Towards an ecological understanding of morphological evolution

Towards an EcologicalUnderstanding of MorphologicalEvolutionALEX T. KALINKA*Institut für Populationsgenetik, Vetmeduni Vienna, Vienna, Austria

THE OBLIQUE NATURE OF EVOLUTIONARY TRAJECTORIESComparative studies of embryology have sought to uncover thedevelopmental origins of differences in morphology betweenextant species. Many biological insights have emerged from thisapproach, but arguably the most important contribution was inproviding a firm foundation for Darwin's (1859) principle ofcommon descent (von Baer, 1828). The discovery that embryospass inexorably through “shadows” of their evolutionary pastinspired a generation of embryologists to pursue an evolutionaryresearch agenda (Hall, 2000). Haeckel's (1866) early caricature ofthis process, formulated in his biogenetic law, slowly gave way toan appreciation that ontogeny, in a seeming paradox (West‐Eberhard, 2003), bears the potential for both evolutionary stasisand future change (Garstang, '22).Despite these successes, a research focus that divorces the

organism from its environment is likely to have unintendedconsequences for how we apprehend the evolutionary process. Inparticular, explaining differences in adult morphologies byrecourse to underlying differences in ontogeny tempts us toview morphology as the ultimate unit of selection—being both thegoal of, and the origin for, evolutionary change. Such anunderstanding presupposes a simple directionality to howmorphologies evolve and diverge between species, and to thedegree that it does, we contend that it is equally misleading. When

attempting to unravel the events that led to a morphologicaldifference between closely related species, starting with themorphology and working backwards requires that we impute aselective benefit to each intermediate step all the way back to thecommon ancestor, and, hence, that we view natural selection asproceeding towards the current state with something akin tointentionality. In contrast to this perspective, natural selectionoperates blindly and in response to the immediate demands of theenvironment, and, thus, we can expect evolutionary trajectories tobe largely oblique and goalless.By stripping organisms of their ecological context, we lose sight

of what they are adapting to in the short‐term. This might seemlike a trivial loss, as it may often be clear how particularmorphologies function in a purely organismal context and so

ABSTRACT The roots of modern evo‐devo can be traced back to the comparative anatomy of the 19th century.Inheriting from this tradition, the field has maintained a mechanistic approach to understandingthe origins of distinct animal morphologies. While this focus has produced a valuable body of work,we argue here that a fuller understanding of why species diverge morphologically must be centeredon the selective forces driving divergence, and these forces ultimately reside in the ecologicalcontext in which organisms live and reproduce. We discuss reasons why we expect manymorphological novelties to evolve largely secondarily to, and often as a by‐product of, primaryselection on life‐history traits. By shifting the focus to proximate evolutionary causes, ourperspective necessarily prioritises selection experiments as a means of empirical testing. We outlineexperimental approaches designed to dissect the role of ecological variables in the evolution ofanimal development and morphology, and we show how methods and advances in fields as diverseas population genomics and ecological stoichiometry can contribute to progress in this direction. J.Exp. Zool. (Mol. Dev. Evol.) 9999B: XX–XX, 2014. © 2014 Wiley Periodicals, Inc.

How to cite this article: Kalinka AT. 2014. Towards an ecological understanding of morphologicalevolution. J. Exp. Zool. (Mol. Dev. Evol.) 9999B:1–10.

J. Exp. Zool.(Mol. Dev. Evol.)9999B:1–10, 2014

Conflicts of Interest: None.�Correspondence to: Alex T. Kalinka, Institut für Populationsgenetik,

Vetmeduni Vienna, Veterinärplatz 1, 1210 Vienna, Austria.E‐mail: [email protected]

Received 6 February 2014; Revised 5 May 2014; Accepted 7 May 2014DOI: 10.1002/jez.b.22578Published online XX Month Year in Wiley Online Library

(wileyonlinelibrary.com).

