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. 2016 Jul 20;172(2):765–775. doi: 10.1104/pp.16.00877

Adaptation of the Long-Lived Monocarpic Perennial Saxifraga longifolia to High Altitude1[OPEN]

Sergi Munné-Bosch 1,2,*, Alba Cotado 1,2, Melanie Morales 1,2, Eva Fleta-Soriano 1,2, Jesús Villellas 1,2,2, Maria B Garcia 1,2
PMCID: PMC5047100  PMID: 27440756

Adaptation of an endemic long-lived monocarpic perennial to high altitude is influenced by multiple mechanisms operating at various levels

Abstract

Global change is exerting a major effect on plant communities, altering their potential capacity for adaptation. Here, we aimed at unveiling mechanisms of adaptation to high altitude in an endemic long-lived monocarpic, Saxifraga longifolia, by combining demographic and physiological approaches. Plants from three altitudes (570, 1100, and 2100 m above sea level [a.s.l.]) were investigated in terms of leaf water and pigment contents, and activation of stress defense mechanisms. The influence of plant size on physiological performance and mortality was also investigated. Levels of photoprotective molecules (α-tocopherol, carotenoids, and anthocyanins) increased in response to high altitude (1100 relative to 570 m a.s.l.), which was paralleled by reduced soil and leaf water contents and increased ABA levels. The more demanding effect of high altitude on photoprotection was, however, partly abolished at very high altitudes (2100 m a.s.l.) due to improved soil water contents, with the exception of α-tocopherol accumulation. α-Tocopherol levels increased progressively at increasing altitudes, which paralleled with reductions in lipid peroxidation, thus suggesting plants from the highest altitude effectively withstood high light stress. Furthermore, mortality of juveniles was highest at the intermediate population, suggesting that drought stress was the main environmental driver of mortality of juveniles in this rocky plant species. Population structure and vital rates in the high population evidenced lower recruitment and mortality in juveniles, activation of clonal growth, and absence of plant size-dependent mortality. We conclude that, despite S. longifolia has evolved complex mechanisms of adaptation to altitude at the cellular, whole-plant and population levels, drought events may drive increased mortality in the framework of global change.

European mountains shelter a huge biodiversity and are home to many endemic plants and animals, i.e. species that occur nowhere else. Global change, and particularly climatic change, is expected to exert a major effect on mountain plant communities, altering their potential capacity for adaptation (Peñuelas and Boada, 2003; Franklin et al., 2016). Under such a scenario of environmental change, populations of organisms must either escape or get quickly adapted, otherwise they go extinct. For instance, certain butterfly species have been migrating north, or to higher altitudes, to escape rising temperatures (Breed et al., 2013). Plants, of course, cannot migrate as fast as animals, and important shifts have already been found among plant communities inhabiting mountain summits (Gottfried et al., 2012). When global air temperatures increase, the number of cooler habitats will shrink, producing a crowding effect and increased competition among some species in the remaining cooler areas; at the same time, however, other habitat types will increase in abundance (Scherrer and Körner, 2011). Alpine habitats could prove more attractive to plant species than lowlands because of their topography providing favorable microhabitats. However, certain rare species may lose out in the long-term competition for space, especially those favoring cooler climates (Körner, 2013).

Leaves of high-mountain plants are highly resistant to photoinhibitory damage. Tocochromanols (particularly tocopherols and plastochromanol-8) are found in thylakoids and play an antioxidant function in protecting lipids from the propagation of lipid peroxidation in chloroplasts. Together with carotenoids, they also prevent PSII damage as a result of singlet oxygen attack (Munné-Bosch and Alegre, 2002; Falk and Munné-Bosch, 2010; Zbierzak et al., 2009; Kruk et al., 2014). A higher tocochromanol content, particularly of α-tocopherol, and an increased capacity for nonradiative dissipation of excitation energy by activation of the xanthophyll cycle have been found in high-mountain plants, thus supporting such a role (Streb et al., 1997, 1998, 2003a, 2003b; García-Plazaola et al., 2015). Furthermore, although the number of studies is still very limited for plants in their natural habitat, high-mountain plants tend to accumulate large amounts of ABA (Bano et al., 2009), a phytohormone that is known to mediate the acclimation/adaptation of plants to temperature extremes by modulating the up- and down-regulation of numerous genes (Gilmour and Thomashow, 1991). The activation of other chemical defenses, such as the accumulation of salicylates and jasmonates, which serve against biotrophs and necrotrophs, can also be affected by extreme temperatures (Kosova et al., 2012; Dong et al., 2014; Miura and Tada, 2014), but it has not been investigated thus far in high-mountain plants.

Some degree of plasticity in physiological traits is ubiquitous among plants so that environmental growth conditions are generally considered essential factors governing the physiological performance of plants and their organs (Larcher, 1994). A number of recent studies with trees, shrubs, and herbs, including vascular epiphytes (Mencuccini and Grace 1996; Zotz, 1997; Schmidt et al., 2001; Schmidt and Zotz, 2001; Munné-Bosch and Lalueza, 2007; Morales et al., 2014), point out, however, to another source of intraspecific variation that many studies in the past have inadvertently missed, i.e. substantial variation in physiological traits related to plant size rather than changing environmental conditions (Zotz et al., 2001). In trees, increased plant size leads to increased hydraulic resistance causing reductions in relative leaf growth rates (Mencuccini and Grace, 1996). Furthermore, photo-oxidative stress has been shown to increase during periods of low precipitation combined with high light in the Mediterranean shrub Cistus clusii as a function of plant size, therefore suggesting an increased vulnerability to photo-oxidative stress in the largest individuals (Munné-Bosch and Lalueza, 2007). To our knowledge, no studies are however available to unveil the possible influence of plant size on photoprotection and activation of chemical defenses in high-mountain plants.

Saxifraga longifolia Lapeyruse (Saxiragaceae) is an endemic species of the western Mediterranean mountains, ranging from the Pyrenees (plus a couple of populations in the Cantabric Mountains) through eastern Spain to reach its southern limit in the high Atlas of Morocco (Webb and Gornall, 1989). This long-lived monocarpic plant develops a basal rosette growing in limestone rocky places, mainly on cliffs, offering a unique sight in years of intensive blooming. Reproduction occurs when plants are at least 6 years old in greenhouse conditions (Webb and Gornall, 1989), and it is thought to be much later in natural conditions. This orophyte plant shows striking variation in plant size, with a diameter of the rosette up to 30 cm in the largest individuals. It has been shown that flower and seed production increase as a function of plant size, with female success being maximum in intermediate sized plants (García, 2003).

In the current study, with the aim of getting new insights into the mechanisms of adaptation to high altitudes and the influence of plant size on this adaptation capacity, we examined the physiological response of S. longifolia growing at three contrasted populations spanning its whole altitudinal range. We described population structure, calculated mortality rates, and analyzed physiological performance, including water contents and activation of photoprotection mechanisms and chemical defenses. We aimed at understanding the effect of varying altitude on the expression of defense mechanisms that govern adaptive processes in high-mountain plants.

