costs of canalization and plasticity in response to neighbors in brassica rapa

Plant Species Biology (2002) 17, 109–118

© 2002 The Society for the Study of Species Biology

Blackwell Science, LtdOxford, UKPSBPlant Species Biology1441 0745The Society for the Study of Species Biology, 2002

17

2 & 3December 2002

081COSTS OF PLASTICITY and CANALIZATION

J. POULTON and A. A. WINN10.1046/j.1441 0745.2002.00081.x

Original ArticleBEES SGML

Correspondence: Alice A. Winn Email: [email protected]

Costs of canalization and plasticity in response to neighbors in

Brassica rapa

JOHN POULTON and ALICE A. WINN

Department of Biological Science, Florida State University, Tallahassee, FL 32306-1100, USA.

Abstract

Variation in patterns of phenotypic plasticity reflects a balance between selection toproduce the optimum phenotype in all environments and forces that limit the evolutionof such ideal plasticity. Theoretical models indicate that a cost of plasticity or canalizationcould counter selection favoring ideal plasticity, but there are few empirical data eitherto support or to refute the existence of such costs. The adaptive significance of plasticityor canalization for plastic responses to the presence of neighbors in an annual plant weredetermined and the costs of plasticity and canalization were quantified. Representativesof 24 paternal half-sibling families of

Brassica rapa

were grown with and without neigh-bors and the plasticities in six traits were measured. Canalization rather than plasticitywas favored by selection by four of the five traits for which there was any selection onplasticity. Of these four, one trait exhibited adaptive canalization and the other threeshowed maladaptive or nonadaptive plasticity. Adaptive plasticity in elongation of thefirst internode was favored, but the observed pattern of plastic response was maladaptive.We found evidence for costs of both plasticity and canalization for some traits, supportingthe plausibility that costs contribute to variation in patterns of phenotypic plasticity.Several studies now support the influence of costs on the evolution of plastic responses,but results do not rule out a role for other factors that limit the evolution of responses tothe environment. A dearth of empirical data continues to limit our understanding of thebasis for variation in patterns of phenotypic plasticity in nature.

Keywords:

adaptive value of plasticity, competition, flowering time, genetic variation for plas-ticity, stem elongation.

Received 15 August 2001; revision received 25 April 2002; accepted 10 May 2002

Introduction

The frequency and magnitude of plasticity in plants hasbeen widely recognized and well documented (reviewedby Bradshaw 1965; Schlichting 1986; Sultan 1987; Travis1994). Patterns of plastic response to the environment dif-fer among traits of the same species, among differentenvironments and among species. Some patterns of plas-ticity reflect adaptive responses to a variable environmentwhere selection favors different values of the same traitunder different environmental conditions (e.g. Dudley &Schmitt 1996; Donohue

et al

. 2000b). Although recentstudies of phenotypic plasticity have focused primarilyon adaptive plasticity, many patterns of plasticityobserved in nature may not be adaptive. Some plasticresponses probably reflect unavoidable consequences of

growing at different resource levels, rather than adap-tive responses to predictable environmental variation(Donohue & Schmitt 1999; Winn 1999; Dorn

et al

. 2000). Ifthe optimum value of a phenotypic trait is the same indifferent environments, then selection may favor canali-zation of the trait rather than plasticity. Plasticity underthese conditions would be maladaptive.

In the absence of evolutionary limits on the evolutionof responses to the environment, species would evolve toproduce the ideal phenotype in all possible environments.If no limits existed, plasticity would never be maladaptiveand all plastic responses would be ideal. Patterns of plas-ticity that we actually observe are determined by a bal-ance between selection favoring plasticity or canalizationand the forces that limit the evolution of trait responsesto the environment. Differences in the point of balancewill contribute to observed variation among traits, envi-ronments and taxa in patterns of plasticity. Consequently,

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an understanding of the forces that limit the evolutionplant responses to the environment will help elucidate thebasis for the diversity of patterns of phenotypic plasticityfound in nature.

One force that could counter selection favoring adap-tive plasticity or canalization is an evolutionary cost. Justas an evolutionary increase in investment in reproductionmay have an associated cost in terms of future survivaland reproduction (Reznick 1985, 1992), the evolution ofincreased plasticity or canalization of a trait may carry afitness cost that counters the benefits of being able toproduce the ideal trait value within each of all possibleenvironments (van Tienderen 1991). A cost of plasticitycould explain why observed patterns of plasticity are notideal, because if such a cost exists, then the pattern ofplasticity that maximizes genotypic fitness will reflect acompromise between always producing the ideal pheno-type and suffering a prohibitive cost. Similarly, maladap-tive plasticity in a trait might persist even when selectionfavors canalization if increased canalization has an evolu-tionary cost.

