitation for the 1984-2013 period was 916 mm (550 to 1549 mm). Mean annual temperature over the same period was 13.2 °C with a minimum in January (5.5 °C) and a maximum in July (22.9 °C). The very shallow bedrock has a hard Jurassic limestone origin.

The volumetric fraction of stones and rocks averages 0.75 in the top 0-50 cm and 0.90 below.

The stone free fine fraction of the soil in the 0-50 cm layer is a homogeneous silty clay loam (USDA texture triangle) comprising 38.8% clay, 35.2% silt and 26% sand.

Monitoring annual stem growth

Stem diameter at breast height (DBH) was measured annually from 1986 to 2013 on 319 trees distributed in eight circular plots (diameter 20 m) within an area of 2 ha. The 319 trees used in this field survey (hereafter FS, Fig. 1) were selected so as to include all the diameter size classes in 1986. DBH was measured every winter under dry conditions using a diameter tape at a height identified by a paint mark on the stem. In January 2008, the stem diameters were distributed as follows: 58% < 8 cm DBH, 24% between 8 and 10 cm and 18% >10 cm.

Because the smaller size classes exhibited a very low growth and a high mortality rate, we restricted our analysis to the 125 trees with a DBH > 8 cm in 2008 to obtain a stronger growth signal (Table 1). From 2004 to 2013, stem growth was measured more frequently and with greater accuracy using automatic band dendrometers on a subset of trees in two neighboring plots (for more details, see [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF]. Automatic dendrometers (ELPA-98, University of Oulu, Finland; hereafter AD) were set up at a height of 1.3 m above the ground on 6 to 12 trees with DBH > 7 cm. The two datasets (FS and AD) were strongly correlated during their overlapping period (Table 1), thus demonstrating the relative accuracy of FS in estimating the annual growth of the largest trees.

Dendrochronological procedures

In 2005 and 2008, 15 and 12 stems, respectively, were selected among the largest size classes of the field survey (FS) sample (10 cm < DBH < 16 cm) and cut down to measure ringwidth chronology. Cross sections were collected at stump height, air dried, sanded and polished (40 to 400 grit). The longest growth radii in each cross section were selected and compared pairwise under a binocular magnifier. In old coppiced oaks, rings are narrow and sometimes (hereafter RW) were obtained from 1942 to 2008 (Table 1, Fig. 1). Cross dating was facilitated by the presence of frost rings corresponding to the severe winters of 1963, 1985 and 1987. These rings, typical for years with severe freezing episodes, were abnormally wide because the cells of their initial area were crushed and dislocated, a typical constraint on tree ring interpretation in woody species under a Mediterranean climate [START_REF] Cherubini | Identification, measurement and interpretation of the tree rings in woody species from mediterranean climates[END_REF]. This particular feature made them useful as markers for cross dating but prevented the reliable measurement of annual stem growth, so these years were excluded from subsequent analyses.

Cross correlation coefficients on annual tree ring indices calculated among the 12 individual series were all higher than 0.6 (P-value<0.05). The ring width measured at stump height and along the longest growth radius was then rescaled to the tree DBH at the time of the cut. This required correction for the tapering between stump height (20 cm) and breast height (130 cm) as well as correction for the bark thickness (see Fig. S1 for details). The average tree ring width after rescaling, which was used to calculate the basal area increment (BAI), was 954 µm (CV = 48%) between 1942 and 2008, and 792 µm (CV = 38%) over the period 1968-2008 used in our study.

Climate variables

The Puéchabon meteorological station (hereafter Puéchabon) is located in a clearing 200 m away from the study plot and has provided daily on-site climate data since 1984 (Fig. 1).

Precipitation was measured with a tipping bucket rain gauge (ARG100; Environmental Measurements, Sunderland, UK) calibrated to 0.2 mm per tip and placed 1 m above the ground, air temperature was recorded with a MP100 sensor (Rotronic, Bassersdorf, Switzerland) at a height of 2 m, and net radiation was measured with a pyranometer (SKS1110; Skye Instruments, UK) at a height of 2 m above the ground.

To extend our analysis before 1984, we used climate variables (daily rainfall, daily minimum and maximum temperature) from the St Martin-de-Londres meteorological station (hereafter SML; 43°47'06''N; 3°43'48''E, altitude 194 m a.s.l., located about 12 km away from the study site; source Meteo-France) over the period 1966-2013 (Fig. 1). The mean daily temperature was calculated as the average of the minimum and maximum daily temperature.

The climate data from SML station were compared to on-site measurements in the overlap period 1984-2013 (Table S1 and Fig. S2). The comparison showed close agreement for temperature but a higher rainfall amount in SML than in Puéchabon. No significant biases (slope not different from 1, and distance to origin not different from 0) were observed between the two meteorological stations in the two crucial variables related to stem growth: the mean temperature from January to March (TJFM) and the sum of precipitation from April to June (PAMJ; Fig. S2).

Daily solar radiation at the SML station was calculated from the processing chain described in [START_REF] Kumar | Modelling topographic variation in solar radiation in a GIS environment[END_REF]. In the first step, theoretical clear sky solar radiation (Ra) was calculated from the daily timing of sunrise and sunset, the bi-hourly sun azimuthal angle and the atmospheric transmittance according to longitude using R Cran Packages 'RAtmosphere' and 'oce'. No topographical effects were taken into consideration. Actual solar radiation (Rs, in MJ m -2 day -1 ) was then calculated from maximum and minimum daily temperature (Tmax and Tmin, respectively) and the clear sky theoretical radiation following the Hargreaves equation:

, where the adjustment coefficient kRs was set to 0.16 for interior location [START_REF] Allen | Crop evapotranspiration-Guidelines for computing crop water requirements-FAO Irrigation and drainage paper 56[END_REF]. Results of the Hargreaves equation were validated against solar radiation measured in Puéchabon since 1998.

Modeling predawn leaf water potential and water stress integral

Soil water storage integrated over the rooting depth, c.a. 4.5 m, was measured during the vegetative periods of 1984-1986 and from July 1998 to August 2009 at approximately monthly intervals, using a neutron moisture gauge (503DR Hydroprobe, CPN, Concord, CA, USA).

Discrete measurements were interpolated at a daily time step using the soil water balance model described by [START_REF] Rambal | The differential role of mechanisms for drought resistance in a Mediterranean evergreen shrub: a simulation approach[END_REF] and further used by [START_REF] Rambal | How drought severity constrains gross primary production(GPP) and its partitioning among carbon pools in a Quercus ilex coppice?[END_REF]. The model was driven by daily values of incoming solar radiation, minimum and maximum temperatures, and rainfall amount. Potential evapotranspiration was computed using the Priestley-Taylor equation [START_REF] Priestley | On the Assessment of Surface Heat Flux and Evaporation Using Large-Scale Parameters[END_REF]. The reduced major axis (RMA) regression between neutron moisture gauge measurements and model simulations yielded an R 2 of 0.93, the slope was 0.94 ± 0.05 (P < 0.0001, n = 91) and the intercept not significantly different from 0. Soil water storage and soil water potential were linked by a Campbell-type retention curve [START_REF] Campbell | Soil physics with basic: Transport models for soil-plant systems[END_REF] whose parameters are strongly dependent on soil texture [START_REF] Saxton | Estimating generalized soil-water characteristics from texture[END_REF][START_REF] Rambal | Drought controls over conductance and assimilation of a Mediterranean evergreen ecosystem: scaling from leaf to canopy[END_REF]. Predawn leaf water potential used for model validation was measured about eight times a year between April and October from 2003 to 2009 on a subsample of four trees among those equipped with automatic dendrometers (see [START_REF] Limousin | Morphological and phenological shoot plasticity in a Mediterranean evergreen oak facing long-term increased drought[END_REF]. RMA regressions between measured and simulated values of predawn leaf water potential yielded an R 2 of 0.84, the slope was 0.93 ± 0.05 (P < 0.0001, n = 54), and the intercept was not significantly different from 0. We used the simulations of predawn water potential rather than soil water content, as the former is more closely linked with plant functioning [START_REF] Rambal | Drought controls over conductance and assimilation of a Mediterranean evergreen ecosystem: scaling from leaf to canopy[END_REF]. The daily simulations of predawn water potential were performed with the climate data from the Puéchabon and SML stations for the periods 1984-2012 and 1966-2012, respectively. The water stress integral (WSI), defined by [START_REF] Myers | Water stress integral-a link between short-term stress and long-term growth[END_REF] as the seasonally or yearly sum of predawn water potential, was used as a drought severity index to quantify annual or seasonal water stress.

  a Rs s R T T k R min max   (Priestley

Duration of spring basal area increment: calculations of dates t0 and t1

The dates of stem growth phenology that bounded the spring growth period, the DOY (day of the year) when stem growth starts (hereafter t0) and the DOY when stem growth stops in early summer (hereafter t1, Fig. S3), were estimated for each year. The relationships between t0 or t1 and climate variables were calibrated by [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] on observations of t0 and t1 obtained with automatic dendrometers from 2004 to 2011. Briefly, t0 was defined as the first day at which basal area exceeded the culmination of the previous year, and t1 as the first day when BAI became null or negative (see [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] for methodological details). t0 and t1 were estimated using climate data from both Puéchabon and the nearby SML station over their corresponding timeframes. t0 was predicted by a nonlinear relationship with the mean temperature from January to March (TJFM). The relationships were fitted between t0 and TJFM measured in Puéchabon (t0 = 849.2*exp(-0.6436 TJFM)+121; R² = 0.95; RMSE = 2.6 days; [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] and between t0 and TJFM measured in SML (t0 = 4300*exp(-0.943 TJFM)+124.4; R² = 0.95; RMSE = 2.3 days; Fig. S4). t1 was predicted by the DOY when the plant water potential simulated using climate data from Puéchabon reached a threshold of -1.1 MPa (R² = 0.75; RMSE = 7 days, [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF], corresponding to a DOY when the plant water potential simulated using climate data from SML reached a threshold of -1.1 MPa + ɛ (with ɛ = -0.1; R² = 0.73; RMSE = 13.5 days) due to the slight differences in temperature and precipitation between the two stations (see Fig. S5). The duration ∆tt0-t1 corresponds to the duration of the period between t0 and t1 and was computed for each pair of simulated phenological thresholds.