PERSPECTIVE AND HYPOTHESIS

© 2014 WILEY PERIODICALS, INC.

knowing the historical process by which they first emerged addslittle to our understanding of how they have continued to evolve.However, short‐term selection to adapt a life‐history trait to suit aparticular environmental challenge can produce several correlatedchanges at the morphological level, changes that were onlyindirectly driven by selection and whichmay initially have little tono adaptive potential. Nonetheless, these morphological changesmay in turn relax constraints that were previously limiting changein one or more directions. Hence, what we lose is perhaps anessential part of the bigger picture of how evolution proceeds withshort‐term life‐history adaptation enabling species to reachunexplored regions of morphospace in which secondary selectionhones their new morphologies via a gamut of both short‐ andlong‐term processes including species sorting and extinction(Reznick and Ricklefs, 2009; Fig. 1). Primates might provide anexample of this type of two‐step evolution towards novelmorphologies. Early primates possessed the unusual combinationof strongly precocial reproductive strategies and small body sizes,most likely adaptations to their arboreal environment. Thiscombination of traits may have been instrumental in theirsubsequent development of high relative brain size by relaxingconstraints that prevent terrestrial mammals from evolving intothis region of morphospace (Shea, 2007).In what follows, we outline a rationale for why life‐history

evolution is important for understanding the evolution of species,not just in the short‐term, but also in the longer term. We discussspecific cases in which understanding life‐history strategies hasshed light on both the current state and the history of species, and,taking inspiration from nature, we propose research directionsthat can contribute to building an ecological basis for under-standing morphological evolution.

LINKING ECOLOGY, LIFE HISTORY, AND MORPHOLOGYHaeckel (1875) coined the term caenogenesis to refer to exceptionsto the recapitulation of phylogeny through ontogeny (thebiogenetic law) that were caused by relative displacements ofembryonic structures in time (heterochrony) or in space (hetero-topy). While for Haeckel such displacements were wrinkles thatneeded to be ironed out of his theory of recapitulation, it laterbecame clear that these types of alterations to the ontogeny of anorganism could represent important adaptive events producing re‐organized morphologies and even major adaptive radiations.Walter Garstang, together with his daughter Sylvia, proposed theterm paedomorphosis to refer to species that retained juvenilecharacters in their adult forms via precocious sexual maturity(progenesis) and retardation of somatic development (neoteny)—both examples of heterochrony (Garstang and Garstang, '26).Rather than viewing this phenomenon as an exception to a generalrule, they argued that it had the potential to produce radically newmorphologies.Alister Hardy ('54) later recognized that the process of

juvenilization was a general means by which species and lineagescould escape specialization that would otherwise lead to whatHuxley ('47) had termed “blind alleys”—evolutionary pathscharacterized by ever‐increasing specialization leading to inevi-table extinction upon environmental change. de Beer ('54) furtherproposed that as prior adult structures are truncated andeventually lost, the genes responsible for these structures wouldbecome “unemployed,” thus releasing substantial genetic materialfor use in future adaptations. Hence, it had been understood fromvery early on that the importance of paedomorphic heterochronieslay precisely in their ability to contribute to the long‐termadaptive potential of evolutionary lineages.

Figure 1. Depiction of an adaptive radiation following a relaxation of a constraint in a species phylogeny. The yellow triangle indicates thepoint when a life‐history adaptation leads to a correlated change in morphology enabling this lineage to explore a region of morphospacethat was previously not possible. This event triggers an adaptive radiation of species, and competition between the resulting lineages leads toextinction events. Over time, the rate of morphological evolution slows to what it was prior to the original adaptive event.

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At the same time, it had also been appreciated that changes suchas the shifting of sexual maturity to earlier periods of developmentrepresented life‐history strategies that must have been adapta-tions to the immediate environmental circumstances of individualspecies. Nonetheless, it was Gould ('77) who undertook the mostthorough exposition of this idea. He argued that species with highrates of intrinsic population growth and low levels of intra‐speciescompetition, such as colonizers encountering abundant resources,would greatly increase their rate of population growth bybecoming sexually mature earlier (progenesis). In contrast tothis scenario, Gould predicted that neoteny (a retardation ofsomatic development) would be a favoured strategy when there isdensity‐dependent competition for limited resources since longerperiods of somatic growth would lead to enhanced adultcompetitiveness. Although both types of change are examplesof paedomorphosis, they occur in response to very differentecological pressures.Gould grounded his argument in the r‐K‐selection theory that