The consideration of costs in many models of theevolution of adaptive plasticity (Via & Lande 1985; vanTienderen 1991; Clark & Harvell 1992; Gomulkiewicz &Kirkpatrick 1992; Moran 1992; Newman 1992; Scheiner1993) may reflect the expectation that plasticity must becostly and that costs are a likely candidate for the factorthat limits the evolution of phenotypic plasticity (seeSultan 1992 for an opposing view). To date, however, littleif any empirical evidence supports the existence of costsof plasticity. Donohue

et al

. (2000a) found no evidence fora cost of adaptive plastic stem elongation in response tocompetitors for the annual plant

Impatiens capensis

. Simi-larly, two analyses of the costs of induced defenses inanimals failed to find convincing evidence for a cost ofplasticity (DeWitt 1998; Scheiner & Berrigan 1998). vanKleunen

et al

. (2000) did find evidence for a cost of plas-ticity in internode length in the perennial plant

Ranuncu-lus reptans

, but plasticity in this trait was not adaptive. Inthe only study to date to quantify costs of canalization,Dorn

et al

. (2000) examined costs of both plasticity andcanalization of traits of

Arabidopsis thaliana

grown at twodifferent densities. They found evidence for costs of canal-ization for leaf length and number of inflorescencebranches but concluded that the small magnitude of thesecosts suggests that they have little biological importance.

Overall, the limited set of empirical results concerningcosts of plasticity and canalization do not provide strongevidence that costs are an important force shaping pat-terns of plasticity, but neither do they convincingly refutethe possibility. If costs turn out to be rare or insufficientto explain observed patterns of plasticity, we will need tobroaden our exploration of the forces that limit the evo-lution of plasticity. As is true for the evolution of any trait,

lack of appropriate genetic variation could also limit theevolution of plasticity or canalization. In addition, recentmodels show that limited reliability of the cues organismsuse to predict future selective environments can have thesame effects as costs on the evolution of plasticity (de Jong1999; Tufto 2000). We need additional empirical estimatesof costs to determine whether costs are important deter-minants of patterns of phenotypic plasticity or whetherwe should focus on other forces that might be moreimportant in shaping the evolution of plasticity.

Here we present an analysis of plastic responses of anannual plant to neighbor density for traits whose plastic-ity was expected to range from adaptive to maladaptive.We measured plasticity in stem elongation, which hasbeen shown to be an adaptive response to competition inseveral plant species (Schmitt

et al

. 1995; Dudley &Schmitt 1996; Weinig 2000). We also measured severalsize-related traits for which plasticity seems likely to bemaladaptive (Donohue & Schmitt 1999). We measuredspecific leaf weight, expecting that it would be plastic butthat the magnitude of plasticity might be too small to havefunctional consequences (Winn & Evans 1991). For eachtrait, we used the pattern of selection in environmentswith and without neighbors and the observed pattern ofplastic response to diagnose the adaptive significance ofplasticity. We also tested the hypothesis that plasticity orcanalization for each trait entails a cost. Finding signifi-cant costs of plasticity for adaptive plastic responses orcosts of canalization for maladaptive plastic responseswould be consistent with the hypothesis that costs con-tribute to the diversity of patterns of phenotypic plasticity.

Materials and methods

Brassica rapa

(Brassicaceae) is a self-incompatible, annualplant that exhibits strong phenotypic responses to compe-tition (Miller & Schemske 1990; Miller 1995). We obtaineda bulk collection of seeds of

B. rapa

from a natural popu-lation near Irvine, CA, USA. To provide genetic replicatesnecessary to measure plasticity, we executed a paternalhalf-sibling breeding design in the greenhouse. Four hun-dred individuals were grown and 24 were chosen at ran-dom to serve as sires. Each of these sires was crossed withthree randomly chosen dams. Although this species pro-duces perfect flowers, each individual was used only as adam or as a sire.

Eight seeds from each of the 72 full-sibling familiesgenerated by hand crosses were planted in each of twoenvironments. Each seed assigned to the no-neighborenvironment was planted alone in a 4-inch pot. Each seedassigned to the neighbor treatment was planted in thecenter of a 4-inch pot in which four additional (unrelated)seeds were planted, one in each corner of the pot. Plantswere grown in a standard greenhouse potting mix (Pro-

C O S T S O F P L A S T I C I T Y A N D C A N A L I Z AT I O N

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Mix) in a greenhouse at Florida State University’s MissionRoad Research Facility in Tallahassee, FL, USA. Pots werearranged in blocks on four greenhouse benches and wererepositioned randomly each week to minimize effects ofenvironmental variation. Natural light was supplementedby sodium halide lamps (14 h/day), which provided thelong-day cue necessary to induce flowering.