Data processing and statistical analyses

Annual stem growth (DBH and ring widths) was converted into annual basal area increments (BAI, expressed in mm² year -1 ) for the two data series. Analyses were performed with averaged BAI values of 125 and 5-12 individual trees for FS and RW, respectively. The year 1992 presented two periods when predawn plant water potential fell below the threshold of -1.1 MPa, in spring and autumn (Fig. S6). This unusual bi-modal drought prevented us from determining the duration ∆tt0t1 so 1992 was excluded from subsequent analyses. The growth series BAIFS included measurements that appeared to be outliers compared to BAIRW or BAIAD for the years 2002 and 2010 (see Fig. S7). In 2002, BAIFS was particularly low when BAIRW was close to the expected value, and in 2010, BAIFS was clearly higher than the BAIAD value.

Because BAIFS is less accurate than the other two methods, and was calculated as the difference between two subsequent measurements of DBH, these two values were considered to be unreliable and excluded from the analyses performed on BAIFS.

To investigate the links between tree growth and climate, we computed the correlations between our two different growth datasets (FS and RW) and a set of relevant climate predictors.

For each correlation tested, we reported the Pearson's correlation coefficient and its significance based on the standard bootstrap method with 1000 samples taken from the original distribution of climate and tree ring data. Within each sample, the number of observations was random and followed a geometric distribution. We used the 'treeclim' R package built upon the DENDROCLIM2002 statistical tool dedicated to tree ring analysis [START_REF] Biondi | DENDROCLIM2002: A C++ program for statistical calibration of climate signals in tree-ring chronologies[END_REF][START_REF] Zang | treeclim: an R package for the numerical calibration of proxy-climate relationships[END_REF]. Climate predictors were derived from temperature and precipitation at monthly, seasonal and annual time scales, and from our functional index of spring growth duration (∆tt0t1). To test for the stationarity of the drought and temperature controls over the studied period, we also performed the same bootstrap sampling Pearson correlation analysis on a 10-year moving window along the whole series.

The temporal trends in stem growth and climate variables were characterized using both trend tests and breakpoint analyses. Temporal trends in time series were estimated using the Theil-Sen test [START_REF] Sen | Estimates of the Regression Coefficient Based on Kendall's Tau[END_REF]) from the 'openair' R cran package, applied on a 1000 block bootstrap simulations to account for auto-correlated variables [START_REF] Kunsch | The jackknife and the bootstrap for general stationary observations[END_REF], with the block length set to n/3, n being the length of the time series. Temporal breakpoints in the time series were assessed by computing the yearly F-statistic (sequential F-test) and the supF statistic was then used to test for their significance.

RESULTS

Sensitivity of stem growth to the dual temperature-drought drivers

Fig. 2 Time series of annual stem basal area increment (BAI) for the ring width series (RW;1942-2008), the longterm field survey (FS;[START_REF] Saxton | Estimating generalized soil-water characteristics from texture[END_REF][START_REF] Koutavas | CO2 fertilization and enhanced drought resistance in greek firs from Cephalonia Island, Greece[END_REF] and the automatic dendrometer series [START_REF] Piovesan | Drought-driven growth reduction in old beech (Fagus sylvatica L.) forests of the central Apennines, Italy[END_REF]2004[START_REF] Koutavas | CO2 fertilization and enhanced drought resistance in greek firs from Cephalonia Island, Greece[END_REF]. Error bars are standard errors among the sampled trees (see Table 1 for sample size).

Annual stem growth (BAIRW) from 1942 to 2008 exhibited two distinct phases, with a significant breakpoint around 1967 (Fstat = 81.96; P-value<0.0001; Fig. 2). The first phase lasted 25 years after the clearcut and showed a linear increase of 5.5 (±0.62) mm² y -1 in annual BAI. This phase, which corresponds approximately to the cut frequency formerly used in the traditional management of this Q. ilex coppice [START_REF] Floret | Dynamics of holm oak (Quercus ilex L.) coppices after clearcutting in southern France[END_REF], was characterized by incomplete canopy cover and notable self-thinning. The second phase showed a stabilization of BAIRW around a mean value of 203.8 (±54.84) mm² y -1 . The influence of climate on stem growth was consequently only considered from 1968 on, to avoid the confounding effects caused by changes in competition, space-filling and self-thinning during the first phase.

The two long-term growth series BAIRW and BAIFS were significantly correlated with each other during their overlap period 1986-2008 (r = 0.55, P-value < 0.05, Table 1). Both series of BAI exhibited large between year variations (coefficient of variation, CV = 27% and 29% for in 1995 to 95 days in 2008 (Fig. 3). BAI was linearly correlated with ∆tt0-t1 for both stem growth series (R² = 0.56; P-value < 0.0001 and R² = 0.35; P-value < 0.01 for RW and FS respectively; Fig. 3). Moreover, ∆tt0-t1 was the best explanatory variable for the inter-annual variations in BAIRW compared to other climate variables over the period 1984-2008 (Table 2; 3). We thus concluded that the dual control of annual stem growth by temperature and precipitation, stated by [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] for the period 2004-2011, remained valid for a longer retrospective period and when growth is measured with less accuracy than with automatic dendrometers. The intercepts of the linear relationships between BAI and ∆tt0-t1, which represent the residual autumnal growth [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF], differed significantly between the two BAI series (F = 17.97; P-value < 0.001), with 122.9 (±34.42) mm² y -1 and 95.5 (±39.09) mm² y -1 for RW and FS, respectively (Fig. 3). By contrast, the slopes of these relationships did not differ significantly between RW and FS (F = 0.46; P-value > 0.05), and the common slope equals 1.23 mm² day -1 . The sensitivity of BAIRW to ∆tt0-t1 was thus identical to the sensitivity of BAIFS.

Furthermore, neither the extension of the study period to 1968-2008 nor the switch in meteorological stations from Puéchabon to SML, removed the significant correlation between BAIRW and ∆tt0-t1 (R² = 0.31; P-value < 0.001; Table 2).

Stem growth response to climate trends from 1968 to 2013

Over the longer period 1968-2008, ∆tt0-t1 was not a better predictor of BAI than the sum of spring precipitation (PAMJ) or t1 alone (r = 0.75; P-value < 0.001 and r = 0.66; P-value < 0.001 respectively; Table 2, 3), and we also observed a loss of correlation between BAI and t0 (r = 0.18; P-value = 0.31; Table 2).

PAMJ was the main explanatory climate variable for t1 (over the period 1968-2008: r = 0.80; P-value<0.001) and it exhibited no temporal trend between 1968 and 2013 (Pvalue = 0.31; Fig. 4b). However, spring water limitation increased (WSI in spring: -0.48 MPa day y -1 ; P-value<0.001; Fig. 4d, Table S3) as a result of increasing spring potential evapotranspiration (PET; +0.028 mm day -1 y -1 ; P-value<0.001; Fig. 4c, Table S3). This trend in PET was mainly due to the significant warming trend in spring temperatures throughout the period 1968-2013 (+0.07°C y -1 ; P-value<0.001; Fig. 4a, Table S3). As a result, drought onset t1 exhibited a temporal trend toward earlier dates (-0.57 day y -1 ; P-value<0.01; Fig. 5). The correlation between BAIRW and PAMJ remained significant, however, throughout most of the 1968-2008 period (Fig. 6a). In parallel, TJFM increased significantly throughout the period (+0.04 °C y -1 ; P-value<0.05; Fig. 4a) which led to a significant trend of t0 toward earlier dates (-0.21 day y -1 ; P-value<0.05; Fig. 5). However, a pause in the atmospheric winter warming occurred in the last decade. This warming pause is apparent in the discrepancy between actual and expected TJFM obtained from the temporal trend built over the period 1978-1998. The year 1978 was chosen because it corresponds to the onset of a rapid warming phase in the Northern Hemisphere [START_REF] Mann | Northern hemisphere temperatures during the past millennium: Inferences, uncertainties, and limitations[END_REF]. Similarly, the year 1998 was considered as the onset of the climate hiatus observations (IPCC 2014). We used the period 1978-1998 to build the reference trend line and extrapolate expected TJFM values over the 1999-2013 period. We then tested for a breakpoint in the anomalies between observed and expected TJFM over the whole period. Such a breakpoint was identified in 2002, indicating a significant slowing down in winter warming after this date 1 9 7 0 1 9 8 0 1 9 9 0 2 0 0 0 2 0 1 0

-5 0 0 -4 0 0 -3 0 0 -2 0 0 -1 0 0 0 A M J Y E A R ( d )
(Fstat = 21.52; P<0.001; Fig. 4a). This temporal pattern was also observed at seven other meteorological stations in the region surrounding our study site (Table S4 and Fig. S8), thus confirming the regional occurrence of the globally observed climate hiatus [START_REF] Trenberth | An apparent hiatus in global warming?[END_REF]. The date of growth onset (t0) was very sensitive to the warming pause in TJFM and a breakpoint in the t0 trend was observed in 2002 (Fstat = 21.52; P<0.001; Fig. 5). We also observed that the start of the winter warming pause coincided with a change in the climate controls on annual stem growth. Indeed, the correlation between BAI and TJFM, which was only significant over one 10-year window between 1968 and 1994, became constantly significant from the time window 1995-2004 on (Fig. 6b). The temporal trends in t0 and t1, both toward earlier dates (-10 and -26 days, respectively) offset each other to some extent until 2002 and resulted in a non-significant trend in ∆tt0-t1. After 2002, the prolonged trend in t1 continued to decrease while t0 stabilized, thereby leading to a sudden shortening of the growing season (Fig. 5). The timing of this phenological reduction was concomitant with the appearance of a significant relationship between TJFM and BAI (Fig. 6b). The decline in ∆tt0-t1 induced by the winter warming pause resulted in a decrease in BAIRW after 2002. This decreasing trend over time become significant after the 10-year time window [START_REF] Kumar | Modelling topographic variation in solar radiation in a GIS environment[END_REF]-2006 (Fig. 7a,b) (Fig. 7a,b). BAIFS exhibited a similar decreasing trend in the same period, but of lower amplitude and not significant.

