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(American Journal of Botany. 2001;88:1401-1408.)
© 2001 Botanical Society of America, Inc.

Population Biology

Susceptibility to UV damage in Impatiens capensis (Balsaminaceae): testing for opportunity costs to shade-avoidance and population differentiation1

Peter Dixon, Cynthia Weinig2 and Johanna Schmitt

Department of Ecology and Evolutionary Biology, Box G-W, Brown University, Providence, Rhode Island 02912 USA

Received for publication August 22, 2000. Accepted for publication January 30, 2001.


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 LITERATURE CITED
 
Plastic increases in leaf secondary compounds may be an adaptive strategy that reduces the damaging effects of high-energy, ultraviolet radiation (UV). Here, we examine (1) the relationship between fitness and anthocyanin and flavonoid concentrations in experimental, UV environments, (2) the effects of UV on Impatiens capensis plants derived from woodland and clearing sites, and (3) whether susceptibility to UV damage is reduced by exposure to high ratios of red : far-red wavelengths (R : FR), which also stimulate the production of leaf compounds. Seedlings from each site were exposed to either high R : FR typical of sunlight or low R : FR characteristic of foliar shade, after which plants were moved into ambient UV or UV-removal treatments. Ultraviolet radiation stimulated the production of anthocyanins and flavonoids. However, higher anthocyanin concentrations were associated with lower biomass in the UV environment. Relative to the clearing population, reproductive output of the woodland population was more detrimentally affected by exposure to UV, despite its higher concentration of anthocyanins. Increased anthocyanin production may therefore be a stress response rather than an adaptive one. The greater tolerance of the clearing population to UV suggests that populations with an evolutionary history of UV exposure evolve mechanisms to limit damage. The R : FR pretreatments did not influence susceptibility to UV damage.

Key Words: anthocyanins • flavonoids • Impatiens capensis • local adaptation • phenotypic plasticity • R : FR • shade avoidance • ultraviolet (UV) radiation


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 LITERATURE CITED
 
Phenotypic plasticity is commonly viewed as a evolutionary strategy enabling individuals to adaptively match their phenotype to local conditions and avoid opportunity costs (or trade-offs) associated with fixed trait expressions (Levins, 1963 ; Bradshaw, 1965 ; Lively, 1986 ; Sultan, 1987 ). In plants, certain phenotypic responses to ultraviolet radiation (UV) may be explicitly adaptive, reducing the damaging effects of high-energy, UV radiation (Robberecht and Caldwell, 1978 ; Greenberg et al., 1997 ; Rozema et al., 1997 ). Upon exposure to UV, plants increase the production of leaf flavonoids and anthocyanins (Wellman, 1971; Tevini, Braun, and Fieser, 1991 ; Krizek, Britz, and Mirecki, 1998 ). Flavonoids absorb strongly in the UV region (Robberecht and Caldwell, 1978 ; Teramura, 1986 ) and appear to reduce the negative effects of UV on photosynthesis (Robberecht and Caldwell, 1983 ; Caldwell, Teramura, and Tevini, 1989 ; Wilson and Greenberg, 1993 ; Teramura and Sullivan, 1994 ) and plant growth (Robberecht and Caldwell, 1978 ; Caldwell, Robberecht, and Flint, 1983 ; Li et al., 1993 ; Kraus, Markstädter, and Riederer, 1997 ). Similar protective effects have also been suggested for anthocyanins (Takahashi, Takeda, and Ohnishi, 1991 ; Burger and Edwards, 1996 ; Coley and Kursar, 1996 , but see Gould, 1994 ; Woodall and Stewart, 1998 ). Other phenotypic responses to UV, e.g., decreases in biomass, may simply reflect the detrimental effects of a stressful environment on plant growth. To determine the role of UV as a selective agent, it is necessary to test whether selection for a trait is stronger in UV than in non-UV environments (Wade and Kalisz, 1991 ). Few studies have examined the relationship between fitness and anthocyanin or flavonoid concentrations for wild species growing under different levels of UV (Tevini and Teramura, 1989 ).