For each plant, the number of days between plantingand emergence was recorded and the leaves present attwo weeks of age were counted. The date on which eachplant began flowering was also recorded and once flow-ering began, open flowers were randomly cross-pollinatedevery second day with a feather duster (Stanton

et al

.2000). Plants were harvested when they began to senesce.

At the time of harvest, the first internode was mea-sured, dried to a constant weight and weighed to thenearest 0.001 mg. Elongation of this internode was calcu-lated as its length divided by its dry mass. The area of thelargest leaf produced by each plant was also measured.This leaf was dried and weighed, and its specific leafweight was calculated as leaf dry mass divided by leafarea. The height of each plant was measured and the totalnumber of flowers produced was determined at the timeof harvest.

Determining the adaptive significance of plasticity

Diagnosis of the selective value of phenotypic plasticityin a trait requires knowledge of both the pattern of selec-tion on the trait within each environment and theobserved pattern of plasticity (cf. Dorn

et al

. 2000). If thereis directional selection on a trait in two environments, andthe signs of the selection coefficients are opposite, thenselection favors adaptive plasticity. If the observed plasticresponse of the trait is in the correct direction (e.g. theresponse is to increase the trait value in the environmentwhere an increase is favored), then the observed patternof plasticity is adaptive. If there is no directional selectionon the trait in either environment, then plasticity is non-adaptive regardless of the pattern expressed.

If there is selection on a trait in only one environment,there may or may not be selection on plasticity in the trait.In this case, the selective value of plasticity can be diag-nosed from a regression of family mean relative fitness inthe two environments on plasticity calculated as the dif-ference in family mean trait expression between the twoenvironments. A significant regression would indicateadaptive plasticity if the response matched the patternof selection (e.g. the trait value was greater in the envi-ronment where selection favors a greater value) andmaladaptive plasticity if the direction of response isopposite to the pattern of selection. A non-significantregression would indicate that the observed plasticity isnonadaptive.

If there is significant directional selection on a trait intwo environments and the signs and magnitudes of theselection coefficients are the same, then fitness would bemaximized by maintenance of a constant appropriate traitvalue. Under these conditions, selection favors canaliza-tion rather than plasticity, so any plasticity observed islikely to be maladaptive. Selection against plasticitywould be confirmed by a significantly negative regressionof mean genotypic fitness in the two environments onplasticity. If no genotype is canalized for the appropriatelymaximal or minimal value in the two environments, plas-ticity could be adaptive. This diagnosis would be con-firmed by a positive regression of mean relative fitness onplasticity. If the two environments differ in magnitude ofselection, the diagnosis of the adaptive significance ofplasticity would depend on the relative frequency of alter-nate environments and the magnitude of the differencebetween the selection coefficients (see Via & Lande 1985;van Tienderen 1991).

Statistical analyses

We performed a mixed-model nested

ANOVA

for eachtrait, including the effects of Environment (fixed), Sire(random) and Dam nested within sire (random). For eachtrait, the presence of plasticity is indicated by a significanteffect of Environment. A significant Sire–Environmentinteraction indicates the presence of additive genetic vari-ation for plasticity. To test for the presence of additivegenetic variation within each environment, we conducteda nested, random-effects

ANOVA

for each trait within eachenvironment. The main effects in these analyses were Sireand Dam nested with Sire. A significant Sire effect indi-cates the presence of additive genetic variation for a trait.

We quantified the selective value of each trait in eachenvironment by regressing half-sibling family mean rela-tive fitness on the family mean for each trait value.Regression coefficients from these analyses indicate themagnitude and direction of directional selection on eachtrait in each environment (Lande & Arnold 1983). We usedthe number of flowers produced, which is strongly, posi-tively related to biomass in this species (Miller 1995), asour estimate of fitness. Relative fitness was calculated foreach individual within an environment as the total num-ber of flowers produced divided by the mean number offlowers produced by all individuals in that environment.

We determined relationships between traits andrelative fitness from family means rather than individualphenotypic values to avoid possible bias due to environ-mental covariance (Rausher & Simms 1989; Rausher1992). Because of the much smaller number of familiesthan of individuals, the analysis of family means hasconsiderably less power than the analysis of phenotypicvalues. For this reason, we also conducted analyses

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describing relationships between traits and relative fitnessusing phenotypic values. These results are reported in thetext in cases where statistical significance of coefficientsfrom the family means analysis was marginal.