DISCUSSION

Growth duration as a predictor of annual stem growth

The duration of spring stem growth (∆tt0-t1) was the best explanatory variable of yearly basal area increment for both BAIFS and BAIRW after 1984. This confirms earlier results obtained with automatic dendrometers over a shorter period and a smaller sample of trees [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF], and therefore validates ∆tt0-t1 as a robust estimator of Q. ilex annual stem growth over long time scales and large samples. The two independent samplings we used here differed markedly in the number of trees measured, the measurement method and its precision. FS corresponds to a large number of trees (125 trees) representing a broader tree size distribution but measured with a less precise resolution (0.3 mm of DBH), while RW was measured on a small sample of large dominant trees (12 trees) but with a better resolution (0.01 mm of radius). Average stem growth was lower for BAIFS than for BAIRW, but both responded linearly and with the same sensitivity to ∆tt0-t1 (Fig. 3).

The spring stem growth duration ∆tt0-t1 is defined as the length of the period between the onset of spring stem growth (t0) and the date of growth cessation caused by summer drought (t1). [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] observed that t0 was well predicted by winter temperature, and that t1 occurred when the leaf predawn water potential dropped below a threshold of -1.1 MPa. A similar approach was used by [START_REF] Rossi | Predicting xylem phenology in black spruce under climate warming[END_REF][START_REF] Rossi | Lengthening of the duration of xylogenesis engenders disproportionate increases in xylem production[END_REF] who linked the stem growth of boreal black spruce with the duration of xylem growth delimited by cold temperatures in spring and autumn, and by Delpierre et al. (2016a) who linked the growth of temperate oak to the timing of the summer growth cessation controlled by water limitation. By identifying the climatic limits for cell division and elongation, the temporal delimitation of tree growth duration is an innovative way of improving simulation of forest carbon sink response to climate change [START_REF] Fatichi | Moving beyond photosynthesis: from carbon source to sink-driven vegetation modeling[END_REF]. Tree growth has generally been simulated using two types of models: statistical dendroclimatological models, which are empirical and may lead to marked uncertainties when extrapolated to future climate scenarios (e.g. [START_REF] Gea-Izquierdo | Growth projections reveal local vulnerability of Mediterranean oaks with rising temperatures[END_REF][START_REF] Babst | Above-ground woody carbon sequestration measured from tree rings is coherent with net ecosystem productivity at five eddycovariance sites[END_REF]; and process based models that link tree growth to carbon assimilation (e.g. [START_REF] Sitch | Evaluation of ecosystem dynamics, plant geography and terrestrial carbon cycling in the LPJ dynamic global vegetation model[END_REF][START_REF] Friedlingstein | Climate-Carbon Cycle Feedback Analysis: Results from the C4MIP Model Intercomparison[END_REF][START_REF] Gaucherel | Changes of the potential distribution area of French Mediterranean forests under global warming[END_REF][START_REF] Keenan | Predicting the future of forests in the Mediterranean under climate change, with niche-and process-based models: CO2 matters![END_REF], ignoring experimental evidence showing a direct, and generally more limiting, climate control of plant growth [START_REF] Körner | Carbon limitation in trees[END_REF][START_REF] Muller | Water deficits uncouple growth from photosynthesis, increase C content, and modify the relationships between C and growth in sink organs[END_REF]. Projecting tree growth based on growth phenology as in [START_REF] Rossi | Predicting xylem phenology in black spruce under climate warming[END_REF] and [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] merges the biological realism of the first approach, i.e. a direct link between climate and growth, with the mechanistic understanding of tree physiology used in process based models, and be a significant step forward in projecting the response of forest ecosystems to climate change (Delpierre et al., 2016b). Showing that growth duration is an adequate predictor of tree growth for long time series datasets (40 years in our study) and coarse temporal resolution (annual diameter measurements) is thus an important preliminary.

Nevertheless, the spring growth duration approach remains limited by both its conditions of applicability and the large proportion of the growth variability not explained by ∆tt0-t1. The particular case of the year 1992, with its bi-modal drought in spring and autumn and its wellwatered summer (Fig. S6), suggests that ∆tt0-t1 can only be used as a descriptor of stem growth phenology in years with typical Mediterranean seasonality and a drought period occurring in summer. While the lowest threshold of predawn water potential below which stem growth is prevented certainly remains valid for any drought seasonality [START_REF] Hsiao | Sensitivity of growth of roots versus leaves to water stress: biophysical analysis and relation to water transport[END_REF][START_REF] Muller | Water deficits uncouple growth from photosynthesis, increase C content, and modify the relationships between C and growth in sink organs[END_REF][START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF], we hypothesize that the relationship between stem growth and the duration of periods with adequate growth conditions (the rate of growth) varies in years with peculiar phenology. Variations in the rate of growth may also account for the unexplained variability in the BAI against ∆tt0-t1 relationships (44% and 65% for RW and FS, respectively; Fig. 3). Finally, the relative extent of autumn growth compared to spring growth would deserve more detailed investigation. [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF] observed that autumn growth explained only approximately 30% of annual stem growth and was strongly determined by spring conditions, but its magnitude over long time periods and under climate change conditions remains to be described.

Dual limitation of growth by winter cold and spring-summer drought

Summers in the Mediterranean region are typically characterized by the concomitance of low rainfall, high temperature and high solar radiation, while winters are cold and humid.

Consequently, vegetation functioning is limited by water deficit in summer and by low temperatures in winter [START_REF] Mitrakos | Plant life under Mediterranean climatic conditions[END_REF][START_REF] Terradas | The influence of summer and winter stress and water relationships on the distribution of Quercus ilex L[END_REF]. A cessation of growth during cold and dry periods is frequently observed in Mediterranean trees which display a bi-phasic growth pattern over the year [START_REF] Cherubini | Tree-life history prior to death: two fungal root pathogens affect tree-ring growth differently[END_REF][START_REF] Campelo | Climatic significance of tree-ring width and intra-annual density fluctuations in Pinus pinea from a dry Mediterranean area in Portugal[END_REF][START_REF] Montserrat-Marti | Summer-drought constrains the phenology and growth of two coexisting Mediterranean oaks with contrasting leaf habit: implications for their persistence and reproduction[END_REF][START_REF] Camarero | Plastic bimodal xylogenesis in conifers from continental Mediterranean climates[END_REF][START_REF] Gutiérrez | Climate controls act at different scales on the seasonal pattern of Quercus ilex L. stem radial increments in NE Spain[END_REF].

The sensitivity of growth to water deficit is well established in the literature [START_REF] Lockhart | An analysis of irreversible plant cell elongation[END_REF][START_REF] Hsiao | Sensitivity of growth of roots versus leaves to water stress: biophysical analysis and relation to water transport[END_REF][START_REF] Muller | Water deficits uncouple growth from photosynthesis, increase C content, and modify the relationships between C and growth in sink organs[END_REF], as is the negative impact of drought on ring width [START_REF] Fritts | Tree rings and climate[END_REF]. Previous dendroclimatological studies generally stated that spring to earlysummer precipitation was the main driver of annual stem growth of Q. ilex [START_REF] Gutiérrez | Climate controls act at different scales on the seasonal pattern of Quercus ilex L. stem radial increments in NE Spain[END_REF]. More precisely, it is the start of the dry season, defined by a threshold of soil water deficit that was identified as the main determinant of variations in inter-annual growth [START_REF] Maselli | Start of the dry season as a main determinant of inter-annual Mediterranean forest production variations[END_REF][START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF]. The predominant effect of the timing of drought induced growth cessation on annual stem growth is not limited to Mediterranean ecosystems, as a similar effect has been observed in mesic temperate forests [START_REF] Brzostek | Chronic water stress reduces tree growth and the carbon sink of deciduous hardwood forests[END_REF]Delpierre et al., 2016a). In our study, we defined the start of the dry season as a critical threshold of predawn plant water potential of -1.1 MPa. This threshold is biologically meaningful because water potential affects the cell turgor pressure necessary for cell growth [START_REF] Hsiao | Sensitivity of growth of roots versus leaves to water stress: biophysical analysis and relation to water transport[END_REF], and it has the other advantage of comprehensively accounting for climate and local soil conditions in a single metric [START_REF] Ruffault | Differential regional responses in drought length, intensity and timing to recent climate changes in a Mediterranean forested ecosystem[END_REF]. It is, however, a more complex index to calculate than climatic indices of drought as it requires substantial knowledge of soil and vegetation characteristics, and accurate meteorological data at the daily time scale. Consequently, the uncertainty on t1 was higher than the uncertainty on t0, especially when meteorological data from SML (located at a distance of 12 km from the study site) was used instead of on-site data, but t1 nonetheless remained a good predictor of annual stem growth for whichever period (Table 2).