had grown in popularity in the 1970s following the work ofMacArthur and Wilson ('67) and Pianka ('70). However, at aroundthe same time that Gould published these ideas, it was starting tobecome clear that the neat dichotomy of life‐history traitspredicted by the r‐K model was at best a useful heuristic and atworst a gross and misleading oversimplification (Stearns, '77;Charlesworth, '80; Reznick et al., 2002), and Gould's decision touse the r‐K framework as an explanatory device may havecontributed to these particular ideas being somewhat overlooked.Despite the demise of r‐K theory, Gould's ideas continue to havemerit as they provide a solid developmental basis for the evolutionof important life‐history strategies, thereby connecting ecologyand the evolution of ontogeny. Moreover, by stressing the easewith which juvenile and adult traits could be combined viarelatively minor alterations in the hormonal control of develop-mental transitions, Gould had conceived of plausible micro‐evolutionary routes to macro‐evolutionary change. Recentcomparative studies of vertebrate metamorphosis have largelyconfirmed Gould's intuition, showing that evolutionary tinkeringof the hormonal control of metamorphosis can result in majorchanges in both the life history and development of an organism(Laudet, 2011).While it is conceptually appealing to build connections between

fields that are usually treated as disparate and unrelatedphenomena, it is important to ask how empirical studies canhelp to sharpen our understanding of their underlying relation-ships. We propose that there are, broadly speaking, two types ofapproaches that can be undertaken and which are complementaryto each other. The first is to select closely‐related species that havebeen well‐characterized ecologically, and to study the life‐historyadaptations of each species and how these relate to morphologicaldifferences at the developmental level. The second approach is toperform laboratory evolution experiments in which differentecological pressures are used to select for divergent life‐history

traits in separate populations, and to study the correlated changesin morphology and how these have been driven by allelicsubstitutions. Common to both of these approaches is the need tounderstand how different ecological pressures, both individuallyand in combination, can influence the evolution of morphologyvia immediate life‐history adaptation, and, further, how life‐history and morphological traits are linked in the underlyinggenetics of an organism. We discuss each of these approachesin turn.There are several advantages associated with studying species

known to have experienced differential ecological pressures in the(relatively) recent past and for which clear life‐history signaturesare evident. Most importantly, the experiment has already beenconducted by nature, which circumvents the need for theresearcher to reconstruct appropriate ecological conditions in alab setting with all of the uncertainties and unintended side‐effects associated with that enterprise. Additionally, the period ofevolutionary time separating species will for the most part besubstantial enough to measure reasonably large differences intheir morphologies and how these might scale allometrically.However, the longer the time separating species, the greater theuncertainty in reconstructing the sequence of events leading to thecurrent state, and, therefore, in unraveling the relationshipbetween life‐history traits and morphology.An intriguing example of such a natural experiment is

furnished by the swamp rabbit, Sylvilagus aquaticus. This northAmerican species of cottontail rabbit is semi‐aquatic, inhabitingswamps and wetlands in the Southern United States (Terrel, '72). Itis a long‐lived species, which is characterized by a large body sizeand later age at first reproduction relative to other rabbit species,and typically produces a few, large offspring each breeding season(Swihart, '84). This combination of life‐history strategies isunusual by rabbit standards, and may be indicative of a majorchange in the ecological constraints experienced by this species,such as a significant reduction in extrinsic mortality rates. Swamprabbits share a common ancestor with the marsh rabbit, S.palustris, around 1 million years ago (Ge et al., 2013), yet themarsh rabbit does not share any of the unusual life‐historystrategies of the swamp rabbit (Swihart, '84). In contrast, thisspecies has a small body size and is characterized in particular byunusually small ears and short legs. Their species ranges also donot overlap, with the swamp rabbit living in a region spanningAlabama to Texas and the marsh rabbit living mainly in theFlorida peninsula and along coastal regions to the north‐east(Fig. 2). The combination of divergent life‐history and morpho-logical traits, close but non‐overlapping ranges, and recentevolutionary divergence make this species pair ideal for askingquestions about how ecological variables can drive divergence ofboth life‐history and morphological traits, and, with theavailability of a reference rabbit genome (Lindblad‐Tohet al., 2011), how these differences are determined at the geneticlevel. These two species also provide a good study system for