RESULTS

Physiological Performance and Mortality in Populations at Various Altitudes

S. longifolia is a monocarpic species; therefore, plants die as a consequence of reproduction. We wondered, however, whether mortality of juveniles is also influenced by stressful conditions across an altitudinal range by monitoring three plant populations growing at 570, 1100, and 2100 m a.s.l. The population occurring at the highest altitude (Las Blancas) had the largest plants, the least frequency of small ones (Supplemental Fig. S1A), and the lowest mortality rate in juveniles (average across 4 years: 4.8%, Table I). Mortality of juveniles was higher in the lowest population (Pantano de la Peña, 6.9%) and highest in the intermediate one (San Juan de la Peña, 11.5%, Table I), both of them showing a similar population size distribution (Supplemental Fig. S1A). Therefore, altitude, which, as expected, resulted in lower temperatures (Supplemental Fig. S2), does not seem to be associated to mortality rates in juveniles. Rather, mortality rates in juveniles seemed to be more associated with reduced soil water contents. Among the three populations studied, the volumetric soil water content was the highest in the high population and lowest in the intermediate one (Supplemental Fig. S2). Plants growing in San Juan de la Peña were exposed to more stressful conditions during the day of measurements compared to the other two populations, as indicated not only by reduced soil water contents, but also higher solar radiation and air temperatures, which may in turn contribute to drought stress. Mortality due to flowering was much lower than among juvenile plants (Table I), and flowering rates were rather stochastic across years and populations, with the highest population showing the higher flowering rates during 2013-14, but also the smaller ones during 2014-15 (Table I).

Table I. Mortality rates (numbers and percentage) during the last 4 years in the long lived monocarpic plant, S. longifolia.

Monitoring of individual plants was carried out between June and August of the given periods (indicated in parentheses). Mortality rates refer to juveniles (Juv) and flowering (R).

Pantano de la Peña San Juan de la Peña Las Blancas
Juv R Juv R Juv R
N (2011-2012) 194 204 281
Dead 6 2 14 12 5 0
% mortality
 3.1
1.0
 6.9
5.9
 1.8
0.0
N (2012-2013) 230 205 285
Dead 33 0 22 0 13 3
% mortality
 14.3
0.0
 10.7
0.0
 4.6
1.1
N (2013-2014) 211 201 276
Dead 11 1 30 0 13 42
% mortality
 5.2
0.5
 14.9
0.0
 4.7
15.2
N (2014-2015) 204 198 217
Dead 10 12 27 10 18 2
% mortality
 4.9
5.9
 13.6
5.1
 8.3
0.9
Average (2011-2015) 6.9 1.8 11.5 2.8 4.8 4.3
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Physiological stress indicators, including leaf water, pigment and lipid peroxidation levels (Fig. 1), antioxidant protection (Fig. 2), and stress-related phytohormones (Fig. 3) revealed that the physiological performance in juvenile plants differed in the three populations, some parameters changing as a function of altitude and others following the mortality rate pattern more linked to drought stress (the intermediate population showed the lowest leaf water contents among the three populations, Fig. 1). Intriguingly, the intermediate population was the one showing the highest pigment levels, including those of chlorophylls (Fig. 1), carotenoids, and anthocyanins (Fig. 2). However, when antioxidants were expressed on a chlorophyll basis, the intermediate population showed the highest carotenoid/chlorophyll ratio but the lowest α-tocopherol/chlorophyll ratios (Supplemental Fig. S3), which was observed together with the lowest chlorophyll a/b ratios (Fig. 1). Furthermore, the intermediate population was the one showing the highest levels of all stress-related phytohormones, including ABA, salicylic acid, and jasmonic acid, with the lowest levels of the jasmonic acid precursor, 12-oxo-phytodienoic acid (Fig. 3), thus confirming that the intermediate population was the one experiencing the highest physiological stress in juvenile plants, which is in agreement with the highest mortality rates observed (Table I).

Figure 1.

Figure 1.

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Plant size (estimated as rosette diameter), relative leaf water content (RWC), leaf mass per area ratio (LMA), chlorophyll (Chl) a+b, Chl a/b ratio, and lipid hydroperoxide levels (an estimation of lipid peroxidation) in plants of the long-lived monocarpic plant, S. longifolia, growing at three altitudes (570, 1100, and 2100 m a.s.l. in Pantano de la Peña, San Juan de la Peña, and Las Blancas, respectively). Data shows the mean ± se of n = 65, 64, and 83 juvenile individuals for the three populations at increasing altitude, respectively. Results of one-way ANOVA are shown in the inlets. Different letters indicate significant differences between populations (P < 0.05) using Bonferroni post hoc tests. NS, Not significant.

Figure 2.

Figure 2.

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Antioxidant protection, including levels of anthocyanins, carotenoids, and α-tocopherol in plants of the long-lived monocarpic plant, S. longifolia growing at three altitudes (570, 1100, and 2100 m a.s.l. in Pantano de la Peña, San Juan de la Peña, and Las Blancas, respectively). Data show the mean ± se of n = 65, 64, and 83 juvenile individuals for the three populations. Results of one-way ANOVA are shown in the inlets. Different letters indicate significant differences between populations (P < 0.05) using Bonferroni post hoc tests.

Figure 3.

Figure 3.

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Endogenous concentrations of stress-related phytohormones, including abscisic acid (ABA), salicylic acid (SA), oxo-phytodienoic acid (OPDA), and jasmonic acid (JA) in plants of the long-lived monocarpic plant, S. longifolia, growing at three altitudes (570, 1100, and 2100 m a.s.l. in Pantano de la Peña, San Juan de la Peña and Las Blancas, respectively). Data shows the mean ± se of n = 65, 64, and 83 juvenile individuals for the three populations at increasing altitude, respectively. Results of one-way ANOVA are shown in the inlets. Different letters indicate significant differences between populations (P < 0.05) using Bonferroni post hoc tests.

Levels of photoprotective molecules (α-tocopherol, carotenoids, and anthocyanins) increased significantly in response to high altitude (1100 relative to 570 m a.s.l., Fig. 2), which was paralleled by reduced leaf water contents (Fig. 1) and increased ABA levels (Fig. 3). The more demanding effect of high altitude on photoprotection was, however, abolished (except for α-tocopherol increases) at very high altitudes (2100 m a.s.l.), these plants showing improved water contents (Fig. 1), and a reduced need for photoprotection (driven by anthocyanins and carotenoids, Fig. 2) and activation of chemical defenses (including the three aforementioned classes of stress-related phytohormones, Fig. 3). α-Tocopherol levels increased (Fig. 2), and lipid hydroperoxide levels (an indicator of lipid peroxidation) decreased (Fig. 1) as a function of altitude. Furthermore, Las Blancas was the population showing the lowest leaf mass per area ratio (Fig. 1), carotenoid/chlorophyll ratio (Supplemental Fig. S3), and levels of ABA and jasmonic acid (Fig. 3).