In addition to direct responses to UV, responses to the ratio of red : far-red wavelengths (R : FR) may influence susceptibility to UV damage. Responses to environmental R : FR have recently received attention from evolutionary biologists, because the responses provide a clear example of adaptive phenotypic plasticity (Smith, 1982 ; Ballaré, Scopel, and Sanchez, 1990 ; Schmitt and Wulff, 1993 ; Schmitt, 1997 ). For plants, low R : FR reliably indicates the presence of neighboring plants and the onset of competition for sunlight (Smith, 1982 ; Ballaré, Scopel, and Sanchez, 1990 ; Schmitt and Wulff, 1993 ; Smith and Whitelam, 1997 ), because vegetative canopies selectively absorb in the red region of the spectrum while transmitting far-red (Kasperbauer, 1971 ; Smith, 1982 ; Smith, Casal, and Jackson, 1990 ). Photomorphogenic responses to low R : FR, such as increased stem elongation, enhance fitness among crowded plants (Schmitt, McCormac, and Smith, 1995 ; Dudley and Schmitt, 1996 ; Weinig, 2000a ) by increasing leaf exposure (Weinig, 2000a) . Dudley and Schmitt (1996) also demonstrated that elongation is explicitly maladaptive under uncrowded conditions; Impatiens capensis plants stimulated to elongate by low R : FR had reduced fitness relative to shorter conspecifics when planted in low-density plots. The reduced fitness of elongated relative to nonelongated plants was only partially explained by differences in height, demonstrating an intrinsic opportunity cost to elongation. One possibility is that elongated plants suffer greater UV damage because high R : FR stimulates the production of flavonoids and anthocyanins (Lange, Shropshire, and Mohr, 1971 ; Schmidt and Mohr, 1981 ), whereas low R : FR mediates elongation responses but not the production of leaf compounds. It is of interest to test whether increased UV susceptibility constitutes an opportunity cost of elongation because such costs influence evolutionary responses, countering the evolution of genotypes with fixed shade-avoidance phenotypes. Moreover, plants growing in dense stands would be subject to these opportunity costs as they initially experience low R : FR through lateral shading and subsequently emerge into high-UV environments when they overtop their neighbors.

Both UV and R : FR are likely to vary among populations of many species. Studies have now shown genetic differentiation for plasticity to R : FR among populations from different canopy environments (e.g., overhead, forest canopies vs. open, old fields) (Bain and Attridge, 1988 ; Dudley and Schmitt, 1995 ) and among populations occurring with different species of herbaceous competitors (Weinig, 2000b) . Those studies examining population susceptibility to UV have focused on populations located over an altitudinal or latitudinal gradient that experience graded levels of UV (e.g., Sullivan, Teramura, and Ziska, 1992 ; Ziska, Teramura, and Sullivan, 1992 ). Differences between natural populations from different canopy environments, which typically are either exposed to or sheltered from UV, remain to be investigated.

The study organism used here, Impatiens capensis Meerb. (Balsaminaceae), touch-me-not or jewelweed, occurs in both forest understories and old fields and exhibits microgeographic genetic differentiation in response to light availability (Schmitt, 1993 ) and R : FR (Dudley and Schmitt, 1995 ; Donohue and Schmitt, 1999 ). Adaptive plasticity or susceptibility to UV may show similar patterns of differentiation. This study therefore addresses four questions: (1) What are the morphological and fitness effects of ambient UV in I. capensis? (2) What is the relationship between anthocyanin or flavonoid concentrations and fitness under UV and non-UV conditions? (3) Do populations with different evolutionary histories of exposure to UV (clearing vs. woodland) differ in their responses to UV? (4) Does the seedling R : FR environment affect subsequent phenotypic responses to UV, i.e., is there an opportunity cost of shade-avoidance responses in terms of UV susceptibility?


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 LITERATURE CITED
 
Study system
Impatiens capensis is an annual species that commonly occurs in both moist, sunlit clearings and deciduous or evergreen woodlands. Source populations for this experiment were derived from the Haffenreffer Reserve of Brown University (Bristol, Rhode Island, USA); one of the populations was located at the margin of a deciduous forest and never experienced overhead shading, whereas the second was within the forest understory (Dudley and Schmitt, 1995 ; Donohue and Schmitt, 1999 ). The two populations will be referred to as clearing and woodland, respectively. Seeds used in this experiment were collected in the late winter of 1998 from greenhouse-grown, inbred lines derived from the two populations. Seeds were placed in microtitre trays filled with distilled water and stratified for 4 mo at 4°C (N = 394).