LINKING ECOLOGY, LIFE‐HISTORY, AND MORPHOLOGY 3

unraveling the relative contributions of random genetic drift andselection to the evolution of morphological divergence since thelife history traits of the swamp rabbit produce smaller populationsizes than that of the marsh rabbit, and population genomicapproaches could be used to this end (Lachance and Tishkoff,2013).Unlike experiments conducted by nature, laboratory experi-

mental evolution studies allow for the precise control of theconditions under which different populations are adapting andthus enable stronger inferences about causative variables. Forthis reason, they offer an attractive means to investigate themorphological consequences of different ecological pressures,especially when combined with the re‐sequencing of evolvedpopulations (Turner et al., 2011; Turner and Miller, 2012).However, artificial selection for particular traits can producedramatically different responses to selection relative to experi-ments that recapitulate specific ecological pressures. A goodexample of this discrepancy can be seen in selection experimentsconducted on Drosophila melanogaster. In two different experi-ments, populations of flies were successfully selected for fasteregg‐to‐adult development times, and this was achieved byevolving the lines under high larval densities (Joshi andMueller, '96) and by directly selecting flies that developed fasterwhile maintaining the larvae at low densities (Prasad et al., 2001).However, quite strikingly, this was the only trait that evolved inthe same direction in the two selection treatments. In the highdensity treatment, larvae evolved a higher feeding rate, whereas inthe low density treatment the larval feeding rate decreased

(Joshi, 2001). Several other indicators of larval competitiveness,such as urea tolerance and foraging path length, increased at highdensities and decreased at low densities suggesting that selectingdirectly for an important life‐history trait, such as developmenttime, is unlikely to produce a set of correlated traits that arerepresentative of evolution in natural populations. Nonetheless, itwas shown that selection for faster development led to a dramaticreduction in body size which was associated with reproductiveisolation from the control populations and, thus, selection purelyon life‐history traits can lead to major consequences for thecontinued evolution of a lineage (Ghosh and Joshi, 2012).By applying high extrinsic mortality rates to the adults of

D. melanogaster populations, the life‐history theory predictionsof evolution of higher intrinsic mortality rates, decreasedlifespans, and earlier age at first reproduction were confirmed(Stearns et al., 2000). In addition, there were correlated changes inbody size, growth rate, and ovariole number (Gasser et al., 2000).Therefore, to best understand how life‐history traits evolve andinfluence morphological traits, it is essential that experimentalevolution studies closely reproduce ecological pressures known tobe experienced in the wild.One way in which this can be readily achieved is by conducting

evolution experiments in the field. Typically, these experimentsare possible only for species that are naturally confined to certainfixed regions, such as freshwater fish, or species which have slowor limited migration and dispersal. By conducting controlledperturbation experiments with Trinidadian guppies, Reznick et al.('90) found, during an 11‐year experiment, that fish transplanted

Figure 2. Species ranges for the swamp rabbit (left) and the marsh rabbit (right) in the southern United States. Note that the ranges areneighboring but non‐overlapping. These two species are relatively recently separated (1 million years ago) and have since evolved a suite ofdistinct life‐history and morphological traits.


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from rivers in which they experienced high adult predation torivers where they instead experienced high juvenile predationevolved delayed sexual maturation combined with the productionof fewer, larger offspring, a result that is in agreement withtheoretical predictions. Subsequent density‐manipulation experi-ments have shown that their life‐history evolution is influencedby population density with higher densities leading to greaterinvestment in juvenile growth and higher reproductive investmentby females (Reznick et al., 2012). The rate of evolution of life‐history traits in the original experiment was found to beunexpectedly high, up to seven times higher than rates ofevolution inferred from palaeontological data (Reznick et al., '97).Similar rapid evolution of life‐history traits was recently found infield experiments involving the common evening primrose plant(Agrawal et al., 2013). In eight replicate populations, it was foundthat shorter lifespan and later flowering times consistentlyevolved over just three generations in response to low competitionand seed predation by the larvae of a single moth species.Controlled experimental evolution studies have demonstrated

that life‐history traits can respond rapidly to selection in newenvironments, and are often associated with correlated morpho-logical changes, particularly in body size. Long‐term studies ofthese populations will provide better opportunities to assess howmorphologies change in response to life‐history adaptation. Inparticular, it would be extremely valuable to determine to whatextent the shifting of sexual maturation to earlier stages might becontributing to juvenilization, or the mixing of adult and juveniletraits, in the evolving populations, andwhether this is enabling theexploration of new regions of morphospace. Amphibian studieshave found strong evidence that paedomorphic strategies areassociated with enhanced fitness in specific ecological settings,such as low juvenile predation and cave environments (Semlitschet al., '90; Bonett et al., 2014; Denoel and Ficetola, 2014), but to beable to study the emergence of paedomorphic characters in realtime in experimental populations would bring the possibility ofdisentangling the primary selected traits from those that evolvesecondarily. In addition, the use of controlled experimentalpopulations will facilitate genomic approaches that can ultimatelyprovide the basis for evidence of adaptation at the genetic level(Barrett and Hoekstra, 2011). Whole‐genome re‐sequencing ofexperimentally evolved lines has helped to shed light on thegenomic basis of life‐history traits such as aging (Remolinaet al., 2012), and studies such as these have the potential to unravelthe relationship between life‐history traits and morphology.Finally, experimental evolution studies can be combined withgenetic manipulations, such as gene replacement, mutagenesis,and transgenic alterations (Kawecki et al., 2012), and candidategenes or pathways can become the focus of individual experi-ments (Bettencourt et al., '99); the increased forethought andcontrol associated with these types of experiments can helpto sharpen the prior hypotheses of otherwise “blind” evolutionexperiments.