Depending on the species and on the local bio-climate, a minimum temperature threshold ranging from +4 °C to +7 °C is required for stem growth to occur [START_REF] Körner | Carbon limitation in trees[END_REF][START_REF] Rossi | Evidence of threshold temperatures for xylogenesis in conifers at high altitudes[END_REF][START_REF] Deslauriers | Cambial phenology, wood formation and temperature thresholds in two contrasting years at high altitude in southern Italy[END_REF][START_REF] Gruber | Impact of drought on the temporal dynamics of wood formation in Pinus sylvestris[END_REF][START_REF] Swidrak | Effects of environmental conditions on onset of xylem growth in Pinus sylvestris under drought[END_REF][START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF]. The onset of stem growth can be assessed from winter temperature (Delpierre et al., 2016b), thus the mean winter temperature directly impacts the duration of cambium activity and wood formation [START_REF] Rossi | Predicting xylem phenology in black spruce under climate warming[END_REF]. However, there is no consensus in dendrochronological studies of the Mediterranean Q. ilex that winter temperature is a good predictor of annual BAI, and both negative [START_REF] Zhang | Variations de la croissance radiale de Quercus ilex L en fonction du climat[END_REF]Paton et al., 2009;[START_REF] Gea-Izquierdo | Climate-growth variability in Quercus ilex L. west Iberian open woodlands of different stand density[END_REF] and positive correlations [START_REF] Campelo | Dendrochronology of Quercus ilex L. and its potential use for climate reconstruction in the Mediterranean region[END_REF][START_REF] Nijland | Relating ring width of Mediterranean evergreen species to seasonal and annual variations of precipitation and temperature[END_REF][START_REF] Gea-Izquierdo | Tree-rings reflect the impact of climate change on Quercus ilex L. along a temperature gradient in Spain over the last 100years[END_REF] between annual stem growth and winter temperature have been reported. In our study, a positive correlation between growth and winter temperature was observed over the period 1984-2008 (r = 0.48, P<0.001; Table 3), in accordance with [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF], but the relationship was not significant for the period 1968-2008, either with t0 or with TJFM.

Taken together, these results suggest that summer drought is the main limiting factor for Q. ilex growth, which is to be expected under the dry Mediterranean climate. Nevertheless, the contrasted growth responses to winter temperature depending on the study concerned, or on the period of interest, led us to investigate the impact of recent climate change on Q. ilex growth in more detail.

Can warmer winter temperature compensate for earlier summer drought under climate change?

From 1968 to 2008, the annual and spring amounts of precipitation were stable (Fig. 4b), but water limitation increased (Fig. 4d) due to increasing potential evapotranspiration with rising temperature in spring and summer (Fig. 4a andc), in accordance with regional observations [START_REF] Ruffault | Differential regional responses in drought length, intensity and timing to recent climate changes in a Mediterranean forested ecosystem[END_REF]. Consequently, we simulated an earlier occurrence of drought onset (t1, -26 days on average) along with an earlier growth onset, although to a lesser extent (t0, -10 days; Fig. 5), which, taken together resulted in a non-significant decrease in growth duration. The positive effect of warming on ecosystem functioning and tree growth through a longer growing season has been widely observed in temperature limited boreal and temperate forests [START_REF] Keeling | Increased activity of northern vegetation inferred from atmospheric CO2 measurements[END_REF][START_REF] Menzel | European phenological response to climate change matches the warming pattern[END_REF][START_REF] D'arrigo | On the 'divergence problem' in northern forests: A review of the tree-ring evidence and possible causes[END_REF][START_REF] Dragoni | Evidence of increased net ecosystem productivity associated with a longer vegetated season in a deciduous forest in south-central Indiana, USA[END_REF]. However, in water limited regions, like the Mediterranean, warming is generally considered to be an aggravating factor for drought, mainly because of increased evaporative demand [START_REF] Angert | Drier summers cancel out the CO2 uptake enhancement induced by warmer springs[END_REF][START_REF] Zhao | Drought-induced reduction in global terrestrial net primary production from 2000 through[END_REF][START_REF] Williams | Temperature as a potent driver of regional forest drought stress and tree mortality[END_REF]. Consequently, climate warming in the Mediterranean usually reduces tree growth [START_REF] Jump | Rapid climate change-related growth decline at the southern range edge of Fagus sylvatica[END_REF][START_REF] Sarris | Recent decline in precipitation and tree growth in the eastern Mediterranean[END_REF][START_REF] Peñuelas | Twentieth century changes of tree-ring δ13C at the southern range-edge of Fagus sylvatica: increasing water-use efficiency does not avoid the growth decline induced by warming at low altitudes[END_REF][START_REF] Piovesan | Drought-driven growth reduction in old beech (Fagus sylvatica L.) forests of the central Apennines, Italy[END_REF]Martin-Benito et al., 2010). Our results thus mitigate this widely accepted conclusion and illustrate the peculiar parallel controls driven by temperature, such that the benefit of an earlier stem growth is cancelled out by earlier drought onset mostly caused by increasing evaporative demand.

The effect of temperature on t1, mediated by PET, may also explain why the correlation with growth was better for t1 than for PAMJ over the period 1984-2008, when temperatures increased significantly, but not over the longer period 1968-2008. Actually, spring PET, like temperature, did not increase significantly until the early 1980s (Fig. 4a,c), suggesting that, in the past, inter-annual variability in precipitation may have been a stronger driver of drought onset. Alternatively, the lack of on-site precipitation measurements before 1984 may mean that t1 estimates based on SML are less closely linked to on-site conditions than three-month cumulated precipitation. The correlation in precipitation amounts between Puéchabon and SML actually increases with longer temporal resolutions.

Winter warming may also have a positive impact on stem growth by delaying growth cessation in autumn, thereby partly compensating for earlier drought onset. However, the phenology of autumn growth cessation appears to be less variable than that of t0 [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF], possibly because it is concurrently driven by photoperiod. Moreover, warm winters may even impact Q. ilex growth negatively if the species requires winter chilling, as is the case of the deciduous Quercus species [START_REF] Fu | Declining global warming effects on the phenology of spring leaf unfolding[END_REF].

A Mediterranean "divergence problem"

When looking at the temporal variations in the correlations between stem growth and climate variables, we observed an abrupt and significant increase in the sensitivity of stem growth to temperature in the early 2000s, while at the same time, its response to precipitation weakened (Fig. 6a,b). Temporal changes in the response of tree growth to climate have been observed in a wide range of climates and tree species in recent decades (e.g. [START_REF] Briffa | Reduced sensitivity of recent tree-growth to temperature at high northern latitudes[END_REF][START_REF] Büntgen | Summer temperature variations in the European Alps, a.d. 755-2004[END_REF][START_REF] Carrer | Long-term change in the sensitivity of tree-ring growth to climate forcing in Larix decidua[END_REF][START_REF] Jump | Climate relationships of growth and establishment across the altitudinal range of Fagus sylvatica in the Montseny Mountains, northeast Spain[END_REF][START_REF] D'arrigo | On the 'divergence problem' in northern forests: A review of the tree-ring evidence and possible causes[END_REF]. The 'divergence problem' in northern forests has been defined as the tendency for tree growth at previously temperature limited sites to undergo a weakening of their temperature response concurrent with an increasing sensitivity to drought [START_REF] D'arrigo | On the 'divergence problem' in northern forests: A review of the tree-ring evidence and possible causes[END_REF]. Our results suggest a Mediterranean 'divergence problem' according to which tree growth in water limited Mediterranean ecosystems undergo a weakening of their response to spring-summer precipitation and an increasing sensitivity to winter temperatures. Similar reports of temporal changes from water driven to increasingly temperature driven tree growth in water limited ecosystems have already been reported for beech forests in northeast Spain [START_REF] Jump | Climate relationships of growth and establishment across the altitudinal range of Fagus sylvatica in the Montseny Mountains, northeast Spain[END_REF], black pine forests in Spain (Martin-Benito et al., 2010), and Scots pine, European larch and black pine in Switzerland [START_REF] Feichtinger | Growth adjustments of conifers to drought and to century-long irrigation[END_REF]. These observations differ from ours by reporting an increase of the overall negative effect of warming on tree growth. Nevertheless, they collectively point to the increasing influence of temperature on the growth of previously water limited trees.