Experimental design
In a combined greenhouse and field experiment, we exposed seedlings of the two populations to one of two R : FR pretreatments (high vs. low) and then transplanted plants into one of two UV treatments (ambient environmental UV vs. UV removal). On 30 June 1998, seeds from the clearing and woodland populations were sown into four 128-cell trays filled; four trays were used in order that all seedlings in each of two could be transplanted into the UV treatments. Planting in multi-celled trays results in seedling densities that reduce R : FR and induce shade-avoidance responses among Impatiens grown under normal, sunlit conditions (Dudley and Schmitt, 1996 ). Equal numbers of seeds from each population were planted into random positions within the four trays. Two trays were placed under high R : FR conditions, while two experienced low R : FR conditions.

R : FR light treatments
The high R : FR conditions were established by filtering sunlight through solutions of copper sulfate (30 g/L) in plastic Ziploc bags on plexiglass trays. This solution selectively removes far-red wavelengths, resulting in constitutively high R : FR despite high seedling densities. The treatment effectively suppresses shade-avoidance responses (Ballaré, Scopel, and Sanchez, 1991 ; Dudley and Schmitt, 1996 ; Donohue and Schmitt, 1999 ). In the low R : FR treatment, seedlings were allowed to experience reductions in R : FR ensuing from the high-density planting. The solution of copper sulfate reduced levels of photosynthetically active radiation (PAR) in addition to lowering the levels of FR wavelengths. To control for the morphogenic effects of irradiance, neutral-shade cloths were placed over seedling trays in the low R : FR treatment. Photosynthetically active radiation and R : FR in the two light treatments were measured using a LI-COR-1800 Spectroradiometer (LI-COR, Inc., Lincoln, Nebraska, USA). Both light treatments reduced PAR by 60% of light in the greenhouse to 150 µmol·m–2·sec–1. Under the copper sulfate solution R : FR = 13, whereas initial R : FR equaled 1.25 (similar to natural sunlight) under the neutral-shade cloths. In a comparable experiment with Impatiens, R : FR below the seedling canopy was reduced only to 0.91 in the copper-sulfate treatment, while R : FR was lowered to 0.23 in the neutral-shade controls (as cited in Dudley and Schmitt, 1996 ). Studies in several plant species have shown that R : FR within a stand ranges from 1.1 to 0.2 and decreases with increasing neighbor proximity and density (Ballaré, Scopel, and Sanchez, 1990 ; Smith, Casal, and Jackson, 1990 ). A barrier of aluminum foil was placed around each of the experimental light treatments to minimize edge effects and to prevent nonfiltered light from shining on the plants. Because we varied R : FR while holding PAR constant, phenotypic differences observed among experimental light treatments reflect the effects of light quality rather than levels of PAR.

Seedling trays remained in the R : FR treatments for a total of 3 wk, at which point hypocotyl, first internode length, total height, and the length of the longest leaf were measured. A subsample of plants from both populations within each R : FR treatment (2 R : FR treatments x 2 populations x 10 plants; N = 40) was destructively harvested to measure leaf flavonoid and anthocyanin content. Following the census at 3 wk, some of the remaining experimental plants (N = 144) were transplanted into 10-cm pots filled with Metromix 350 and placed under the UV treatments.

Ambient UV and UV-removal treatments
The experimental UV treatments were set up in three plots outside the Brown University greenhouse. Each plot contained an ambient-UV and a UV-removal treatment. To establish the ambient-UV treatment, plastic, Acrylite OP-4 panels (Cryo Industries, Woodcliff Lake, New Jersey, USA) were attached to polyvinyl chloride (PVC) frames. The OP-4 filter transmits 80% of incident light at 300 nm and 89% of light at 380 nm, the upper end of the UV region. The UV-removal treatment was set up using a similar arrangement of Acrylite OP-3 panels (Cyro Industries), which transmit <1% of light at wavelengths lower than 380 nm. The UV-removal treatment therefore reduces both UV-A and UV-B. Recent studies examining the effects of global change have attempted to exclude UV-B without altering levels of UV-A, because it is the shorter, UV-B wavelengths that are most damaging and most relevant in determining the effects of ozone depletion (Caldwell, 1979 ). In addition, plants are known to respond both to UV-A and UV-B independently and to the relative proportion of the two (Middleton and Teramura, 1993 ; Krizek, Britz, and Mirecki, 1998 ). The broad, UV-removal treatment used here is appropriate to test our experimental questions of opportunity costs and differences in UV susceptibility between sun and shade populations because plants from different canopy environments will experience decreases in both the UV-A and -B regions of the spectrum. Neither of the two filters absorbs wavelengths outside the UV region. Photosynthetically active radiation and R : FR were therefore equivalent to natural, sunlit conditions.