Size Influences Physiological Performance and Plant Death

Logistic models showed that for juveniles, the effect of individual plant size on mortality was significantly important for the low and intermediate populations, where smaller plants had a higher probability to die than larger ones (estimated β1 parameter: −0.12 and −0.05, respectively; P < 0.001 in both cases; Fig. 4). Mortality rate in the highest population was independent on plant size, with dead plants less concentrated in small plants and more uniformly distributed across total size distribution (Fig. 4; Supplemental Fig. S4).

Figure 4.

Figure 4.

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Logistic mortality regression models for the three populations studied. The “x” axis corresponds to the diameter (measured in mm) of plants in year “t”, and the “y” axis to the recorded fate in year “t+1” (0 = alive, 1 = dead). Dots show individual yearly events (dead or alive) from 2011 till 2015. A total of 824, 786, and 1012 events are plotted in the low, intermediate, and high population, respectively. All dots should fit the “0” or “1” values but were not forced to lie on a line for illustrative purposes.

Correlation analyses revealed that the relative leaf water content varied as a function of plant size in the high population (r = 0.528, P < 0.001), but not in the intermediate and low populations. The larger the plant, the higher the leaf water content in Las Blancas (Table II; Fig. 5), a correlation that was not significant at lower altitudes. In should be noted, however, that the lowest leaf water contents were observed in San Juan de la Peña (with values around 50% lower than in the other two populations), although the large variability observed prevented the correlation to be significant (Fig. 5). Other, more moderate, correlations were observed between plant size and leaf mass area ratio for the intermediate and high populations, and between plant size and the chlorophyll a/b ratio for the low population (r = 0.34–0.35, P < 0.05, Table II).

Table II. Spearman rank's correlation analyses between plant size (estimated as rosette diameter) and all measured parameters in the long-lived monocarpic plant, S. longifolia.

All data from juvenile plants, including the three populations, was pooled for analyses, but also analyzed separately. rho and P values are indicated in bold when correlations were significant (Bonferroni adjusted, P < 0.0033). RWC, relative water content; LMA, leaf mass per area ratio; Chl, chlorophyll. LOOH, lipid hydroperoxides; Ant, anthocyanins; Car, carotenoids; α-Toc, α-tocopherol; ABA, abscisic acid; SA, salicylic acid; OPDA, oxo-phytodienoic acid; JA, jasmonic acid.

All data Pantano de la Peña San Juan de la Peña Las Blancas
RWC 0.370 (< 0.001) 0.293 (0.009) 0.256 (0.021) 0.528 (< 0.001)
LMA 0.347 (< 0.001) 0.331 (0.004) 0.347 (0.003) 0.344 (0.001)
Chl a+b −0.028 (0.345) −0.186 (0.071) −0.170 (0.092) 0.243 (0.014)
Chl a/b 0.098 (0.080) 0.354 (0.002) 0.244 (0.027) −0.084 (0.227)
LOOH −0.166 (0.009) 0.016 (0.451) 0.120 (0.178) 0.291 (0.004)
Ant −0.034 (0.314) −0.206 (0.052) −0.172 (0.089) 0.221 (0.023)
Ant/Chl −0.025 (0.359) −0.084 (0.255) −0.032 (0.401) −0.039 (0.364)
Car −0.081 (0.123) −0.313 (0.006) −0.087 (0.249) 0.169 (0.064)
Car/Chl −0.093 (0.090) −0.261 (0.019) 0.067 (0.301) −0.203 (0.034)
α-Toc −0.147 (0.017) −0.331 (0.004) −0.224 (0.039) 0.091 (0.207)
α-Toc/Chl −0.046 (0.253) 0.013 (0.459) −0.057 (0.327) −0.089 (0.213)
ABA −0.037 (0.298) −0.044 (0.366) −0.152 (0.130) −0.073 (0.257)
SA 0.030 (0.334) 0.183 (0.074) −0.220 (0.050) 0.086 (0.221)
OPDA −0.005 (0.471) 0.070 (0.292) −0.079 (0.281) −0.032 (0.387)
JA 0.008 (0.453) −0.010 (0.468) −0.115 (0.196) −0.018 (0.437)
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Figure 5.

Figure 5.

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Spearman rank's correlation analyses between plant size of juvenile plants (estimated as rosette diameter) and the relative water content (RWC) in three populations of the long-lived monocarpic plant, S. longifolia. rho (r) and P values are indicated in the inlets (correlation was significant in the population at the highest altitude only, Las Blancas, P < 0.0033, Bonferroni adjusted).

Plant Maturity and Death

As a monocarpic species, S. longifolia dies right after blooming. During flowering, leaves serve as an important source of photoassimilates, but then the plant enters into a programmed, senescing process leading to death. We were interested in evaluating possible differences in plant physiological performance between juvenile and mature plants, and particularly between mature plants growing at different altitudes. With this purpose, we measured stress indicators in both juvenile and mature plants (at a flowering stage) in the low and intermediate populations (no sufficient individuals could be sampled for analyses in the high population due to extremely low reproductive events during 2015). Plant maturity increased the leaf mass area ratio, and the levels of antioxidants (carotenoids and α-tocopherol), ABA and jasmonic acid, while decreased those of 12-oxo-phytodienoic acid in the two populations studied (Table III), thus indicating that plant maturity led to enhanced physiological stress. Furthermore, mature plants of the intermediate population showed lower leaf water contents, higher α-tocopherol levels, and a lower extent of lipid peroxidation, as estimated by lipid hydroperoxides, compared to mature plants from the low population (Table III).

Table III. Influence of maturity for all measured parameters in the long-lived monocarpic plant, S. longifolia.

Data correspond to the mean ± se. n = 64 juvenile and n = 8 mature plants in Pantano de la Peña, and n = 64 juvenile and n = 10 mature plants in San Juan de la Peña. An asterisk indicates differences between juvenile and mature plants (Student's t test, P < 0.05). Different letters indicate differences between populations, either in juvenile or mature plants, the latter indicated in capital letters (Student's t test, P < 0.05). No sufficient plants in a mature stage were found in Las Blancas, so data are not available for the population at the highest altitude. RWC, relative water content; LMA, leaf mass per area ratio; Chl, chlorophyll. LOOH, lipid hydroperoxides; Ant, anthocyanins; Car, carotenoids; α-Toc, α-tocopherol; ABA, abscisic acid; SA, salicylic acid; OPDA, oxo-phytodienoic acid; JA, jasmonic acid.