When evidence of selection on a trait was confined toone environment, we regressed mean family relative fit-ness for the two environments on plasticity calculated asthe difference between the family mean values of the traitexpressed in the two environments. The sign of the regres-sion coefficient for plasticity in this analysis indicates theselective value of plasticity. When the coefficients from theregression of relative fitness on a trait in the two environ-ments were both significantly different from zero and ofthe same sign, we tested the hypothesis that the two coef-ficients were of significantly different magnitudes usingan analysis of covariance. Relative fitness was the depen-dent variable, environment was the class variable and thetrait was treated as a covariate. A significant interactionbetween the trait and environment in this analysis indi-cates that the two environments differ in the magnitudeof the effect of the trait on fitness.

Regression coefficients from the univariate regressionof relative fitness on each trait describe the relationshipbetween the value of the trait and relative fitness, includ-ing both the direct effect of the trait on relative fitness andindirect effects by way of any correlated traits (Lande &Arnold 1983). We diagnosed the adaptive significance ofplasticity in a trait on the basis of these values, whichreflect the net force of directional selection on the traits.We also conducted a multiple regression of relative fitnesson the set of traits measured within each environment toreveal the role of covariances among traits in determiningpatterns of selection. As an aid to interpreting indirectselection through trait covariance, we also estimatedgenetic correlations between traits within each environ-ment from Pearson product moment correlations amonghalf-sibling family trait means.

Costs of plasticity and canalization

The regression analysis described by van Tienderen (1991;cf. DeWitt

et al

. 1998; Scheiner & Berrigan 1998) was usedto test for costs of plasticity and canalization of traits ineach environment. In this analysis, family mean relativefitness within one environment was regressed on the fam-ily mean trait value and the plasticity in the trait mea-sured as the difference between environments in thefamily mean value of the trait. The regression coefficientfor plasticity measures the relationship between the mag-nitude of plasticity and relative fitness in one environ-ment, taking into account the expressed value of the traitin that environment. This coefficient will be negative iffamilies that exhibit the greatest degree of plasticity havelower fitness than less plastic families. A coefficient for

plasticity that is significantly less than zero indicates acost of plasticity in the trait. A coefficient for plasticitysignificantly greater than zero indicates a cost of canaliza-tion of the trait (cf. Dorn

et al

. 2000).All statistical analyses were performed with

SAS

version 8.1 (SAS Institute; Cary, NC). The assumptionof normal distribution of data was evaluated byKolmogorov–Smirnoff tests. The violation of normalitywas detected for time to flowering and for internodeelongation in the environment without neighbors.Analyses conducted with and without log transformationof these variables produced qualitatively similar results.We present the results of analyses of untransformeddata because they are easier to interpret. All other traitdistributions met the assumption of normality.

Where necessary, we tested homoscedasticity usingLevene’s procedure. Variance heterogeneity was detectedfor internode elongation and number of flowers pro-duced. Log transformation was successful in makingvariances homogeneous. Results of analyses of log-transformed data were qualitatively similar to those fromuntransformed data, so results for untransformed data arepresented for ease of interpretation.

We used a sequential Bonferroni procedure (Holm1979) to correct significance values for multiple tests ofsignificance for correlations among traits and for the coef-ficients that reflect costs of plasticity or canalization. Forthese analyses, we present both the original significancelevels and the Bonferroni corrected results.

Results

Plasticity and additive genetic variation

All traits showed significant plasticity except time to flow-ering, in which the two environments differed by less thana day (Table 1). Plants grown with neighbors emergedslightly earlier and produced significantly fewer leavestwo weeks after emergence than those grown alone.Plants with neighbors also exhibited nearly fourfoldgreater elongation of the first internode than those grownwithout neighbors. At the time of harvest, plants grownwith neighbors were significantly shorter, produced onlyone third as many flowers and produced leaves more than20% thinner than those of plants grown without neigh-bors (Table 1). Plant survival to harvest was high (90%)and equivalent in the two environments.

Significant additive genetic variation was detected inboth environments for elongation of the first internode,time to flowering and number of flowers produced(Table 2). For height at harvest and number of leaves attwo weeks, we found significant additive genetic varia-tion in one environment and marginally significant vari-ation in the other. There was additive genetic variation for


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specific leaf weight in the environment with no neighborsonly. No additive genetic variation for time to emergencewas detected in either environment (Table 2).

Additive genetic variation for plasticity was detectedfor internode elongation, number of leaves at two weeks(

P

=

0.054), height and number of flowers produced(Table 2). We detected no significant additive genetic vari-ation for plasticity in time to emergence, days to flower-ing, or specific leaf weight (Table 2).