Concurrent increase in drought and warming: a keystone aspect of climate change revealed during the recent warming hiatus

A keystone result of our study is the sudden significance of the growth-temperature relationship occurring from the 1995-2004 time window on (Fig. 6b). In parallel, we observed lower winter temperatures than would have been expected on the basis of continuous climate warming (Fig. 4a). This regional pattern is therefore comparable to the globally observed "warming hiatus" [START_REF] Easterling | Is the climate warming or cooling?[END_REF][START_REF] Trenberth | An apparent hiatus in global warming?[END_REF]. Our results suggest that these recent cooler winter temperatures resulted in later growth onset and led to an increased temperature constraint on the duration of stem growth. In the meantime, the constraint exerted by the water deficit increased constantly from 1968 onward with neither breakpoints nor changes in the trend line (Fig. 4d,5). As a result, the stem growth of Q. ilex was significantly correlated only with water deficit in the past, but the shorter growth period after the 2000s revealed the dual control by winter temperatures and spring-summer water deficit. The pause in climate warming in turn disrupted the precarious balance between increasing winter temperatures and increasing spring-summer water deficit, which temporarily sustained stem growth until the end of the 1990s (Fig. 7). This is, to our knowledge, the first example of a 'divergence problem' in the tree growth-climate relationship triggered not by a continuous climate change but instead by the warming pause. This recent warming pause, a still controversial aspect of climate change [START_REF] Lewandowsky | On the definition and identfiability of the alleged "hiatus" in global warming[END_REF][START_REF] Wehner | The global warming hiatus's irrelevance[END_REF], is mainly caused by the variations in Atlantic and Pacific multidecadal oscillations [START_REF] Steinman | Atlantic and Pacific multidecadal oscillations and Northern hemisphere temperatures[END_REF], and produced globally heterogeneous patterns of breakpoint in the recent warming trend [START_REF] Ying | The recent hiatus in global warming of the land surface: Scale dependent breakpoint occurrences in space and time[END_REF], with an enhanced effect on winter temperature in Eurasia [START_REF] Li | Eurasian winter cooling in the warming hiatus of 1998-2012[END_REF]. It is however, likely to be reversed in the coming decades [START_REF] Steinman | Atlantic and Pacific multidecadal oscillations and Northern hemisphere temperatures[END_REF]. [START_REF] Gao | Increased aridity in the Mediterranean region under greenhouse gas forcing estimated from high resolution simulations with a regional climate model[END_REF][START_REF] Ruffault | Projecting future drought in Mediterranean forests: bias correction of climate models matters![END_REF], which are likely to move the onset of drought forward more strongly than the onset of spring growth [START_REF] Lempereur | Growth duration is a better predictor of stem increment than carbon supply in a mediterranean oak forest: implications for assessing forest productivity under climate change[END_REF]. Together with the current growth limiting winter temperatures, these future trends in drought features could lead to a sharp reduction in forest productivity and an increase in tree mortality in Mediterranean Q. ilex forests. Table S4 Breakpoints and changes in mean winter temperature for seven meteorological stations in the region Languedoc-Roussillon.

Fig. 1

 1 Fig. 1 Chronological timeline of the main data sources used in this study. Stem growth series: automatic dendrometers (AD), field survey (FS) of stem diameter at breast height (DBH) and ring width series (RW).Climatic data: Puéchabon meteorological station and St Martin-de-Londres (SML) meteorological station.

  not clearly visible after 40 years of cambial age; consequently only 12 of the sampled stems could be cross dated with confidence, seven stems in 2005 and five stems in 2008. The first ring was formed in 1942 after the clearcut in winter 1941-1942, so data in the ring width series

  , respectively). The minimum value of BAI was recorded in 2006 for both series: BAIRW = 114 mm² y -1 (SE = 36.5 mm² y -1 ) and BAIFS = 58 mm² y -1 (SE = 13.2 mm² y - 1 ). The maximum BAI was recorded in 1977 for RW (BAIRW = 348 mm² y -1 , SE = 22.1 mm² y - 1 ) and in 2001 for FS (BAIFS = 246 mm² y -1 ; SE = 20.8 mm² y -1 , vs. BAIRW = 213 mm² y -1 ; SE = 24 mm² y -1 .2).

Fig. 3

 3 Fig. 3 Relationship between yearly basal area increment (BAI) and the duration of the spring growth period (∆tt0-t1). Ring width (RW, dark grey circle) is shown for the period 1984-2008 and field survey (FS, light grey square) for the period 1986-2013. The linear relationships between ∆tt0-t1 and BAIRW (dark grey line; BAIRW = 1.34*∆tt0-t1 + 122.9; R² = 0.56; P-value < 0.001) or BAIFS (light grey line; BAIFS = 1.09*∆tt0-t1 + 95.5; R² = 0.35; Pvalue < 0.01) are represented. Error bars are standard errors.

Fig. 4

 4 Fig. 4 Temporal trends in (a) spring (AMJ) temperature (grey circles) and winter (JFM) temperature (light grey inverted triangles), (b) mean annual precipitation (dark grey triangles) and AMJ precipitation (grey circles), (c) mean annual potential evapotranspiration (ETP; dark grey triangles) and AMJ ETP (grey circles) and (d) annual water stress integral (WSI, dark grey triangles) and AMJ three months WSI (grey circles). The solid lines represent significant temporal trends (P-value<0.05). The dashed lines represent the extended trend for temperature after 1998, or before 1978 for PET. The effect of the warming pause on the JFM temperature (panel a) is shown by the hatched area between the points and the regression line, the vertical dotted lines indicate the start of the winter warming pause in 2002.

Fig. 5

 5 Fig.5Onset of phenological stem growth (t0, pale grey inverted triangles) and end of stem growth (t1, grey circles) from 1966 to 2013, expressed as day of the year (DOY).The effect of the warming pause on t0 is shown by the hatched area between the points and the regression line over the 1978-1998 period.

Fig. 6 Fig. 7

 67 Fig. 6 Temporal variations in the bootstrapped Pearson's correlation coefficients between (a) BAIRW and PAMJ, and (b) BAIRW and TJFM over 10-year moving windows. The black curve represents the correlation coefficient and the grey curves the upper and lower confidence intervals at a significance level α = 0.05. Asterisks identify the time windows with significant correlations.

  Whether a future re-acceleration of climate warming in the Mediterranean would compensate for increasing water deficit by phenological stimulation of earlier tree growth again, or on the contrary, further exacerbate drought stress remains an open and important question. Studying trends in targeted and concurrent climate variables such as temperature and drought should fully capture the complexity of climate change impacts (Mazdiyani & Aghakouchak 2015). Climate projections for the Mediterranean region forecast an increase in potential evapotranspiration and a decrease in summer precipitation by the end of the 21 th century

Fig. S8

 S8 Fig. S8 Temporal trends in winter temperature for seven meteorological stations in the region Languedoc-Roussillon.

Table 1

 1 Main characteristics of the three growth series used in the study, automatic dendrometer (AD), field survey

	(FS) and ring width (RW), over their respective complete temporal series and their overlap periods. Coefficients
	of correlation (r) between FS and AD over the period 2004-2013 and between RW and FS over the period 1987-
	2008 are given (P-value < 0.05: *; P-value < 0.001: ***).

Characteristics of the growth series Method Period Mean BAI (mm²) DBH 2008 (cm) Sample size AD

		Auto Dendro	2004-2013	185.8(±70,00)	11.1 (±2.45)	12
	FS	Diameter tape	1987-2013	154.4 (±44,47)	10.5 (±1.85)	125
	RW	Ring width	1968-2008	204.6 (±55,39)	12.7 (±1.75)	12

Correlations on overlap periods Ovelap period Growth series Mean BAI (mm²) r AD vs. FS 2004

	-2013	AD FS	185.8(±70.00) 142.3 (±55.37)	0.90 (***)
	FS vs. RW 1987-2008	FS RW	158.2 (±44.68) 196.2(±45.00)	0.55 (*)

Table 2

 2 Mean Pearson's correlation coefficients (and significance derived from a 1000 classical bootstrap sampling) between chronologies of annual stem growth (BAI) and the main phenology explanatory variables. The

	correlations are given using climate variables measured in Puéchabon field surveys (FS) over the period 1986-
	2013 and ring width (RW) over the period 1984-2008, and climate data from the SML meteorological station over
	the period 1968-2008 for RW (see Fig. 1). The explanatory variables tested were the start and stop growth dates
	(t0 and t1, respectively) and the duration of stem growth (∆tt0-t1). The coefficient of correlation (r) and the level of
	statistical significance (*P-value < 0.05; **P-value < 0.01; ***P-value < 0.001) are given. Significant correlations
	are in bold.
		FS	RW	RW
	Period	1986-2013	1984-2008	1968-2008
	t0	-0.43**	-0.51***	0.18
	t1	0.56***	0.68***	0.66***
	∆tt0-t1	0.60***	0.73***	0.56***

Table 3

 3 Mean Pearson's correlation coefficients and significance (derived from a 1000 classical bootstrap sampling) between annual stem growth (BAI) or spring growth duration (Δtt0t1) and monthly or seasonal precipitation (P, in mm) and temperature (T, in °C) for RW data series. The climate data from Puéchabon and

	Saint Martin-de-Londres meteorological stations were used for the periods 1984-2013 and 1966-2013,
	respectively. The coefficients of correlation (r) and the level of statistical significance (*P-value < 0.05; **P-
	value < 0.01; ***P-value < 0.001) are given. Significant correlations are in bold.
				BAI				Δtt0t1
		1984-2008	1968-2008	1984-2013	1966-2013
		T	P	T	P	T	P	T	P
	November (t-1)	0.06	-0.08	-0.23*	-0.03	0.07	0.24	-0.31*	-0.03
	December (t-1)	0.30**	0.08	-0.07	-0.04	0.25	0.04	-0.06	-0.03
	January	0.50*** -0.06	0.02	0.15	0.51*** 0.03	0.34*** 0.14
	February	0.37*	-0.27*	-0.02	0.06	0.20	-0.16	0.22	0.02
	March	0.34**	-0.17	-0.10	0.21	0.41*** -0.10	0.11	0.19*
	April	0.05	0.23	0.00	0.33*	-0.11	0.48***	-0.07	0.24*
	May	0.12	0.47*** -0.23	0.57***	0.08	0.37	-0.25*	0.48**
	June	-0.46** 0.21	-0.65*** 0.44***	-0.01	0.00	-0.24*	0.22
	July	-0.56*** 0.32**	-0.54*** 0.36**	-0.26	-0.13	-0.42*** 0.01
	August	-0.08	-0.04	-0.30** 0.07	0.03	0.24*	-0.37*** 0.08
	September	-0.39*	-0.39*	-0.42*** -0.25*	-0.18	-0.35	-0.34*** -0.10
	October	-0.13	-0.13	-0.14	-0.04	-0.18	-0.34**	-0.22	-0.15
	Jan. Feb. March	0.48*** -0.20	-0.05	0.18	0.47*	-0.11	0.32**	0.15
	Apr. May June	-0.15	0.53*** -0.38*** 0.75***	-0.02	0.49*	-0.23	0.55***
	June Aug. Sept.	-0.42** -0.34	-0.51*	-0.14	-0.18	-0.30	-0.45*** -0.05
	from Nov(t-1) to Oct	0.04	-0.11	-0.37*	0.25*	0.15	0.01	-0.21*	0.17

ACKNOWLEDGEMENTS

M. Lempereur benefited from a doctoral research grant provided by the French Environment and Energy Management Agency (ADEME). Long term meteorological data were provided by Météo France. The Puéchabon experimental site belongs to the SOERE F-ORE-T, which is supported annually by Ecofor, Allenvi and the French national research infrastructure ANAEE-F. The authors would like to thank Alain Rocheteau, Karim Piquemal and David Degueldre for their technical assistance, and the two anonymous reviewers for their detailed and helpful reviews of the manuscript.