Pantano de la Peña
 San Juan de la Peña
Juvenile Mature Juvenile Mature
Diameter (mm) 72.1 ± 3.2a 96.3 ± 20.0A 71.8 ± 3.9a 116.0 ± 14.6*A
RWC (%) 92.4 ± 0.8a 93.0 ± 2.5A 78.9 ± 1.5b 65.0 ± 5.6*B
LMA (g/m2) 156.1 ± 4.7a 72.5 ± 7.9*A 161.1 ± 5.1a 78.0 ± 6.6*A
Chl a+b (μmol/g DW) 1.33 ± 0.08a 1.39 ± 0.25A 1.70 ± 0.07b 1.61 ± 0.22A
Chl a/b 2.26 ± 0.02a 1.88 ± 0.04*A 2.01 ± 0.02b 1.79 ± 0.07*A
LOOH (μmol/g DW) 6.94 ± 0.49a 11.86 ± 2.84A 5.84 ± 0.71a 5.33 ± 1.23B
Ant (μmol/g DW) 0.61 ± 0.03a 0.60 ± 0.11A 0.71 ± 0.04b 0.78 ± 0.12A
Ant /Chl 0.49 ± 0.03a 0.45 ± 0.04A 0.42 ± 0.01b 0.52 ± 0.10A
Car (μmol/g DW) 0.29 ± 0.02a 0.43 ± 0.07*A 0.44 ± 0.02b 0.56 ± 0.06*A
Car/Chl 0.23 ± 0.01a 0.32 ± 0.02*A 0.26 ± 0.01b 0.37 ± 0.03*A
α-Toc (μmol/g DW) 0.28 ± 0.01a 0.39 ± 0.05*A 0.32 ± 0.01b 0.56 ± 0.03*B
α-Toc /Chl 0.26 ± 0.02a 0.34 ± 0.06A 0.20 ± 0.01b 0.40 ± 0.05*A
ABA (ng/g DW) 424.5 ± 21.3a 937.3 ± 153.5*A 598.3 ± 38.7b 848.8 ± 63.9*A
SA (ng/g DW) 375.9 ± 12.0a 674.4 ± 89.9*A 472.6 ± 24.6b 568.4 ± 58.3A
OPDA (ng/g DW) 2788 ± 225a 440 ± 103*A 1289 ± 158b 256 ± 38*A
JA (ng/g DW) 187.5 ± 7.9a 379.3 ± 74.6*A 216.2 ± 14.4b 550.1 ± 273.7A
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Clonal Growth, Reproduction, and Death

Rosettes might split into two or many more smaller rosettes in a given year, in a kind of clonal growth process, but without the presence of rhizomes because the single axonomorphic root stays inside the crevice (see Supplemental Fig. S1B). The frequency of multiple-rosette individuals ranged from 7–11% across the 4 years of study in Las Blancas, whereas it was as low as 2% in San Juan de la Peña and absent in Pantano de la Peña. Although clonal growth might happen at any plant size, it is more frequent among large plants (average diameter of nonclonal and clonal plants: 48.2 mm and 83.4 mm, respectively; n = 264 and 18 plants recorded in 2015, respectively). Survival did not differ between individuals with or without clonal reproduction (Fisher's exact test, P = 0.141, n = 1075), and none of the physiological parameters measured differed between nonclonal and clonal plants, except for the relative leaf water content, which was significantly lower in clonal plants (Table IV).

Table IV. Influence of clonal growth for all measured parameters in the highest population (Las Blancas) of the long-lived monocarpic plant, S. longifolia.

Data correspond to the mean ± se of n = 74 nonclonal and n = 9 clonal plants. An asterisk indicates differences between juvenile and mature plants (Student's t test, P < 0.05). RWC, relative water content; LMA, leaf mass per area ratio; Chl, chlorophyll. LOOH, lipid hydroperoxides; Ant, anthocyanins; Car, carotenoids; α-Toc, α-tocopherol; ABA, abscisic acid; SA, salicylic acid; OPDA, oxo-phytodienoic acid; JA, jasmonic acid.

Non-Clonal Clonal
Diameter (mm) 67.63 ± 2.72 66.71 ± 7.75
RWC (%) 84.01 ± 1.04 75.74 ± 1.81*
LMA(g/m2) 144.24 ± 3.55 131.31 ± 7.52
Chl a + b(μmol/g DW) 1.52 ± 0.06 1.34 ± 0.11
Chl a / b 2.17 ± 0.02 2.24 ± 0.04
LOOH(μmol/g DW) 5.38 ± 0.37 3.94 ± 0.60
Ant(μmol/g DW) 0.53 ± 0.02 0.57 ± 0.10
Ant / Chl 0.36 ± 0.01 0.46 ± 0.13
Car (μmol/g DW) 0.29 ± 0.01 0.30 ± 0.01
Car / Chl 0.20 ± 0.01 0.22 ± 0.01
α-Toc (μmol/g DW) 0.34 ± 0.01 0.30 ± 0.02
α-Toc / Chl 0.25 ± 0.01 0.24 ± 0.02
ABA(ng/g DW) 336.22 ± 15.49 349.76 ± 28.53
SA(ng/g DW) 383.11 ± 10.38 415.69 ± 46.77
OPDA(ng/g DW) 2428.69 ± 216.78 1609.05 ± 357.17
JA(ng/g DW) 119.57 ± 5.02 100.95 ± 15.27
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DISCUSSION

Our study of three populations of the long-lived monocarpic Mediterranean S. longifolia located at different altitudes in the Pyrenees has revealed that despite the complexity of mechanisms of adaptation to high altitude, which operate at the cellular, whole-plant, and population levels, this species may be vulnerable to drought stress events (periods of low precipitation combined with high solar radiation and high temperatures) during the summer in the framework of climate change.

Physiological Adaptation to Altitude: Photo- and Antioxidant Protection

Exposure to high solar radiation is known to induce photo-oxidative stress, particularly when it is accompanied by other stress conditions, such as extreme temperatures, as it occurs at high altitude (Streb et al., 1997, 2003a, 2003b; Pintó-Marijuan and Munné-Bosch, 2014). Furthermore, global change may increase the frequency of drought events, which occur irregularly and may therefore affect plant populations from a given species in a rather different way just depending on its specific location, leading to increased photo-oxidative stress. In chloroplasts, production of reactive oxygen species under excess light conditions is mainly mediated by the triplet excitation state of chlorophyll, which can lead to singlet oxygen formation, as well as by the photoreduction of oxygen through photosynthetic electron transport in the Mehler reaction, leading to the production of superoxide anions (Asada 2006). Plants have developed a variety of protective systems, which allow them to control ROS levels, so that oxidative damage can be prevented. Among them, carotenoids, acting as scavengers of triplet chlorophyll and singlet oxygen, and mediating the harmless dissipation of excess excitation energy as heat (Demmig-Adams and Adams 1993; Demmig-Adams et al., 2013, 2014); anthocyanins, acting as a filter for high-level energy from the blue and UV light region of the spectrum (Landi et al., 2015); and tocochromanols, both quenching and scavenging singlet oxygen, and inhibiting the propagation of lipid peroxidation (Havaux et al., 2005; Munné-Bosch, 2005; Triantaphylidès and Havaux, 2009; Falk and Munné-Bosch, 2010), play a key role. Results shown here for an orophyte endemism of the western Mediterranean illustrates that despite the complex mechanisms evolved by plants at the cellular level to survive high-mountain conditions, drought stress is one of the main triggers of mortality in S. longifolia. The intermediate population was the one showing the highest mortality rates, which paralleled with the lowest soil and leaf water contents and the activation of defense responses (e.g. ABA accumulation). Increased water availability in the high population (compared to the other ones) most likely led to a reduced need for photoprotection despite increased high light exposure due to the altitudinal gradient. Interestingly, however, α-tocopherol levels increased as a function of altitude, their biosynthesis being mostly governed by high light exposure (Havaux et al., 2005). This pattern was paralleled with reductions in lipid hydroperoxides, thus indicating the protective role of vitamin E in preventing the propagation of lipid peroxidation in the chloroplasts, which is in agreement with previous studies on other high-mountain plants (Streb et al., 1997, 2003a, 2003b). Unfortunately, chlorophyll fluorescence measurements could not be performed in intact leaves from this species in the field due to the high reflectance of the epidermis, an aspect that warrants further investigation.