Adaptive significance of plasticity

Detection of a relationship between family mean relativefitness and family mean trait values within each envi-ronment requires variation in both relative fitness (esti-mated from number of flowers produced) and theexpressed value of the trait. Sire had no significant effectin one environment on the number of leaves at twoweeks or specific leaf weight; it had no significant effecton time to emergence in either environment (Table 2). Weproceeded with analysis of selection for these traits

despite the lack of evidence of additive genetic variation,because the regression analysis could be more sensitivethan the

ANOVA

and might detect a relationship eventhough the

ANOVA

could not.The analysis of selection indicated that the earliest-

emerging plants had the greatest relative fitness in bothenvironments, though selection was only marginally sig-nificant in the absence of neighbors (Table 1). The twoenvironments did not differ in the magnitude of selection(

F

environment

¥

time to emergence

=

0.02,

P

<

0.20). Genotypes able tomaintain consistent early emergence in both environ-ments should be favored. In other words, canalizationrather than plasticity in this trait would be favored byselection. Although plants emerged significantly faster inthe environment with neighbors, the difference was a mat-ter of a few hours (Table 1) and plasticity had no signifi-cant effect on relative fitness (regression coefficient

=

0.20,

P

<

0.38). These results support the conclusion that plas-ticity in time to emergence was nonadaptive.

Elongation of the first internode was the only trait forwhich the analysis suggested selection favoring adaptive

Table 1

Means (standard deviations) for plant traits for replicates of 24 paternal half-sibling families grown with and without neighbors,and selection differentials for each trait in each environment.

Trait

Trait means Selection differentials Pattern favored by selection

Significance of plasticity

Noneighbors Neighbors

F

Noneighbors Neighbors

Days to emergence 3.34 (0.18) 3.23 (0.18) 6.8*

-

0.71

t

-

0.77*** canalization nonadaptiveElongation of first internode (cm/g) 6.45 (1.76) 24.82 (9.48) 116*** 0.11**

-

0.005 plasticity maladaptiveNumber of leaves at 2 weeks 5.64 (0.31) 4.72 (0.35) 207*** 0.38

t

0.35* canalization maladaptiveDays to flowering 33.96 (4.79) 33.73 (3.80) 0.32

-

0.68***

-

0.37** canalization adaptiveSpecific leaf weight (g/cm

2

) 5.53 (1.00) 4.25 (0.60) 76*** 0.10 0.027 none nonadaptiveHeight (cm) 102.4 (12.8) 81.8 (10.6) 55*** 0.007 0.013** canalization nonadaptiveNumber of flowers 336 (110) 103 (28) 111*** – – – –

t

0.5

<

P

<

0.10, *

P

<

0.05, **

P

<

0.01, ***

P

<

0.001.

F

-values are for the effect of Environment in

ANOVA

for effects of environment, sire,and dam nested within sire for each trait.

Table 2

Effects (

F

-values) of Sire from

ANOVA

for effects of sire and dam nested within sire for each trait in each environment and thesire–environment interaction for each trait from

ANOVA

including effects of environment, sire, dam nested within sire, and theirinteractions.

TraitSire effect

Sire–by–Environment interactionNo neighbors Neighbors

Days to emergence 0.62 0.88 0.81Elongation of first internode (cm/g) 3.87*** 1.76* 2.2***Number of leaves at 2 weeks

t

1.24 1.72 1.54*Days to flowering 4.4*** 3.22*** 1.21Specific leaf weight (g/cm

2

) 5.54*** 1.14 0.7Height (cm) 1.66

t

3.81*** 1.82*Number of flowers 4.1*** 2.72** 4.39***

t

0.5

<

P

<

0.10, *

P

<

0.05, **

P

<

0.01, ***

P

<

0.001

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, 17, 109–118

plasticity. Directional selection was significantly positivein the absence of neighbors and negative but not signifi-cant in the presence of neighbors (Table 1). In the pheno-typic analysis, directional selection on elongation wassignificantly negative in the environment with neighbors(selection differential

= -

0.078,

P

<

0.001). Despite selec-tion favoring adaptive plasticity, the observed plasticresponse in elongation was the opposite of the pattern ofselection. Plants were significantly less elongated in theabsence of neighbors (Table 1), that is, in the environmentin which greater elongation was favored by selection(Table 1). The observed pattern of plasticity in elongationwas therefore maladaptive.