SUPPORTING INFORMATION

Table S3 Results of the Theil-Sen tests for the trends of temperature, precipitation, evapotranspiration and drought index.


Nitin S Dange

Frédéric Robert

Yannick Landais

Free-radical Carbo-cyanation of Cyclopropenes. A Stereocontrolled Access to All-Carbon Quaternary Stereocenters in Acyclic Systems

t-BuO

Sn-Sn base

ABSTRACT:

The free-radical carbo-cyanation of cyclopropenes offers straightforward access to tetrasubstituted cyclopropanes in satisfying yields and moderate diastereoselectivity. The incorporation of various functional groups on the cyclopropane ring, allows a subsequent base-mediated ring-opening, leading to functionalized acyclic systems having an all-carbon quaternary stereocenter.

Polysubstituted cyclopropanes are useful intermediates in organic synthesis, 1 which have attracted a lot of interest due mainly to their easy ring opening under acidic-, basic or freeradical conditions. 2 Cyclopropanes are readily available in enantiopure form using now well-established organometallic and organocatalyzed processes. 3 Recent progress in the stereocontrolled synthesis of cyclopropanes should also include the functionalization of substituted cyclopropenes. 4 Organometallic (1) and free-radical additions (2) onto the π-system of cyclopropenes thus offer a complementary route to polysubstituted cyclopropanes, which may be functionalized further (Figure 1). It is worth of note that cyclopropenes may also be prepared in enantiopure form through rhodium-mediated cyclopropanation of the corresponding alkynes, using commercially available diazo esters. 5 Recent work by Marek et al. 6 has elegantly shown that syn-carbometallation of cyclopropenes (Figure 1, eq 1) led to the corresponding metallated cyclopropane I, which could be trapped by various electrophiles or undergo selective ring opening. Free-radical atom-transfer processes using cyclopropenes have been reported earlier by Saicic, 7a Zard, 7b et al. and more recently by Miyata, Ueda et al. 8 leading to cyclopropanes in moderate to good yields. In contrast, radical addition of two carbon fragments across cyclopropene π-system has, to our knowledge, not been explored so far. This may be attributed to the high reactivity of the cyclopropyl radical II, which is involved in fast atom-transfer processes (ATRA product IV), [START_REF]For instance BrCCl3 bromine atom transfer to a cyclopropyl radical was estimated to occur with a rate constant of 2[END_REF] preventing further reaction of II with other radical traps. Steric hindrance in the neighborhood of the radical center may also preclude any approach of reagents onto II. Finally, the weaker nucleophilicity of the cyclopropyl σ-radical as compared to that of other alkyl radicals is another factor, which may hamper carbo-functionalization of cyclopropenes. 10 We report here on a new multicomponent carbo-cyanation addition onto substituted cyclopropenes, which gives rise to tetrasubstituted cyclopropyl nitriles III, bearing a quaternary center, in satisfying yields and moderate diastereocontrol. The structure of these cyclopropanes also allow further base-mediated ring opening, offering a straightforward and stereodivergent access to acyclic systems having functionalized all-carbon quaternary stereocenters, present in many biomolecules and natural products. Preliminary experiments were carried out using iodoester 1a as a radical precursor, cyclopropene 2a and commercially available p-tosyl cyanide 3 as a cyanide source. Initiation of the process was performed using di-t-butoxyhyponitrite (DTBHN) and (Bu3Sn)2 as a chain carrier. 11 First reaction attempts by mixing the three components, DTBHN and ditin, produced less than 20% of the desired nitrile as a mixture of two stereoisomers 4aa and 4ab, in the presence of a large amount of the iodine atom transfer product (not shown). Both diastereomers were easily separated through chromatography and their structure assigned through 1 H NMR. Slightly better yield was obtained by adding slowly to the medium, a mixture of p-TsCN and DTBHN through a syringe pump (Table 1, entry 1). Considering that the atom transfer product (ATRA) (i.e. IV, X = I, Figure 1) was likely the intermediate in the reaction process, it was envisioned that the Bu3Sn radical might be sterically too hindered to abstract the iodine atom and regenerate the cyclopropyl radical II for further cyanation. This proved to be the case, as reaction using instead the smaller Me3Sn radical led to much improved yield, cyclopropyl nitriles 4aa-4ab being formed in 60% yield, albeit with modest diastereocontrol (Table 1, entry 2). Reducing the rate of addition had surprisingly a deleterious effect (Table 1, entry 3). The best yield was observed when DTBHN and p-TsCN were added at a rate of 1 ml per hour (Table 1, entry 4). Variation in the nature of the solvent using DCE or CH3CN led to a reduced yield as compared to benzene or no conversion (Table 1, entries 5-6). Finally, an attempt at lowering the reaction temperature to 45 °C, instead of 65 °C, to slow down the decomposition of DTBHN, also led to a lower yield ( a Reaction was carried out mixing 1a (1 equiv), 2a (2 equiv) and (Me3Sn)2 (unless otherwise indicated) (2 equiv) in the solvent, then a mixture of DTBHN (40 mol %) and 3 (2 equiv) was added at 65 °C through a syringe pump. b Isolated yields. c Isolated ATRA product (~ 10-15%). d Estimated through 1 H NMR of the crude reaction mixture after a short filtration column. d (Bu3Sn)2 was used. e Reaction performed at 45 °C.

R 1 R 2 R [M] R 1 R 2 R [M] R 1 R 2 R E E + (1) [M]: MgBr (2) R 1 R 2 R R 1 R 2 R R 1 R 2 R R 1 R 2 R R X
Optimized conditions above (Table 1, entry 4) were then applied to a series of cyclopropenes 2a-i, leading to cyclopropyl nitriles 4a-l in moderate to good yields and modest diastereocontrol (Scheme 1). Variation in yields depending on the nature of the substituents on the cyclopropane ring suggests that steric hindrance is the main issue this process. For instance, variation of the size of the added ester group (i.e. 1a-c) as in 4a-c led to decreased yield going from CO2Et to CO2Ph. Similarly, increasing bulk of the resident R 2 group in cyclopropene as in 4j-k also led to lower yields and more importantly to a loss of diastereocontrol. In all cases, a small amount of the ATRA product was also isolated (from 10 to 15% yield). This approach allows an access to orthogonal carbonyl functions as in 4b-e, which will be useful for further functionalizations. Finally, it is worth noticing that, although diastereocontrol is modest, diastereoisomers of 4b-l are easily separable through silica gel chromatography. Scheme 1. Scope of the carbo-cyanation of cyclopropenes 2ai. The structure of the two isomers of cyclopropanes 4a-l, assigned through 1 H NMR, showed that the addition of the radical precursor occurred anti relative to the cyclopropene ester group, leading to a pyramidalized cyclopropyl σ-radical, known to rapidly invert (10 8 s -1 ). 10 The latter then abstracts the iodine atom of the precursor 1a-d, providing the corresponding ATRA product (i.e. IV, Figure 1) as an intermediate (vide infra). Reaction of this iodide with the trimethyltin radical then returns the cyclopropyl σ-radical (i.e. II, Figure 1), which may react under two configurations A and B, the p-tosyl cyanide 3 preferably approaching anti relative to the resident ester group in configuration A (Figure 2). 8a Configuration B is less favored due to steric interactions between the R group and the CH2CO2Et chain. When R substituent becomes too bulky as in 4j-k, steric interactions with CO2Et develop and configurations A and B likely have similar energies, resulting in a loss of diastereocontrol. Interestingly, cyanation of diastereomerically pure iodocyclopropane 5 [START_REF]Iodocyclopropane 5 was obtained in 87% yield (dr 88:12) through addition of α-iodoester 1[END_REF] with p-tosyl cyanide 3 under the above conditions led to the cyclopropane 4a with the same diastereomeric ratio observed in the above one-pot reaction, indicating that the inversion at the radical center is faster than the cyanation, in good agreement with earlier studies (Scheme 2). 10 Scheme 2. Cyanation of iodocyclopropane 5. The high reactivity of the cyclopropyl radical was further illustrated during carbo-cyanation of cyclopropene 6. Treatment of the latter under the above carbo-cyanation conditions led to the desired cyclopropyl nitrile 7 in low yield along with nitrile 8 (as a mixture of two diastereomers), resulting from a 1,5-hydrogen shift (Scheme 3). This unexpected C-H activation is driven by radical stability, secondary radicals being much more stable than cyclopropyl ones, as suggested by radical stabilization enthalpies (RSE), [START_REF] Hioe | Encyclopedia of Radicals in Chemistry, Biology and Materials[END_REF] measured for cyclopropyl and isobutyl radicals.