Activation of chemical defenses, such as the biosynthesis of salicylates and jasmonates, are known to be influenced by both biotic and abiotic stress factors (Davies, 2010). In the current study, both groups of compounds increased with altitude (comparing the intermediate and low populations), but its accumulation was abolished at the highest altitude, most likely due, at least in part, to improved soil and leaf water contents (compared to the other two populations). Other factors may, however, also influence the accumulation of salicylates and jasmonates. In the high population, salicylic acid accumulation was similar to that of the low population, and that of jasmonic acid was even smaller, thus indicating a reduced need for chemical defense against necrotrophs at 2100 m a.s.l. (Davies, 2010). It is also likely that reduced jasmonic acid levels result from a trade-off between activation of different defense pathways in plants (photoprotection versus potential chemical defense to necrotrophs through jasmonates), so that enhanced vitamin E accumulation at the highest altitude may negatively influence the biosynthesis of jasmonates, which is in agreement with previous studies (Demmig-Adams et al., 2013, 2014; Morales et al., 2015; Simancas and Munné-Bosch, 2015). Enhanced jasmonic acid accumulation in the intermediate population may reflect activation of acclimation responses, but also increased cell death, as shown in other studies (Shumbe et al., 2016). Enhanced jasmonic acid levels may be triggered by increased biotic stress, but also by abiotic factors, such as drought stress (Brossa et al., 2011; de Ollas et al., 2013). Thus, it is very likely that enhanced ABA, salicylic acid, and jasmonic acid levels all respond to an increased drought stress that activates acclimation responses but that ultimately lead to increased cell death and mortality in the intermediate population. Results suggest that enhanced physiological stress and mortality in the intermediate population was caused by increased drought stress during 2015. If more drought events occur in the other two populations, which are indeed likely to increase in the frame of global change (IPCC, 2014), it is expected they will also result in an increased mortality. More frequent snowfalls lead, however, to an increased water availability in the highest population. It may therefore be anticipated that, as precipitation patterns suggest (Supplemental Fig. S2), the populations found at the two lowest altitudes will be the ones showing the highest sensitivity to drought stress-induced mortality.

Adaptation at the Population Level: Plant Size, Clonal Growth, and Population Size Structure

The population at highest altitude showed some demographic differences compared to the other two sampled populations: lower frequency of small individuals, size-independent mortality rate, the largest plants of all recorded across populations, and a particular trait that was almost absent in the other two—clonal growth (Supplemental Fig. S1B). The lower frequency of small-sized plants in the high population is the consequence of lower recruitment (very few new individuals enter each year in the monitoring plots; M.B. García, unpublished data). Interestingly, small rosettes at high altitude survive better than at lower altitudes (Supplemental Fig. S4), and mortality was more uniformly distributed with size in the high population compared to the other two populations. Higher survival in the highest population may be associated with improved soil and leaf water contents despite being exposed to higher light intensity. Furthermore, plants from this species seem to escape from increased size-dependent mortality, as it has been shown in other plant species, mostly woody perennials (shrubs and trees), in which increased plant size make larger individuals more vulnerable to environmental constraints (Mencuccini et al., 2005, 2007; Baudisch et al., 2013; Salguero-Gomez et al., 2013­; Munné-Bosch, 2014, 2015). This was not observed here in either studied population. Small sizes throughout their lifespan, as it happens in perennial herbs (García et al., 2011; Morales et al., 2013; Morales and Munné-Bosch, 2015), may protect S. longifolia plants from the potential negative effects of aging. It appears, therefore, that whole-plant senescence in this species may be attributed to reproduction and extrinsic factors, such as drought stress, but not to ageing, as only very small individuals (< 30 mm diameter) die more frequently in the two lowest populations. Interestingly, those populations are the more exposed ones to summer drought. It may be speculated that an increase in temperatures and drought events in the framework of global change may be a serious threat for this species. Furthermore, population recruitment is lowest at the highest population, and reduced recruitment would translate into a negative population dynamics, a real limitation for adaptation in the framework of global change.

The fact that plants get larger in the high population could be related to the existence of clonal growth, another interesting mechanism observed mainly at the highest altitude. Newly formed rosettes never become fully independent because they all share the same root system, but they can behave independent in the sense that not all daughter rosettes die or reproduce at the same time (see Supplemental Fig. S1B). Considering that this is a monocarpic plant, forming new rosettes might help the plant to reduce mortality as a consequence of reproduction and extend the fecundity period like a polycarpic organism, spreading fitness over time. Flowering plants with one single rosette inevitably die the same year of reproduction, whereas 31% of multiple-rosette individuals survived. Therefore, having more than one rosette allows the plant to decide which ones to “sacrifice,” which translates into survival of the individual if not all rosettes synchronously flower in a given year. Clonal growth, thus, constitutes an additional process operating at the individual level in terms of enhancing survival, which could also help to explain the lower mortality rate of this population compared to the other ones. The particular habitat or environment has been shown to be an important selective factor in predicting the evolutionary stable reproductive strategy in natural populations of monocarpic plants (Hesse et al., 2008), and S. longifolia constitutes a clear example of reproductive strategy variability along an altitudinal gradient.

It is concluded that, despite the endemic Mediterranean plant, S. longifolia has evolved complex mechanisms of adaptation to altitude (including e.g. enhanced α-tocopherol levels or changes in reproductive strategy like activation of clonal growth), this species is rather sensitive to drought stress, and consequently, drought events may drive increased mortality in populations from this species in the framework of global change. Further research is needed, however, to better understand the mechanisms underlying the influence of altitude and drought on size-dependent mortality, how they interact, and how this will in turn be affected by global warming in this and other endemic plants in the near future.