For the number of leaves at two weeks, directionalselection was positive in both environments but statisti-cally significant only in the environment with neighbors(Table 1). In the analysis of selection on phenotypic val-ues, there was significant selection for plants with moreleaves in the no-neighbors environment (regressioncoefficient

=

0.31,

P

<

0.001). This result, along with theabsence of evidence for a difference between environ-ments in the magnitude of selection on the number ofleaves (

F

environment

¥

number of leaves

=

0.02,

P

<

0.88), supports theconclusion that selection would favor genotypes thatmaintained a large number of leaves in both environ-ments. Plants actually produced significantly fewer leavesin the environment with neighbors (Table 1). Theregression of mean family fitness on plasticity wassignificantly negative (regression coefficient

= -

0.30,

P

<

0.04) indicating that the observed pattern of plasticityin number of leaves at two weeks was maladaptive.

There was significant selection favoring earlier flower-ing both with and without neighbors (Table 1), and theenvironments did not differ in magnitude of selection(

F

environment

¥

time to flowering

=

2.23,

P

<

0.14). This pattern ofselection indicates that canalization of flowering timewould be favored. The absence of significant plasticity intime to flowering (Table 1) indicates that time to floweringexhibited adaptive canalization.

Specific leaf weight showed significant plasticity(Table 1) but had no effect on fitness in either environment(Table 1), and no significant relationship between familymean fitness and plasticity in specific leaf weight wasapparent (regression coefficient

= -

0.003,

P

>

0.07). Theseresults indicate that plasticity in specific leaf weight wasnonadaptive.

Selection favored taller plants in the environment withneighbors (Table 1). Selection on height in the no-neighbors environment was positive but not significant inthe analysis of family means, but it was significant in theanalysis of phenotypic values (regression coefficient

=

0.07,

P

<

0.001). The environments did not differ signifi-cantly in magnitude of selection (

F

environment

¥

height

=

0.8,

P

<

0.37), indicating that canalization of height was

favored. Plasticity of height was significant (Table 1), aswas variation among families in plastic response of height(Table 2), but family mean fitness and plasticity in heightshowed no relationship (regression coefficient

=

0.002,

P

<

0.58). Together these results indicate the observedplasticity in height has no consequences for fitness. Plas-ticity in height was therefore nonadaptive.

Multivariate regression analysis of the relationshipsbetween traits and relative fitness in the no-neighborsenvironment revealed significant direct selection on timeto flowering only (Table 3). The sign of this relationshipwas negative, as it was in the univariate analysis. In theanalysis for the competitive environment, time to flower-ing was again negatively related to relative fitness(Table 3), as it was in the univariate analysis (Table 1).Significant direct selection also favored taller plants in theno-neighbors environment (Table 3), as it did in theunivariate analysis (Table 1).

In the analysis of correlations among family means forthe no neighbors environment, elongation of the firstinternode and specific leaf weight were both negativelycorrelated with days to flowering (Table 4). Specific leafweight and height were positively correlated. In the com-petitive environment, elongation and specific leaf weightwere again negatively related to days to flowering, as wasthe number of leaves at two weeks (Table 4). After Bon-ferroni correction, only the relationships between elonga-tion and days to flowering and between specific leafweight and total height in the no-neighbors environmentwere significantly different from zero.


Four costs of plasticity or canalization were significantlydifferent from zero (Table 5). The significant positiveregression coefficient for plasticity of elongation in the no-neighbors environment indicates a cost of canalization inthis trait expressed in the absence of neighbors. A similar

Table 3

Coefficients from multiple regression of relative fitnesson plant traits (selection gradients) within each of twoenvironments

TraitNo

neighbors Neighbors

Days to emergence

-

0.002

-

0.355Elongation of first internode (cm/g)

-

0.079

-

0.007Number of leaves at 2 weeks 0.056 0.030Days to flowering

-

0.081***

-

0.050**Specific leaf weight (g/cm

2

) 0.015

-

0.103Height (cm)

-

0.0004 0.010**

**

P

<

0.01, ***

P

<

0.001


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, 17, 109–118

result for height suggests a cost of canalization in this traittoo. In the environment with neighbors, we found evi-dence of a cost of maladaptive plasticity in both the num-ber of leaves at two weeks and elongation. AfterBonferroni correction, only the cost of canalization formaladaptive plasticity in elongation and the cost of plas-ticity for maladaptive plasticity in the number of leavesat two weeks were significant (Table 5).

Discussion

Other than days to flowering, all traits of

Brassica rapa

thatwe measured exhibited plasticity in response to neigh-bors. The responses we observed are consistent with thosefrom other studies of the effects of density on plant traits(e.g. Dudley & Schmitt 1996; Donohue

et al

. 2000a,b, 2001;van Kleunen

et al

. 2000; Weinig 2000). Plants grown withneighbors were generally smaller and exhibited greaterelongation of the first internode than those grown alone(Table 1). The absence of plasticity in days to flowering isalso consistent with some previous studies. Dorn

et al

.(2000) found no effect of density on time to flowering inArabidopsis thaliana. Donohue et al. (2000a) found thatImpatiens capensis at high density flowered earlier thanplants at low density at a sunny location, but no effect ofdensity on time to flowering was apparent at a shadierwoodland site.