The UV treatments were maintained for 4 wk, at which point the lengths of the first five internodes, total height, number of nodes, number of branches, number of flowers, number of pedicels, number of fruits, and longest leaf length were recorded for each experimental plant. The total number of pedicels, fruits, and flowers was used to estimate total reproductive output. Plants were then harvested, dried at 40°C for 5 d, and weighed to determine aboveground biomass. The value calculated for reproductive output reflects only early reproductive effort rather than lifetime fecundity, because the experiment was concluded prior to natural senescence of the plants. Approximately 1–2 g of leaf material (petiole and leaf blade) were removed from each plant at the time of harvest to estimate flavonoid and anthocyanin levels.

Anthocyanin and flavonoid extraction
Plant material was ground in liquid nitrogen and placed in a –80°C freezer. Samples were then removed successively from the freezer to complete the anthocyanin and flavonoid measurements. One gram of ground, thawed leaf material was placed in test tubes containing 3 mL of extraction buffer (81% distilled water, 18% methanol, and 1% glacial HCl by volume). Samples were then boiled for 3 min in a hot water bath and incubated in the dark at room temperature for 24 h.

Following the 24-h incubation, samples were centrifuged at 6000 g and 4°C for 40 min. The supernatant of each sample was pipetted into a spectrophotometer cuvette transparent to light at wavelengths >285 nm (Fisher Scientific, Springfield, New Jersey, USA). The samples were scanned for absorbances between 285 and 700 nm in a Spectronic 20 (Bausch and Lomb, Rochester, New York, USA). Anthocyanin contents were determined by scanning the 0.33 g/mL samples, and absorbance values were calculated as Absorbance535 = Absorbance535 – 0.33Absorbance650 to correct for absorbance by chlorophyll, following Lindoo and Caldwell (1978). Samples were further diluted to 0.005 g/mL to measure levels of compounds absorbing exclusively in the UV range, because absorbance values of the undiluted samples were too high to provide accurate estimates of concentrations. Peaks in absorbance falling between 280 and 380 nm were attributed to the presence of flavonoids or related phenolic compounds, because these substances are known to absorb in the UV region and because the extraction process used here selectively isolates these compounds (Wellmann, 1971 ; Robberecht and Caldwell, 1983 ; Mirecki and Teramura, 1984 ; Li et al., 1993 ).

Effects of the R : FR pretreatment on hypocotyl length and longest leaf length were determined using MANOVA with R : FR and population as fixed effects. Hypocotyl was natural-log transformed to reduce heteroscedasticity. No analyses were performed for the concentrations of anthocyanins and flavonoids measured following the R : FR pretreatment, because seedlings failed to produce detectable levels of either compound. Traits measured after the UV treatment (morphological characters, biomass, total reproductive structures, flavonoid concentrations, and anthocyanin content) were analyzed using MANOVA with UV, R : FR treatment, population, and plot(UV) as main effects and population x UV, population x R : FR, R : FR x UV and UV x POP x R : FR as interaction effects. The data were analyzed in a split-plot design with UV tested over plot(UV). Univariate tests were then performed for individual traits to examine the source of significant effects in the MANOVA. Branch number was natural log-transformed, leaf length and biomass were squared, and total reproductive output was square-root transformed to meet assumptions of ANOVA. All analyses were carried out with SPSS 8.0 (SPSS, 1998 ).

To test whether UV-mediated decreases in height were beneficial in the ambient-UV treatment, we performed an ANCOVA for reproductive output with R : FR, population, UV, and plot(UV) as class variables and final leaf size and height as covariates. The interaction of height x UV was used to test whether the relationship between height and reproductive output differed between the UV treatments. To examine the relationship between fitness and leaf anthocyanin and flavonoid concentrations in the UV-removal and ambient-UV treatments, we performed ANCOVAs for biomass accumulation including R : FR and population as class variables and final leaf size, height, anthocyanin levels, and flavonoid concentrations as covariates. Leaf size and height were included covariates to control for the effects of individual plant size. ANCOVA was chosen rather than simple regression, because the populations proved to differ in many of the focal traits (e.g., reproductive output, height, anthocyanin levels, flavonoid concentrations). ANCOVA factors out population differences and differences due to seedling R : FR, which could otherwise contribute to the relationships being tested (e.g., Dudley and Schmitt, 1996 ).