In general, the availability of affordable sequencing technologieshas made population genomics a viable approach for understand-ing how species evolve, and in concert with experimental evolutionstudies promises to bridge the genotype‐phenotype gap for life‐history andmorphological traits. A broad range of phenotypes havebeen studied in this setting so far, such as hypoxia‐tolerance (Zhouet al., 2011), resistance of temperature extremes (Orozco‐terWengelet al., 2012), courtship song (Turner and Miller, 2012), andparasitoid resistance (Jalvingh et al., 2014). While these studiesexemplify the use of population‐level sequencing for uncoveringthe genetic basis of complex traits, the technology will ultimatelyenable the simultaneous measurement of genomic and phenotypicchanges during controlled evolution experiments thereby provid-ing a near‐complete picture of how an evolving population changesin specific ecological contexts.When considering which questions are best to study and what

biological systems are most tractable, it is often possible to deriveboth insight and inspiration from examples proffered by natureitself.With this inmind, we nowdiscuss what can be learned aboutshort‐ and long‐term evolutionary dynamics from the oddity thatis island evolution, and we ask to what extent life‐history trade‐offs are being illuminated by our growing knowledge of howorganismal energy budgets are managed across different biologi-cal levels.

Island EvolutionThe unusual and often fascinating evolution of island species hascaptured the imagination of biologists since Darwinfirst discussedthe adaptive radiation of finches on the Galapagos islands. In astudy of 116 island species of mammals, Foster ('64) concludedthat the body size of island species either increased or decreasedrelative to their mainland equivalents, and that the direction ofthis change depended on the difference in ecological conditionsthat were experienced on the island compared to the mainland. Heargued that small‐bodied mainland species tended to increase insize on islands due to an absence of predation, and large‐bodiedspecies tended to decrease in size due to experiencing a reductionin resource availability. It has further been observed that islandbirds tend to have reduced clutch sizes (Cody, '66), and male birdsoften reduce their investment into secondary sexual ornaments(Mayr, '42). In all of these cases, it is the unique ecology of theisland environment that is the driving force behind shifts in life‐history traits relative to the mainland (Case, '78). While thesestudies focused on true islands, Macarthur and Wilson ('67)extended the concept to include any habitable region surroundedby uninhabitable or unsuitable regions, such as grasslands orwater bodies surrounded by desert. What is crucial for producingthe unique patterns of evolution seen on islands is that apopulation is relatively isolated from gene flow coming fromoutside populations.It has long been noted that island populations evolve rapidly

relative to mainland populations. A recent study combining data



from both fossils and extant species, found that island mammalshave tended to evolve rapidly, at the morphological level, oversurprisingly short time‐scales (Millien, 2006). Such rapid evolu-tion has also been documented during a single observationalstudy; it was found that the beak morphology of a species ofDarwin's finches diverged significantly from that of a competitorjust 22 years after the competitor species had arrived on the island(Grant and Grant, 2006). Thus, the constraints generated by inter‐species competition, or predation, can have strong effects on theevolutionary divergence of species whenever these constraints areeither relaxed or newly imposed. Therefore, to observe high ratesof evolution in experimental settings, it may be optimal toassemble a base population from the wild, as opposed to startingwith lab‐adapted populations. In addition, it is important to knowsomething about the ecological constraints experienced undernatural conditions so that these constraints can be relaxed,imposed, or maintained during the experiment depending onwhatoutcome is desired. A clear example of this strategy ofmanipulating ecological constraints is provided by the experimentof Reznick et al. ('90) in which they switched a population ofguppies from a regime of adult predation to one of juvenilepredation. In general, understanding the constraints that limit thenumber of possible evolutionary trajectories available to speciescan help both in interpreting the variation in morphological traitsacross extant species and in uncovering the molecular changesresponsible for this variation (Lewitus and Kalinka, 2013; Lewituset al., 2013).Another feature of island evolution is that it often involves an