Although the plastic responses to density we observedare common to many species, none was found to be adap-tive (cf. Dorn et al. 2000). Selection favored canalizationrather than plasticity in four of the five traits for whichwe detected any selection on plasticity (Table 1).Observed patterns of response to neighbors in these traitsranged from adaptive canalization to both maladaptiveand nonadaptive plasticity. Selection did favor adaptiveplasticity in elongation of the first internode, but the pat-tern of plasticity actually expressed was maladaptive.

Selection favoring canalization combined with theabsence of plasticity in time to flowering indicates thatthis trait exhibited adaptive canalization. This is the samepattern reported by Dorn et al. (2000) for plastic responseof flowering time to density in Arabidopsis. Donohue et al.(2000a, b) report similar findings for Impatiens capensisgrown in the field at a woodland site, but they foundadaptive plastic acceleration in time to flowering inresponse to high density for the same species in a sunnylocation. It appears that both the pattern of plasticity intime to flowering and the adaptive significance of thisplasticity can differ among environments.

Selection also favored canalization of time to emer-gence in our study. This trait showed significant plasticity,but the magnitude of plasticity was very small (Table 1)and plasticity did not influence fitness. Although we clas-sified plasticity in time to emergence as nonadaptive, it

Table 4 Pearson product moment correlations among paternal half-sibling family means for traits of plants grown alone (above thediagonal) and with neighbors (below the diagonal)

A B C D E F

A. Days to emergence 1.0 -0.13 -0.16 0.36 0.11 -0.16B. Elongation of 1st internode 0.42 1.0 0.03 -0.84a,*** 0.56* 0.27C. Number of leaves at 2 weeks -0.23 -0.07 1.0 -0.28 -0.05 0.28D. Days to flowering 0.06 -0.52* -0.46* 1.0 -0.43* -0.33E. Specific leaf weight 0.08 0.34 0.11 -0.47* 1.0 -0.08F. Height -0.29 -0.31 0.14 -0.04 0.14 1.0

aSignificant at P <0.01 after sequential Bonferroni correction (k = 15 within each environment). *P < 0.05.

Table 5 Partial regression coefficients(standard errors) for plasticity from multi-ple regression of family mean relative fit-ness in one environment on family meantrait value in that environment and theplasticity of the trait

Trait No neighbors Neighbors

Days to emergence -0.070 (0.26) 0.470 (0.26)Elongation of first internode (cm/g) 0.018a,** (0.006) -0.080* (0.04)Number of leaves at 2 weeks -0.213 (0.22) -0.496a,** (0.16)Days to flowering -0.009 (0.02) 0.036 (0.02)Specific leaf weight (g/cm2) -0.188 (0.14) 0.098 (0.07)Height (cm) 0.013* (0.006) -0.003 (0.004)

*P < 0.05, **P < 0.01aSignificant (P < 0.01) after sequential Bonferroni correction (k = 6 within each

environment).

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© 2002 The Society for the Study of Species Biology Plant Species Biology, 17, 109–118

might be argued that, because the response was so limitedas to keep time of emergence within a range that did notaffect fitness, this response actually reflects adaptivecanalization.

We found that plasticity in traits that reflect plant size,including number of leaves at two weeks and height, waseither nonadaptive or maladaptive. Donohue et al. (2000a)and van Kleunen et al. (2000) also report patterns of selec-tion suggesting that traits that reflect plant size (numberof nodes, length of the first three stolon internodes, andleaf size) tend to be favored with and without neighborspresent. Although empirical demonstrations remain rare,much of the plasticity observed in traits that reflect overallplant size may not be adaptive (cf. Donohue & Schmitt1999; Winn 1999).

Elongation of the first internode was the only trait forwhich our analysis supported selection favoring adaptiveplasticity. Although we found the expected pattern of aplastic increase in stem elongation in response to the pres-ence of neighbors, the pattern of selection we observedfor plasticity in stem elongation was exactly opposite ofwhat others have reported. Both previous studies and thefunctional explanation for adaptive stem elongation sug-gest that selection should favor greater elongation whencompetitors are present and less elongation when they arenot (Schmitt et al. 1995; Dudley & Schmitt 1996; Donohueet al. 2000a; Weinig 2000). Our observation of selectionfavoring elongation in the absence of neighbors could beexplained by strong negative correlations between elon-gation and time to flowering in our experiment, alongwith selection favoring early flowering.