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 LITERATURE CITED
 
Seedlings in the low R : FR pretreatment had significantly longer hypocotyls and leaves on average than those under the high R : FR pretreatment (Table 1; Fig. 1). The elongation response appeared to start prior to canopy closure when overhead R : FR experienced by the seedlings was still close to initial values. Because the initial, overhead R : FR in both treatments was in a range considered unlikely to elicit shade-avoidance responses (Smith, 1982 ), the early elongation of seedlings in the low R : FR treatment is probably attributable either to strong responses to FR reflected from neighbors or to a cryptochrome-mediated response to low PAR that was overridden by phytochrome-mediated pathways under high R : FR. Seedlings of the clearing site had shorter hypocotyls and smaller leaves than those from the woodland (Fig. 1). There was a population x R : FR treatment interaction for hypocotyl length (Table 1). Hypocotyl lengths of the clearing population were only half that of the woodland population when plants were grown under high R : FR, whereas hypocotyl lengths were approximately equal under the low R : FR treatment (Fig. 1). This result indicates that the clearing population was more responsive to low R : FR than the woodland population.


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  		Table 1. (A) MANOVA and (B) univariate ANOVA for the effects of seedling R:FR pretreatment on leaf size and hypocotyl elongation

 


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  		Fig. 1. Effects of R : FR and population on (A) hypocotyl elongation and (B) leaf length. Means + 1 SE are shown

 
Following the 4-wk UV exposure, plants grown in the low R : FR treatment as seedlings were taller, tended to have less biomass (Table 2A), and produced fewer branches and nodes than plants initially experiencing high R : FR (Table 2B; summarized in MANOVA Tables 3 and 4). Plants initially under low R : FR also had smaller leaves at harvest than those under high R : FR (Table 2A); the effect of R : FR therefore reversed between the pretreatment and final censuses. This reduction in leaf size suggests a cost to shade-avoidance responses in terms of subsequent leaf expansion under direct sunlight. The R : FR pretreatment had no effect on reproductive output, and, again, concentrations of anthocyanin and flavonoid levels were unaffected by R : FR (Table 2A).


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  		Table 2. (A) Effects of R:FR on architecture, growth rate, leaf compounds, and reproductive output as measured when plants were 7 wk of age (following the UV treatment). Means are shown with 95% confidence intervals. Values were back-transformed where necessary. (B) Effects of R:FR and UV on branch number and node number

 

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  		Table 3. MANOVA for effects of R:FR, UV, and population on traits measured following the 4-wk UV treatment (N = 144)

 
In comparison to plants grown in the UV treatment, plants grown in the absence of UV were taller, had larger leaves, accumulated more biomass (Table 5), and tended to initiate a greater number of branches and nodes (Table 2B). Plants growing under UV had higher leaf concentrations of both anthocyanins and flavonoids in comparison to plants in the UV-removal treatment (Table 5).


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  		Table 5. Effects of UV on architecture, growth, reproductive output, and leaf compounds. Means are shown with 95% confidence intervals in parentheses. Values were back-transformed where necessary

 
The effects of UV on node production and branch number were rendered nonsignificant when biomass was included as a covariate in an ANCOVA of UV and plot(UV) (F1,134 = 2.28, P = 0.13 and F1,134 = 0.63, P = 0.43, respectively), demonstrating that differences in resource assimilation and plant size mediate the effects of UV on these architectural traits. The effects of UV on height, however, remained significant in a similar ANCOVA (F1,134 = 11.43, P = 0.001), suggesting that UV-mediated decreases in elongation are a developmental response rather than a simple allometric one. An ANCOVA was performed to test the effects of UV-mediated decreases in height on reproductive output. The ANCOVA included population, R : FR, UV, and plot(UV) as class variables and leaf size and height as covariates. The interaction of UV x height was marginally significant (F1,133 = 3.70, P = 0.06). Regressions of the model residuals on height demonstrated that the relationship between reproductive output was negative in the UV environment and positive in the non-UV environment, but neither of the regressions was significant (UV removal: ß = 0.10, F1,70 = 0.64, P = 0.43; ambient UV: ß = –0.18, F1,70 = 2.38, P = 0.13). Thus, there is no evidence that UV-mediated decreases in height are adaptive.

Interactions of the light treatments (UV x R : FR) were nonsignificant (Table 3). However, given the small sample sizes of this experiment, the statistically marginal effect of UV x R : FR on branch number and node number (Table 4) may be biologically meaningful. The detrimental effects of UV exposure on branching and node production were more pronounced among individuals initially exposed to high R : FR (Table 2B).