MATERIALS AND METHODS

Plant Populations, Treatments, and Sampling

The study was carried out in three natural populations of Saxifraga longifolia Lapeyruse located in central Pyrenees, the area of highest abundance within its distribution range. The three populations were located across an altitudinal range spanning 50 km in straight line. The first population was located in rocky walls of limestone near Pantano de la Peña (570 m a.s.l., coordinates: 42°22'58.4”N 0°44'02.1”W), the second one occurred on a very sloppy conglomerated area near San Juan de la Peña (1100 m a.s.l., 42°30'30.6”N 0°40'21.3”W), and the third one in the uppermost needles of calcareous mountain named Las Blancas (2100 m a.s.l., 42°44'49.3”N 0°33'26.4”W). This long-lived monocarpic perennial plant develops a basal rosette growing in the crevices of limestone rocky places, mainly on cliffs, offering a unique sight in years of intensive blooming.

Samplings in at least 70 individuals randomly selected within each population among plants larger than 30 mm of diameter were performed for biochemical analyses in 2015, including both juvenile and reproductive plants, except in Las Blancas, where too few individuals flowered during 2015. Sixty-five, 64, and 83 juveniles were sampled in Pantano de la Peña, San Juan de la Peña, and Las Blancas, respectively, and additionally, 8 and 10 mature individuals were sampled in the two former populations. Samplings were performed on fully expanded leaves at midday (12 a.m. solar time) on June 22, July 3, and August 18 in Pantano de la Peña, San Juan de la Peña, and Las Blancas, respectively, just after flowering in mature plants, so that all plants were at the same phenological stage. Rosette leaves were used to estimate leaf water contents, pigment concentrations (including chlorophylls, carotenoids, and anthocyanins), levels of vitamin E, the extent of lipid peroxidation, as well as the endogenous concentrations of stress-related phytohormones, including ABA, salicylates, and jasmonates. Samples for biochemical analyses were collected, immediately frozen in liquid nitrogen in situ, and stored at −80°C upon arrival to the laboratory.

Mortality

Between two and three hundred plants per location were marked and annually monitored from 2011 through 2015 to estimate survival rates with the aid of grid plots. In summer each year, all numbered plants were checked, and if alive their diameter were recorded. Annual flowering and mortality rates were compared after pooling all the events recorded along pairs of consecutive years (2011/2012, 2012/2013, 2013/2014 and 2014/2015). Furthermore, in order to explore possible reasons and consequences of clonal reproduction in the highest population, we tested if juvenile and mature plants with multiple rosettes had a different survival probability than singled-rosette ones.

Leaf Water Contents, Pigment Levels, and Lipid Peroxidation

To estimate leaf water contents, samples were collected, kept humid in small bags in darkness during transport to the laboratory, and then weighed to estimate fresh matter (FW). They were immersed in distilled water at 4°C for 24 h to estimate the turgid matter (TW) and then oven-dried at 80°C to constant weight to estimate the dry matter (DW). Relative water content (RWC) was then calculated as 100 × (FW−DW)/(TW−DW).

For pigment analysis, measurements were performed spectrophotometrically on methanolic extracts to estimate chlorophyll and carotenoid levels, as described by Lichtenthaler (1987), which were then acidified with 30% HCl to estimate total anthocyanin levels as described by Gitelson et al. (2001). The extent of lipid peroxidation was estimated by measuring the levels of lipid hydroperoxides in leaves. Lipid hydroperoxide levels were estimated spectrophotometrically following a modified ferrous oxidation-xylenol orange (FOX) assay, as described in DeLong et al. (2002).

Tocochromanols

For analyses of tocochromanol (tocopherols and plastochromanol-8) contents, leaf samples were ground in liquid nitrogen and extracted with cold methanol containing 0.01% butylated hydroxyltoluene using ultrasonication. After centrifuging at 12,000 rpm for 10 min at 4°C, the supernatant was collected and the pellet was re-extracted with the same solvent until it was colorless. Then, supernatants were pooled, filtered, and injected into the HPLC. Platochromanols were separated isocratically on a normal-phase HPLC system using a fluorescent detector as described (Cela et al., 2011). Compounds were identified by coelution with authentic standards and quantified using a calibration curve. From all tocochromanols investigated (α-, β-, γ-, and δ-tocopherols and plastochromanol-8), α-tocopherol was the only compound present at quantifiable amounts in leaves.

Stress-Related Phytohormones

For analyses of ABA, salicylic acid, and jasmonates, leaf samples were ground in liquid nitrogen and extracted with cold methanol using ultrasonication. After centrifuging at 12,000 rpm for 10 min at 4°C, the supernatant was collected and the pellet was re-extracted with the same solvent until it was colorless. Then, supernatants were pooled, filtered, and injected into the UHPLC-MS/MS. Phytohormones were separated using an elution gradient on a reverse-phase UHPLC system and quantified using tandem mass spectrometry in multiple reaction monitoring mode as described (Müller and Munné-Bosch, 2011). Recovery rates were calculated for each hormone on every sample by using deuterated compounds.

Statistical Analysis

To determine the effect of altitude, mean values were tested by one-way factorial ANOVA and additionally Bonferroni posthoc tests. Mean values were compared between clonal and nonclonal plants by means of Student's t test. In all cases, differences were considered significant at a probability level of P < 0.05. Spearman rank's correlation analyses were performed between plant size (estimated as rosette diameter) and all biochemical parameters, and Bonferroni correction applied to determine significant differences. Statistical tests were carried out using the SPSS 20.0 statistical package. The effect of individual size on mortality probability was explored by fitting logistic regression models (logit link function, binomial distribution, in R version 2.15.2, Core Team) for all the annual transitions recorded over that period, for each population separately.

Supplemental Data

The following supplemental materials are available.

  • Supplemental Figure S1. Saxifraga longifolia population structure in 2011, according to rosette diameter (seedlings excluded).

  • Supplemental Figure S2. Monthly average temperatures, soil water contents and precipitation recorded in the three studied populations.

  • Supplemental Figure S3. Levels of anthocyanins, carotenoids and α-tocopherol, expressed per chlorophyll (Chl) unit, in plants of the long-lived monocarpic plant, S. longifolia growing at three altitudes (570, 1100, and 2100 m a.s.l. in Pantano de la Peña, San Juan de la Peña, and Las Blancas, respectively).

  • Supplemental Figure S4. Mortality rate for plants of different sizes.

Supplementary Material

Supplemental Data
supp_172_2_765__index.html (775B, html)

Acknowledgments

We are very grateful to Maren Müller and Serveis Científico-tècnics for technical help with biochemical analyses. We are also indebted to M. Paz Errea and Ricardo García González for providing environmental data, and the fieldwork assistance of Juanlu, Iker, P. Sánchez, and P. Bravo.