CO 2 R 1 CO 2 R 1 R 2 NC EWG I (major) R 2 p-
Scheme 3. 1,5-Hydrogen atom abstraction in carbo-cyanation processes. With cyclopropyl nitriles 4 in hand, the base-mediated ringopening process was then studied. 14 Preliminary attempts using LDA led to low conversion and yield. In contrast, soft enolization of the cyclopropanes 4b, 4f and 4h-i using i-Pr2NEt (DIPEA) and MgBr2 in CH2Cl2 was found to be more appropriate, 15 leading to a clean cyclopropane ring-opening and the formation of highly functionalized acyclic systems 9a-d bearing an all-carbon quaternary center in generally good yields (Scheme 4). 16 Reaction were relatively fast, except with 9b where starting material could not be totally consumed even after 12h. Scheme 4. Base-mediated ring-opening of cyclopropanes 4b, 4f and 4h-i. The carbo-cyanation/cyclopropane ring-opening sequence was finally applied to enantioenriched cyclopropene 2a (er 94:6), readily available through cyclopropanation of pent-1yne using Corey's chiral rhodium catalyst (supporting information). 5c The carbo-cyanation of enantioenriched 2a led to a 7:3 mixture of diastereomers 4aa and 4ab, which were separated through silica gel chromatography (Scheme 5). Separate treatment of both isomers under the soft enolization conditions above, finally led to enantiomers 10a and 10b, the enantiomeric purity of which was shown (chiral HPLC) to be the same to that of the starting cyclopropene 2a, indicating that no erosion of the enantioselectivity occurred during the whole sequence. This straightforward transformation thus provides a stereodivergent access to both enantiomers of diester 10 having all-carbon quaternary stereocenters. 16 In summary, we reported the first free-radical carbo-functionalization of cyclopropenes. This one-pot multicomponent carbo-cyanation offers a straightforward entry to tetrasubstituted cyclopropanes in satisfying yields and moderate diastereoselectivity. This radical addition installs functionalizations allowing further base-mediated cyclopropane ring-opening, leading to acyclic diesters having an all-carbon quaternary stereocenter. This methodology makes use of readily available enantioenriched cyclopropenes and leads to regioselectivity, which complements those observed in carbo-metallation of cyclopropenes. 4,6

CO 2 Et R NC COR' MgBr2 .OEt 2 (3 equiv) DIPEA (3.5 equiv) EtO 2 C NC R 9a (71%) 9c (67%) 9d (

Figure 1 .

 1 Figure 1. Carbometallation and free-radical additions onto cyclopropenes.

Figure 2 .

 2 Figure 2. Transition state model for the carbo-cyanation of cyclopropenes.

Scheme 5 .

 5 Scheme 5. Stereodivergent access to diesters 10a-b having an all-carbon quaternary stereocenter.

Table 1

 1

	,

Table 1 .

 1 Optimization of the carbo-cyanation of cyclopropene.

							CO 2 Et
	EtO 2 C 1a	I	+	CO 2 Et 2a	p-TsCN, DTBHN (R3 Sn) 2 3 solvent, 65 °C	+	NC 4aa CO 2 Et (major) CO 2 Et
							NC	CO 2 Et
							4ab	(minor)
	entry		rate of addition (time)	solvent	yield b,c (%)	dr d
	1 a		0.5 mL/h (5 h)	benzene	24 d	n.d.
	2		0.5 mL/h (5 h)	benzene	60	72:28
	3		0.25 mL/h (10 h) benzene	16	n.d.
	4		1 mL/h (2.5 h)	benzene	64	76:24
	5		1 mL/h (2.5 h)	DCE	33	78:22
	6		1 mL/h (2.5 h)	CH3CN		-	-
	7 e		1 mL/h (2.5 h)	benzene	41	77:23

ACKNOWLEDGMENT

NSD thanks EU for a postdoctoral grant through H2020 Marie-Curie IF program (Leucophyl, N°655527). The University of Bordeaux and CNRS are gratefully acknowledged for financial support. Pierre Bouillac (University of Bordeaux) is thanked for preliminary experiments.

AUTHOR INFORMATION

Corresponding Author * y.landais@ism.u-bordeaux1.fr


Nitin S Dange

Ashique Hussain Jatoi

Frédéric Robert

Yannick Landais

Visible-Light Mediated Addition of Phenacyl Bromides onto Cyclopropenes

CO 2 Et R 2 O + base (2 mol %) R 1 (er = 95:5) (er = 95:5) h υ (455 nm)

ABSTRACT: Visible-light promoted addition of α-bromoacetophenones onto the cyclopropene π-system in the presence of the fac-Ir(ppy)3 catalyst was shown to afford the corresponding 1(4H)-naphthalenones. The syn-carboarylation of the cyclopropene is followed by a cyclopropane ring opening under the basic conditions, allowing the formation of two C-C bonds and the generation of 1(4H)-naphthalenones bearing an all-carbon benzylic quaternary stereocenter.

Cyclopropenes exhibit remarkable reactivity and behave both as electrophilic and nucleophilic species. 1 The high acidity of the olefinic protons, due to ring strain, allows their easy deprotonation and further functionalization with electrophiles, while their electrophilicity is revealed during addition of organometallic species, including hydrides, Grignard reagents, organocuprates, zinc and indium reagents. Cyclopropenes may be accessed by a number of methods and are readily available in enantiopure form through metal-catalyzed cyclopropanation of the corresponding alkynes. 2 These small ring systems thus continue to attract a great deal of interest, but surprisingly, few studies have been reported on their free-radical functionalization. Nakamura et al. first reported cyclopropene radical hydrostannation in 1994. 3a Xanthate-but also bromine atom transfer processes were later described respectively by Saicic, 3b Zard, 3c Miyata, e Recent work from our laboratory further enriched this field, with the development of a tinmediated carbocyanation of cyclopropenes. 4 As a continuation of this work, we report here on the photoredox-catalyzed addition 5 of α-bromoacetophenones II onto cyclopropenes I and the subsequent cyclopropane ring-opening, providing a straightforward access to naphthalenones III bearing a benzylic quaternary stereocenter, which stereochemistry is controlled early during the cyclopropene synthesis (Figure 1). Such a cyclopropene carboarylation should open an access to useful synthons for organic synthesis, the skeleton of which is found in natural products such as labdane terpenoid (-)-kujigamberol 6a or the synthetic analgesic (-)-eptazocine. 6b In contrast with our recent studies, 4,7 the carbocyanation of cyclopropene 2a, using α-iodoacetophenone 1a in the presence of p-TsCN, (Me3Sn)2 in benzene at 65 °C and di-t-butylhyponitrite (DTBHN) as an initiator, did not provide the carbocyanation product, but instead the atom-transfer product 3, and unexpectedly the cyclized product 4a in a 1:1 ratio (Scheme 1). Repeating the experiment without p-TsCN led to the same products along with the naphthalenone 5a, which was shown to result from the cyclopropane ring-opening of 4a under the reaction conditions.

CO 2 R 1 R 2 I II III O Br R 3 R 2 R 1 O 2 C O R 3 IrLn LED NMe HO H OH (-)-Eptazocine (-)-Kujigamberol
Scheme 1.

H

Intramolecular carboarylation of olefins has precedent in the literature but to our knowledge has not been reported with cyclopropenes. 8 Therefore, considering the synthetic value of compounds such as 5 (vide supra), we focused our attention on the development of their one-pot preparation from phenacyl halides and cyclopropenes, relying on an environmentally more benign initiation process than the one used above (Scheme 1). Pioneering studies by Stephenson et al. 5b,8a,9 have established that photoredox-catalyzed atom-transfer radical processes using α-haloacetophenones were efficiently performed under mild conditions using Ru or Ir catalysts and simple LED irradiation. Optimization of the reaction was thus carried out using several photocatalysts, varying the nature of the base and solvents (Table 1). α-Bromoacetophenone 1b was used instead of 1a to minimize the atom-transfer reaction. The transformation was first carried out using cheaper Ru(bpy)3Cl2 (2 mol %), Et3N as a base and LiBr 9 as an additive (Table 1, entry 1). This provided the desired naphthalenone 5a, albeit in low yield. Better yield was observed when the reaction was performed in a DMF-H2O mixture (Table 1, entry 2). In the absence of Et3N, 4a was obtained without any trace of 5a, indicating that the base was required for the ring-opening, but not for the photocatalyst turnover (Table 1, entry 3) (vide infra). fac-Ir(ppy)3 was then tested and displayed lower efficiency under these conditions (Table 1, entry 4). Higher amount of 5a was however isolated with this catalyst, when K2CO3 was used as a base (Table 1, entry 5). Interestingly, we noticed that 5a was decomposed upon long exposure to the LED irradiation. Therefore, the carboarylation process was then performed during 12 h under irradiation, at which time 4a was the major product formed. Then the lamp was switch-off and the ring-opening completed by a gentle heating at 60 °C (Table 1, entry 6). Slightly higher yields were also obtained at higher concentration (0.2 M) (Table 1,entry 7). An increase of the concentration (0.4 M) or a decrease (0.05 M) led to lower yields. Catalyst loading was also varied, indicating that 2 mol % was optimal for this reaction (Table 1, entries 8-9). Finally, the reaction was carried out in the absence of light and photocatalyst, leading in both cases to no product, thus demonstrating the crucial and cooperative role of the Ir complex and visible-light irradiation (vide infra).