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  		Table 4. Univariate tests for effects of R:FR, UV, and population on individual traits measured after the 4-wk UV treatment. Mean square values are shown

 
The clearing and woodland populations differed morphologically, as observed in previous studies (Schmitt, 1993 ; Dudley and Schmitt, 1995 ). Individuals derived from the clearing population were shorter, produced smaller leaves, initiated more branches, had fewer nodes, had lower levels of both flavonoids and anthocyanins (Table 6), and exhibited lower reproductive output (Fig. 2) than the woodland population.


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  		Table 6. Effects of population on phenotype traits measured at 7 wk of age (following the UV treatment). Means are shown with 95% confidence intervals. Values were back-transformed where necessary

 


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  		Fig. 2. Population norms of reaction for reproductive output (the number of reproductive structures present at the time of harvest) under the two UV treatments. Back-transformed means ± approximate 95% confidence intervals are shown

 
Populations differed significantly in sensitivity to UV (summarized in MANOVA Tables 3 and 4). Total reproductive output was substantially decreased among individuals from the woodland population growing in the UV relative to the UV-removal treatments, whereas reproductive output of the clearing population was unaffected by the UV environment (Fig. 2).

Independent of population effects, higher concentrations of anthocyanins were associated with decreased biomass among plants in the ambient-UV environment (Fig. 3; Table 7). However, ANCOVA testing for heterogeneity of slopes failed to detect a significant difference between the two UV treatments (F1,133 = 0.497, P = 0.482).



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  		Fig. 3. Relationship between anthocyanins and fitness (estimated by biomass accumulation) in the ambient-UV and UV-removal treatments. A significant effect of anthocyanins was detected in the ambient-UV treatment but not in the UV-removal treatment (Table 7 ). Values for biomass and anthocyanins are plotted to show the direction of the relationship. Plots of the residuals gave similar results

 

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  		Table 7. ANCOVAs examining the relationship between biomass and anthocyanin and flavonoid concentrations in the UV-removal and ambient-UV treatments

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
LITERATURE CITED
 
Exposure to UV radiation affected plant morphologies and individual fitness, but there was little evidence that the responses were adaptive. Differences in morphology observed between the ambient-UV and non-UV environments appeared to result from the stressful effects of exposure to UV; plants in the UV treatment were smaller than those grown in the absence of UV, and these differences in size explained most morphological differences. There were also several developmental responses to UV; plants exposed to UV were less elongated for their size than those not exposed to UV and had higher levels of anthocyanins and flavonoids. Contrary to prior studies (Takahashi, Takeda, and Ohnishi, 1991 ; Li et al., 1993 ; Middleton and Teramura, 1993 ; Teramura and Sullivan, 1994 ; Burger and Edwards, 1996 ), the results did not show that higher levels of these compounds enhanced growth or fitness under UV radiation. Population differences in fitness did, however, suggest evolutionary responses to UV exposure; reproductive output of the woodland population was more detrimentally affected by UV than that of the clearing population, which has an history of exposure to UV. Finally, pretreatment with low R : FR did not affect susceptibility to UV damage as estimated by biomass accumulation and reproductive output. There is no evidence, therefore, that increased susceptibility to UV poses an opportunity cost to shade-avoidance responses.

Shade-avoidance responses illustrate how photosensory systems enable plants to detect and adaptively respond to environmental conditions (Smith, 1982 ; Smith, Casal, and Jackson, 1990 ; Schmitt and Wulff, 1993 ; Smith and Whitelam, 1997 ); the phytochrome photoreceptors perceive R : FR cues indicative of future competition and elicit competitive, stem-elongation responses. Evidence for an unidentified UV photoreceptor exists, which suggests that plants can detect levels of UV radiation and may respond in a manner that would minimize susceptibility to damage (Wellman, 1983 ; Ballaré, Barnes, and Kendrick, 1991 ; Ballaré, Barnes, and Flint, 1995 ; reviewed in Greenberg et al., 1997 ). Exposure to UV radiation stimulated the production of both anthocyanins and flavonoids, a developmental response hypothesized to be adaptive. However, we found no evidence that either flavonoids or anthocyanins conveyed a benefit in the ambient-UV relative to non-UV environment, as would be expected if UV were a selective agent (Wade and Kalisz, 1991 ). Flavonoids were uncorrelated with fitness, and higher levels of anthocyanins were associated with decreased biomass in the UV environment. Moreover, if the compounds acted to reduce susceptibility to UV damage, we would expect higher average concentrations in the population that typically experiences UV, that is the clearing population, but the opposite pattern was observed. Anthocyanins induced by the UV exposure appear to be a response to stressful growth conditions, rather than an adaptive response. The results of previous studies have given mixed support of the UV-protection hypothesis for anthocyanins; some have found that anthocyanins reduce the detrimental effects of UV (Takahashi, Takeda, and Ohnishi, 1991 ; Burger and Edwards, 1996 ), whereas others suggested little advantage to anthocyanins in UV environments (Woodall and Steward, 1998 ).