Footnotes

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References

  1. Asada K. (2006) Production and scavenging of reactive oxygen species in chloroplasts and their functions. Plant Physiol 141: 391–396 [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bano A, Rehman A, Winiger M (2009) Altitudinal variation in the content of protein, proline, sugar and abscisic acid (ABA) in the alpine herbs from Hunza valley, Pakistan. Pak J Bot 41: 1593–1602 [Google Scholar]
  3. Baudisch A, Salguero-Gómez R, Jones OR, Wrycza T, Mbeau-Ache C, Franco M, Colchero F (2013) The pace and shape of senescence in angiosperms. J Ecol 101: 596–606 [Google Scholar]
  4. Breed GA, Stichter S, Crone EE (2013) Climate-driven changes in northeastern US butterfly communities. Nat Clim Chang 3: 142–145 [Google Scholar]
  5. Brossa R, López-Carbonell M, Jubany-Marí T, Alegre L (2011) Interplay between abscisic acid and jasmonic acid and its role in water-oxidative stress in wild type, ABA-deficient, JA-deficient, and ascorbate-deficient Arabidopsis plants. J Plant Growth Regul 30: 322–333 [Google Scholar]
  6. Cela J, Chang C, Munné-Bosch S (2011) Accumulation of γ- rather than α-tocopherol alters ethylene signaling gene expression in the vte4 mutant of Arabidopsis thaliana. Plant Cell Physiol 52: 1389–1400 [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Davies PJ. (2010) The plant hormones: their nature, occurrence, and functions. In Davies PJ, ed, Plant Hormones: Biosynthesis, Signal Transduction, Action! Dordrecht: Springer, pp. 1–15. [Google Scholar]
  8. DeLong JM, Prange RK, Hodges DM, Forney CF, Bishop MC, Quilliam M (2002) Using a modified ferrous oxidation-xylenol orange (FOX) assay for detection of lipid hydroperoxides in plant tissue. J Agric Food Chem 50: 248–254 [DOI] [PubMed] [Google Scholar]
  9. Demmig-Adams B, Adams WW III (1993) The xanthophyll cycle. In Alscher RG, Hess JL, eds, Antioxidants in Higher Plants, CRC Press, Boca Raton, FL, pp. 91-110. [Google Scholar]
  10. Demmig-Adams B, Cohu CM, Amiard V, Zadelhoff G, Veldink GA, Muller O, Adams WW III (2013) Emerging trade-offs – impact of photoprotectants (PsbS, xanthophylls, and vitamin E) on oxylipins and biotic defense. New Phytol 197: 720–729 [DOI] [PubMed] [Google Scholar]
  11. Demmig-Adams B, Stewart JJ, Adams WW III (2014) Chloroplast photoprotection and the trade-off between abiotic and biotic defense. In B Demmig-Adams, G Garab, WW Adams III, Govindjee, eds, Nonphotochemical Quenching and Energy Dissipation in Plants, Algae and Cyanobacteria. Advances in Photosynthesis and Respiration, Vol. 40. Springer, Dordrecht, The Netherlands. pp. 631–643 [Google Scholar]
  12. de Ollas C, Hernando B, Arbona V, Gómez-Cadenas A (2013) Jasmonic acid transient accumulation is needed for abscisic acid increase in citrus roots under drought stress conditions. Physiol Plant 147: 296–306 [DOI] [PubMed] [Google Scholar]
  13. Dong C-J, Li L, Shang Q-M, Liu X-Y, Zhang Z-G (2014) Endogenous salicylic acid accumulation is required for chilling tolerance in cucumber (Cucumis sativus L.) seedlings. Planta 240: 687–700 [DOI] [PubMed] [Google Scholar]
  14. Falk J, Munné-Bosch S (2010) Tocochromanol functions in plants: antioxidation and beyond. J Exp Bot 61: 1549–1566 [DOI] [PubMed] [Google Scholar]
  15. Franklin J, Serra-Diaz JM, Syphard AD, Regan HM (2016) Global change and terrestrial plant community dynamics. Proc Natl Acad Sci USA 113: 3725–3734 [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. García MB. (2003) Sex allocation in a long-lived monocarpic plant. Plant Biol 5: 203–209 [Google Scholar]
  17. García MB, Dahlgren JP, Ehrlén J (2011) No evidence of senescence in a 300-year-old mountain herb. J Ecol 99: 1424–1430 [Google Scholar]
  18. García-Plazaola JI, Rojas R, Christie DA, Coopman RE (2015) Photosynthetic responses of trees in high-elevation forests: comparing evergreen species along an elevation gradient in the Central Andes. AoB Plants 7: 58. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gilmour SJ, Thomashow MF (1991) Cold acclimation and cold-regulated gene expression in ABA mutants of Arabidopsis thaliana. Plant Mol Biol 17: 1233–1240 [DOI] [PubMed] [Google Scholar]
  20. Gitelson AA, Merzlyak MN, Chivkunova OB (2001) Optical properties and nondestructive estimation of anthocyanin content in plant leaves. Photochem Photobiol 74: 38–45 [DOI] [PubMed] [Google Scholar]
  21. Gottfried M, Pauli H, Futschik A, Akhalkatsi M, Barančok P, Alonso JLB, Coldea G, Dick J, Erschbamer B, Calzado MRF, Kazakis G, Krajči J, et al. (2012) Continent-wide response of mountain vegetation to climate change. Nat Clim Chang 2: 111–115 [Google Scholar]
  22. Havaux M, Eymery F, Porfirova S, Rey P, Dörmann P (2005) Vitamin E protects against photoinhibition and photooxidative stress in Arabidopsis thaliana. Plant Cell 17: 3451–3469 [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hesse E, Rees M, Müller-Schärer H (2008) Life-history variation in contrasting habitats: flowering decisions in a clonal perennial herb (Veratrum album). Am Nat 172: 196–213 [DOI] [PubMed] [Google Scholar]
  24. IPCC (2014) Summary for policymakers. In Field CB, Barros VR, Dokken DJ, Mach KJ, Mastrandrea MD, Bilir TE, Chatterjee M, Ebi KL, Estrada YO, Genova RC et al., eds, Climate change 2014: Impacts, Adaptation, and Vulnerability. Contribution of Working Group II to the Fifth Assessment Report of the Intergovernmental Panel on Climate Change. Cambridge University Press, Cambridge, UK, and New York, NY, pp. 1–32 [Google Scholar]
  25. Körner C. (2013) Alpine ecosystems. In Levin SA, ed, Encyclopedia of Biodiversity, 2nd edition, Vol. 1, Academic Press, Amsterdam, The Netherlands, pp. 148–157 [Google Scholar]
  26. Kosová K, Prášil IT, Vítámvás P, Dobrev P, Motyka V, Floková K, Novák O, Turečková V, Rolčik J, Pešek B, Trávničková A, Gaudinová A, et al. (2012) Complex phytohormone responses during the cold acclimation of two wheat cultivars differing in cold tolerance, winter Samanta and spring Sandra. J Plant Physiol 169: 567–576 [DOI] [PubMed] [Google Scholar]
  27. Kruk J, Szymanska R, Cela J, Munné-Bosch S (2014) Plastochromanol-8: Fifty years of research. Phytochemistry 108: 9–16 [DOI] [PubMed] [Google Scholar]
  28. Landi M, Tattini M, Gould KS (2015) Multiple functional roles of anthocyanins in plant-environment interactions. Environ Exp Bot 119: 4–17 [Google Scholar]