The scope of the reaction was then established using optimized conditions (Table 1, entry 7) on a series of acetophenones 1b-k and cyclopropenes 2a-j, leading to naphthalenones 5a-t, in moderate yields (Scheme 2). Higher yields are obtained with electron-donating substituents (R 2 ) on the phenacyl moiety, illustrating the influence of electronic effects in the process. For instance, when a p-CN substituent was present on the acetophenone, the corresponding unsaturated ketone 5f was not formed, and large amount of p-cyanoacetophenone was recovered instead, indicating that reduction of the bromide was a competing pathway (vide infra). Ru II : Ru(bpy)3Cl2 (2 mol %). b Isolated yields. c DMF-H2O = 4:1. d 4a was isolated (28%). e Ir: fac-Ir(ppy)3 (2 mol %). f The reaction mixture was stirred 12 h under LED irradiation, then heated 24 h at 60 °C without irradiation. g DMF (0.2 M). h fac-Ir(ppy)3 (1 mol %). i fac-Ir(ppy)3 (4 mol %). j Without LED irradiation. k Without photocatalyst.

With m-substituted acetophenones, a mixture of regioisomers was isolated (5g-h Scheme 2). The size of the substituents on the cyclopropene ring has a slight influence on the process efficiency, with lower yields observed with cyclic substituents (5op Scheme 2). The reaction was compatible with the presence of silyl ethers, esters and halides as shown by the formation of 5i, 5k and 5m. Interestingly, naphthalenone 5q was formed from cyclopropene 2j having a mesylate substituent (R 1 = (CH2)2OMs), which was readily displaced in situ by LiBr. It is currently not established if the conversion of the mesylate into the bromide occurred before or after the photocatalyzed process. The reaction could also be performed with heteroarenes, leading to the desired compounds 5r-t, albeit with modest yields. Finally, it is worth noticing that, although yields are generally moderate, final naphthalenones 5 are easily isolated through silica gel chromatography and obtained in only two synthetic operations from the corresponding commercially available alkynes. Scheme 2. Scope of the intramolecular carboarylation of cyclopropenes 2a-j. The stereochemical outcome of the carboarylation/cyclopropane ring-opening sequence was studied, applying the reaction to enantioenriched cyclopropene 2a*, available through cyclopropanation of pent-1-yne using Corey's chiral rhodium catalyst (Scheme 3). 2c 5a* was obtained with the same enantiomeric excess as that of the starting 2a*, indicating that no erosion of the enantioselectivity occurred during the whole sequence. The naphthalenones obtained through this tandem process are useful precursors of polycyclic substrates, as demonstrated with the conversion of 5m into the corresponding polysubstituted pyrane 6 through a simple treatment under basic conditions (Scheme 4). Carbocycles such as 9 may be reached in a straightforward manner starting from mesyl-cyclopropene 7 and 1b, the conversion of which, under optimized conditions above led to the bromide 8. Conversion of 8 into an iodide and then tin-mediated radical cyclization led to 9 as a single diastereomer. A mechanistic pathway was finally proposed to rationalize the course of the tandem process (Figure 2). This would involve, as a first step, a single electron transfer from the photocatalyst excited state (fac-Ir(ppy)3*) to the bromide 1b, affording the αphenacyl radical i. fac-Ir(ppy)3* exhibits a sufficiently strong reduction potential (E1/2 = -1.73V vs SCE) 12a to convert 1b (-0.49V vs SCE) 12b into a radical-anion and then to i. 8a,b The generation of the latter was shown by a radical trapping experiment using TEMPO, which led to the formation of 10. When the same reaction was performed in the presence of cyclopropene 2a, 10 was also formed in 50% yield. i then adds onto the cyclopropene 2 to generate the highly reactive cyclopropyl radical ii, 13 which reacts intramolecularly with the arene moiety to afford the cyclohexadienyl radical iii. When using α-iodoacetophenone, iodine atom transfer reaction was shown to compete with the intramolecular reaction (Scheme 1). 14 The homolytic aromatic substitution (HAS) is then completed through the oxidation of iii into iv, which rearomatizes through deprotonation under the basic conditions, to provide cyclopropane 4. Oxidation of iii into iv is favored [START_REF] Bhatia | Redox potential for cyclohexadienyl radical is E = -0.1 V (vs SCE)[END_REF] due to the high oxidative power of Ir + (E1/2= + 0.77 V vs SCE), 5a which returns the iridium catalyst in its ground state. Single electron transfer from iii to 1 to generate i and iv is another option to sustain the radical chain (but not the Ir catalytic cycle). 16 4 is finally converted in situ into naphthalenone 5 through deprotonation αto the ketone, followed by cyclopropane-ring opening. 4 This mechanism is consistent with the stereochemistry of 4a, resulting from an addition of radical i anti relative to the ester group as in ii, 4 followed by the reaction of the cyclopropyl radical onto the arene from the same face. This syn addition (imposed by geometrical constrains) of the phenacyl radical across the cyclopropene π-system thus ensures a complete transfer of chirality from the stereocenter C1 to the newly created stereocenters at C2 and C3. Chirality at C3 is thus preserved upon base-promoted cyclopropane ring opening as shown in Scheme 3. In summary, we reported a straightforward synthesis of substituted 1(4)-naphthalenones, bearing a benzylic quaternary stereocenter, through a one pot visible light-mediated addition of αbromoacetophenones to cyclopropenes. Although the overall yield is moderate, this methodology offers an access to useful intermediates and constitutes a new way to generate synthons with quaternary stereocenter, still a challenging task in organic synthesis. 4,17 This methodology uses readily available enantioenriched cyclopropenes as starting material, the stereospecific syn carboarylation of the cyclopropene π-system allowing the formation of two new C-C bonds and ensuring a perfect control of the stereochemistry of the quaternary stereocenter. Application of this methodology to the enantioselective synthesis of bioactive targets such as (-)-eptazocine is now underway and will be reported in due course.

CO 2 Et R 1 1b-k 2a-j Ar O Br CO 2 Et R 1 O 5a-t fac -Ir(ppy)3 O CO 2 Et R 2 O MeO O 5a (44%) O MeO 5b , R 2 = Me (40%) 5c, R 2 = Br (37%) 5d, R 2 = OMe (34%) 5e, R 2 = Cl (31%) 5f, R 2 = CN (0%) O CO 2 Et Cl 5i (
+ O Br O CO 2 R 2 R 1 O R 1 CO 2 R 2 O R 1 CO 2 R 2 H O R 1 CO 2 R 2 H O R 1 CO 2 R 2 H base base h υ

Figure 1 .

 1 Figure 1. Visible-light mediated addition of phenacyl bromides to cyclopropenes.

Scheme 3 .

 3 Scheme 3. I n t ra mo l e c u l a r c a rb o a ry l a t i o n o f e n a n t i o e n ri c h e d c y c l o p ro p e n e 2a*.

Figure 2 .

 2 Figure 2. Proposed mechanism for the radical addition of phenacyl bromides to cyclopropenes.

  Addition of α-iodoacetophenone 1a to cyclopropene 2a.

			O		CO 2 Et
				+
			I
			1a		2a
				DTBHN, (Me3 Sn) 2
					p-TsCN
					benzene, 65	°C
		CO 2 Et		CO 2 Et		CO 2 Et
			+		+
	I	3	COPh	O	4a	O	5a
				3	4a	5a
		with p-TsCN	20%	20%	0%
	without p-TsCN	12%	18%	8%

  10

Table 1 .

 1 Optimization of the carboarylation of cyclopropene 2a.

			O		2a		CO 2 Et
				photocat.
			Br	solvent, 20	°C
				blue LED, N 2	O
			1b				5a
					N		N
		Catalysts:	N	Ru	N	Ir
				N		N	N	N
					N
				Ru(bpy)3 Cl 2	fac -Ir(ppy)3
	entry cat.	additive		solvent	Time (h) yield% b
	1	Ru II a Et3N, LiBr		DMF	72	10
	2	Ru II a Et3N, LiBr DMF-H2O c	72	30
	3	Ru II a	LiBr		DMF-H2O c	72	-d
	4	Ir e	Et3N, LiBr DMF-H2O c	72	15
	5	Ir e	K2CO3		DMF	72	34
	6	Ir e K2CO3,LiBr		DMF	12 f	40
	7	Ir e K2CO3,LiBr		DMF g	12 f	44
	8	Ir h K2CO3,LiBr		DMF g	12 f	22
	9	Ir i K2CO3,LiBr		DMF g	12 f	39
	10	Ir e K2CO3,LiBr		DMF g	120 j	-
	11	-k	K2CO3,LiBr		DMF g	120	-
	a

11

  Scheme 4. Further elaboration of naphthalenones.

	EtO 2 C				MeO 2 C
		OAc	K 2 CO 3	O
				MeOH, 20	°C	H
	O			76%	O
	5m				6
	O
		Br		fac -Ir(ppy)3	EtO 2 C
				blue LED	Br
	1b			K 2 CO 3 , LiBr
	+	CO 2 Et	then 60 °C, 24 h DMF, 20 °C, 12 h	O
	MsO			no LED	8
				32%	1. NaI
	7				70% (2 steps)	acetone 2. Bu 3 SnH, AIBN benzene
					80 °C, 12 h
					EtO 2 C
					H
					O
					9

AUTHOR INFORMATION

Corresponding Author * y.landais@ism.u-bordeaux1.fr ACKNOWLEDGMENT ND thanks EU for a postdoctoral grant through H2020 Marie-Curie IF program (N°655527). AH thank the Shaheed Benazir Bhutto University for a PhD grant. The of Bordeaux and CNRS are gratefully acknowledged for financial support.