In addition to UV, high PAR prevailing in the field treatments may have stressed the plants. Although the negative regression of biomass on anthocyanins was significant only in the ambient-UV treatment, a heterogeneity of slopes' test failed to distinguish between the ambient-UV and UV-removal treatments. The similar, negative relationship across the two treatments suggests that there is a stress common to the two treatments. High PAR is a likely candidate because the transition from the greenhouse (R : FR pretreatments) to the field (UV treatments) involved a substantial increase in PAR, from 150 to 1500 µmol·m–2·sec–1. If anthocyanin induction is a stress response, then high PAR could also account for the observed population differences in anthocyanin concentrations; high PAR elicits the production of anthocyanins (Mirecki and Teramura, 1984 ; Gong et al., 1997 ), and the woodland population with little history of exposure to high irradiance may have been more stressed and produced more anthocyanins than the clearing population when moved to the high-light, field treatments. Regardless of whether high PAR or high UV underlie anthocyanin production, the negative relationship between biomass and anthocyanins indicates that the pigment does not reduce environmental light stress.

The morphological responses to UV observed in this study appear to reflect the detrimental effects of UV stress on growth. The reduction in node and branch number observed among individuals in the UV relative to non-UV environments were explained entirely by the smaller size of these plants. Consistent with other studies (Ballaré et al., 1995 ), ambient UV suppressed stem elongation in I. capensis. The observed reduction in height relative to biomass might be a developmental response mediated by an unknown UV photoreceptor. However, decreased height was uncorrelated with reproductive output in the ambient UV treatment, suggesting that the response is not an adaptive one.

The clearing and woodland populations differed in their responses to both UV and R : FR. Most notably, UV had a more detrimental effect on fruit production in the woodland than in the clearing population. The clearing population also showed greater responsiveness of internode elongation to low R : FR, a finding consistent with prior observations (Dudley and Schmitt, 1995 ; Donohue and Schmitt, 1999 ). Population differences in total reproductive output probably result from the characteristic population differences in timing of reproduction and from the comparatively early harvest of this experiment; the woodland population typically flowers earlier than the clearing population (Schmitt, 1993 ) and is likely to have initiated more flowers by the conclusion of the experiment. Had the experiment continued through natural senescence, the clearing population probably would have shown greater total reproductive output, due to its greater branch number and higher number of meristems available to produce flowers (Schmitt, 1995 ).

Consistent with other studies, exposure to UV affected the expression of many phenotypic traits, including fitness. Populations from sun and shade environments differed in their responses to UV exposure, suggesting that mechanisms to minimize the damaging effects of UV have evolved. The nature of these mechanisms, however, was unclear; none of the traits measured here appeared to mitigate UV damage. Relative to plants experiencing high R : FR, early exposure to low R : FR had no effect on subsequent biomass accumulation or reproductive output. Thus, there is no evidence that UV susceptibility constitutes an opportunity cost to elongation responses that might limit the evolution of shade-avoidance genotypes.


   FOOTNOTES
 
1 The authors thank Fred Jackson for construction of the UV treatment plots and care of the experimental plants, Shane Heschel and Neil Hausmann for help with the UV extractions, and Kathleen Donohue for discussions regarding the experimental design. This research was funded by a UTRA grant to P. D. and NSF grant DEB9806858 to J. S. Back

2 Author for reprint requests (tel: 401-863-2897; fax: 401-863-2166; cweinig{at}brown.edu ). Back


   LITERATURE CITED
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 LITERATURE CITED
 
Bain B. T. H. Attridge 1988 Shade-light mediated responses in field and hedgerow populations of Galium aparin. Journal of Experimental Botany 39: 1759-1764[Abstract/Free Full Text]

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