  29. Larcher W. (1994) Okologie der Pflanzen. Stuttgart, Germany, Ulmer [Google Scholar]
  30. Lichtenthaler HK. (1987) Chlorophylls and carotenoids: Pigments of photosynthetic biomembranes. Methods Enzymol 148: 350–382 [Google Scholar]
  31. Mencuccini M, Grace J (1996) Developmental patterns of aboveground hydraulic conductance in a Scots pine (Pinus sylvestris L.) age sequence. Plant Cell Environ 19: 939–948 [Google Scholar]
  32. Mencuccini M, Martínez-Vilalta J, Vanderklein D, Hamid HA, Korakaki E, Lee S, Michiels B (2005) Size-mediated ageing reduces vigour in trees. Ecol Lett 8: 1183–1190 [DOI] [PubMed] [Google Scholar]
  33. Mencuccini M, Martínez-Vilalta J, Hamid HA, Korakaki E, Vanderklein D (2007) Evidence for age- and size-mediated controls of tree growth from grafting studies. Tree Physiol 27: 463–473 [DOI] [PubMed] [Google Scholar]
  34. Miura K, Tada Y (2014) Regulation of water, salinity, and cold stress responses by salicylic acid. Front Plant Sci 5: 4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Morales M, Garcia QS, Siqueira-Silva AI, Silva MC, Munné-Bosch S (2014) Tocotrienols in Vellozia gigantea leaves: occurrence and modulation by seasonal and plant size effects. Planta 240: 437–446 [DOI] [PubMed] [Google Scholar]
  36. Morales M, Garcia QS, Munné-Bosch S (2015) Ecophysiological response to seasonal variations in water availability in the arborescent, endemic plant Vellozia gigantea. Tree Physiol 35: 253–265 [DOI] [PubMed] [Google Scholar]
  37. Morales M, Munné-Bosch S (2015) Secret of long life lies underground. New Phytol 205: 463–467 [DOI] [PubMed] [Google Scholar]
  38. Morales M, Oñate M, Garcia MB, Munné-Bosch S (2013) Photo-oxidative stress markers reveal absence of physiological deterioration with ageing in Borderea pyrenaica, an extraordinarily long-lived herb. J Ecol 101: 555–565 [Google Scholar]
  39. Müller M, Munné-Bosch S (2011) Rapid and sensitive hormonal profiling of complex plant samples by liquid chromatography coupled to electrospray ionization tandem mass spectrometry. Plant Methods 7: 37. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Munné-Bosch S. (2005) Linking tocopherols with cellular signaling in plants. New Phytol 166: 363–366 [DOI] [PubMed] [Google Scholar]
  41. Munné-Bosch S. (2014) Perennial roots to immortality. Plant Physiol 166: 720–725 [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Munné-Bosch S. (2015) Senescence: is it universal or not? Trends Plant Sci 20: 713–720 [DOI] [PubMed] [Google Scholar]
  43. Munné-Bosch S, Alegre L (2002) The function of tocopherols and tocotrienols in plants. Crit Rev Plant Sci 21: 31–57 [Google Scholar]
  44. Munné-Bosch S, Lalueza P (2007) Age-related changes in oxidative stress markers and abscisic acid levels in a drought-tolerant shrub, Cistus clusii grown under Mediterranean field conditions. Planta 225: 1039–1049 [DOI] [PubMed] [Google Scholar]
  45. Peñuelas J, Boada M (2003) A global change-induced biome shift in the Montseny mountains (NE Spain). Glob Change Biol 9: 131–140 [Google Scholar]
  46. Pintó-Marijuan M, Munné-Bosch S (2014) Photo-oxidative stress markers as a measure of abiotic stress-induced leaf senescence: advantages and limitations. J Exp Bot 65: 3845–3857 [DOI] [PubMed] [Google Scholar]
  47. Salguero-Gómez R, Shefferson RP, Hutchings MJ (2013) Plants do not count… or do they? New perspectives on the universality of senescence. J Ecol 101: 545–554 [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Scherrer D, Körner C (2011) Topographically controlled thermal-habitat differentiation buffers alpine plant diversity against climate warming. J Biogeogr 38: 406–416 [Google Scholar]
  49. Schmidt G, Stuntz S, Zotz G (2001) Plant size - an ignored parameter in epiphyte ecophysiology. Plant Ecol 153: 65–72 [Google Scholar]
  50. Schmidt G, Zotz G (2001) Ecophysiological consequences of differences in plant size - in situ carbon gain and water relations of the epiphytic bromeliad, Vriesea sanguinolenta. Plant Cell Environ 24: 101–112 [Google Scholar]
  51. Shumbe L, Chevalier A, Legeret B, Taconnat L, Monnet F, Havaux M (2016) Singlet oxygen-induced cell death in arabidopsis under high-light stress is controlled by OXI1 kinase. Plant Physiol 170: 1757–1771 [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Simancas B, Munné-Bosch S (2015) Interplay between vitamin E and phosphorus availability in the control of longevity in Arabidopsis thaliana. Ann Bot 116: 511–518 [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Streb P, Shang W, Feierabend J, Bligny R (1998) Divergent strategies of photoprotection in high-mountain plants. Planta 207: 313–324 [Google Scholar]
  54. Streb P, Aubert S, Bligny R (2003a) High temperature effects on light sensitivity in the two high mountain plant species Soldanella alpina (L.) and Ranunculus glacialis (L.). Plant Biol 5: 432–440 [Google Scholar]
  55. Streb P, Aubert S, Gout E, Bligny R (2003b) Reversibility of cold- and light-stress tolerance and accompanying changes of metabolite and antioxidant levels in the two high mountain plant species Soldanella alpina and Ranunculus glacialis. J Exp Bot 54: 405–418 [DOI] [PubMed] [Google Scholar]
  56. Streb P, Feierabend J, Bligny R (1997) Resistance to photoinhibition of photosystem II and catalase and antioxidative protection in high mountain plants. Plant Cell Environ 20: 1030–1040 [Google Scholar]
  57. Triantaphylidès C, Havaux M (2009) Singlet oxygen in plants: production, detoxification and signaling. Trends Plant Sci 14: 219–228 [DOI] [PubMed] [Google Scholar]
  58. Webb DA, Gornall RJ (1989) Saxifrages of Europe. London, UK: Christopher Helm Publishers [Google Scholar]
  59. Zbierzak AM, Kanwischer M, Wille C, Vidi PA, Giavalisco P, Lohmann A, Briesen I, Porfirova S, Bréhélin C, Kessler F, Dörmann P (2009) Intersection of the tocopherol and plastoquinol metabolic pathways at the plastoglobule. Biochem J 425: 389–399 [DOI] [PubMed] [Google Scholar]
  60. Zotz G. (1997) Photosynthetic capacity increases with plant size. Bot Acta 110: 306–308 [Google Scholar]
  61. Zotz G, Hietz P, Schmidt G (2001) Small plants, large plants: the importance of plant size for the physiological ecology of vascular epiphytes. J Exp Bot 52: 2051–2056 [DOI] [PubMed] [Google Scholar]

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