The multivariate analysis of selection and the patternsof correlation between time to flowering and elongationsupport the hypothesis that selection for early floweringoverwhelmed selection on elongation in our no-neighborsenvironment. The direct relationship between elongationand relative fitness in the multivariate analysis in thisenvironment was negative (Table 3), in accordance withexpectations described. The positive net effect in theunivariate analysis (Table 1) appears to result from thenegative covariance between elongation and time to flow-ering (Table 4), coupled with selection favoring earlyflowering in the non-competitive environment (Table 1).In other words, plants that accelerated flowering hadhigher fitness despite elongating inappropriately in theabsence of competitors.

In the competitive environment, we observed negativedirect and indirect effects of internode elongation on rel-ative fitness (Tables 1 and 3). It appears that, in this envi-ronment, plants that elongated did not have greaterfitness despite the added benefit of early flowering. Aplausible explanation is that the level of competition wecreated was too intense to be countered by any achievabledegree of elongation (cf. Weinig 2000). If so, any resources

expended to elongate the stem were effectively wasted,resulting in a fitness decrement. At a lower level of com-petition, elongation might certainly have been advanta-geous. As was the case for flowering time, selection forplasticity in internode elongation seems to vary with envi-ronment (cf. Donohue et al. 2000a; Weinig 2000).


The diagnosis of the adaptive significance of plasticity isimportant to the interpretation of the costs of plasticity orof canalization. When plasticity in a trait is adaptive, acost will limit further evolution toward ideal plasticity.For maladaptive or nonadaptive plasticity, a cost of canal-ization may explain why a plastic response persists evenwhen it has no or even negative effects on fitness.

The significant cost of plasticity in elongation expressedin the no-neighbors environment (Table 5) could contrib-ute to the maintenance of maladaptive plasticity observedfor this trait. The same may be true for nonadaptive plas-ticity in height, although this cost was not significant afterBonferroni correction. The absence of a significant cost ofhomeostasis in either environment for maladaptive plas-ticity in the number of leaves at two weeks could indicatethat forces other than costs explain the maintenance ofplasticity in this trait.

Costs expressed in the presence of neighbors includedonly costs of maladaptive plasticity. These indicate nega-tive fitness consequences of plasticity in the number ofleaves at two weeks and possibly elongation above andbeyond the effects of producing an inappropriate traitvalue within this environment. That maladaptive plastic-ity persists, despite its cost, suggests that plasticity inthese traits reflects unavoidable consequences of growthin a poor environment.

Although we did not find evidence for costs associatedwith all responses to the environment, we did find costsexpressed in at least one environment for all traits show-ing significant genetic variation for plastic response(Table 2). Genetic variation in plasticity is a prerequisitefor a convincing test for evolutionary costs, so it is notsurprising that we did not find costs for some traits.Genetic variation for plasticity is also a prerequisite forevolutionary change in the pattern of plasticity. The lackof genetic variation in some traits could explain why pat-terns of plastic response that are not adaptive persist,although we cannot rule out the possibility that a morepowerful test including a larger number of families mighthave found evidence for both genetic variation and forcosts of plastic responses in these traits.

Results from several studies now support the plausibil-ity that costs contribute to the evolution of patterns of traitresponses to the environment (Dorn et al. 2000; vanKleunen et al. 2000), but the only test so far for a cost of

C O S T S O F P L A S T I C I T Y A N D C A N A L I Z AT I O N 117


adaptive plasticity in a natural environment found noevidence of cost (Donohue et al. 2000a). The degree ofcontrol possible in artificial environments can make theseenvironments a valuable tool for exploring the evolutionof plasticity. Work in greenhouse and laboratory environ-ments is useful for testing the plausibility of hypothesesabout patterns of phenotypic plasticity in nature. Butbecause artificial environments are unlikely to recreateaccurately the patterns of selection in natural environ-ments, the results of such work must be confirmed byfield studies in natural environments.

Our current understanding of variation in patterns ofphenotypic plasticity in plants is limited by the paucity ofempirical data on the adaptive significance of plasticresponses and on the nature of limits to the evolution ofplasticity rather than by lack of sophisticated models,methods, or theory. Data from studies conducted in nat-ural environments that consider both plasticity and canal-ization and demonstrate the selective consequences oftrait responses will be most useful. The results of suchstudies may support an important role of costs, or theymay direct future work toward other forces that couldlimit the evolution of plant responses to the environment.Without more empirical data, the basis for the diversityof patterns of plasticity in plants will remain enigmatic.

Acknowledgements

We thank Tom Miller for providing the Brassica seeds andCecilia Gallup for assistance in the greenhouse. DavidHoule and Carl Schlichting provided helpful commentson an earlier draft of the paper.

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