initial reduction in population size as islands are usually foundedby a small number of colonizing individuals. Mayr ('54, '63)viewed these founder events as central to the processes drivingdifferentiation and eventually speciation at the periphery ofspecies ranges. He argued that in large, panmictic populationsthere would be substantial genetic variation and, hence, allelesthat reached high frequencies in these populations would need tobe “good mixers,” that is, they would need to have high fitnessacross a broad range of genetic backgrounds; in contrast, after afounder event, and the ensuing reduction in genetic variance,alleles that succeed would instead be those that have high fitnesson a much more homozygous genetic background. This change ingenetic environment, Mayr suggested, would lead to a knock‐oneffect across the genome as a new suite of alleles would sweep tofixation, instigating what he termed a “genetic revolution.” InMayr's view, such founder effects would quickly lead tospeciation.Although the validity and importance of founder‐effect specia-

tion has proved controversial (Barton and Charlesworth, '84;Gavrilets andHastings, '96; Coyne andOrr, 2004; Templeton, 2008),it remains true that island species tend to evolve rapidly andfounder effects may contribute to this process (Provine, 2004). Inaddition, theoretical studies support Mayr's intuition that in large,sexual populations alleles that have high average fitness across a

range of genetic backgrounds will be those that reach higherfrequencies (Livnat et al., 2008), and, hence, we might expect verydifferent sets of alleles to succeed in populations that haveexperienced a reduction in genetic variation. Therefore, it remainsan important and open question as to whether employing restrictedpopulation sizes in evolution experiments, as was practiced byStearns et al. (2000) (using populations of 200 flies), mightcontribute to elevated rates of evolution and in this way greatlyfacilitate our ability to observe both life‐history and morphologicalevolution within reasonable periods of time.

Trade‐Offs in Energy AllocationLife‐history theory has enjoyed a great deal of success inpredicting the shapes of trade‐offs and the optimal strategies thatorganisms are expected to employ (Roff, '92; Stearns, '92; Charnovand Downhower, '95), but there has been comparatively little workdevoted to deciphering the genetic and molecular mechanismsthat mediate these trade‐offs. As trade‐offs are ultimately basedon the acquisition and partitioning offinite energy into competingdemands, such as reproduction and growth, it makes sense tofocus efforts on studying pathways already known to be involvedin shuttling resources into specific physiological processes,especially those that are rate‐limiting in their effects. When anorganism invests resources into growth, this will inevitably detractfrom the resources that can be invested into reproduction, and themolecular determinants of this investment strategy mustultimately be responsible, in some measure, for both the bodysize of the organism and the scheduling of its reproduction.However, the extent to which this is true will depend strongly onwhich nutrients are limiting in particular environments.Organismal growth depends on protein synthesis, and protein

synthesis depends largely on the translational machinery of thecell (rRNA, tRNA, mRNA, and ribosomal protein); these facts formthe basis for the growth‐rate hypothesis (GRH), which posits thatthe growth rate of an organism is an increasing function of itsphosphorous (P) concentration as this is the nutrient limiting RNAcontent (Sterner, '95). It has been found that the GRH holds acrossa wide variety of organisms, from bacteria to D. melanogaster,with a close relationship being uncovered between growth rate,RNA content, and biomass P (Elser et al., 2003). An importanttrade‐off in this system is between the rate and the yield of theprotein synthesis machinery (Dethlefsen and Schmidt, 2007;Matzek and Vitousek, 2009). There is evidence that rRNA genes(rDNA) contribute to faster developmental rates in several species(Cluster et al., '87; Gorokhova et al., 2002) and of particularimportance is the rDNA copy number (White and McLaren, 2000).There is also a strong positive correlation between rDNA copynumber and genome size across a range of eukaryotes(Prokopowich et al., 2003), suggesting a correlated effect on celland, therefore, body size, and evidence that rDNA copy numbervaries according to geographical location, and, hence, mayunderlie local adaptation within species (Long et al., 2013). In an


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artificial selection experiment onDaphnia pulex, it was found thatjust two generations of selection for low weight‐specific fecundityled to increased juvenile growth rates, higher RNA and P content,and an increase in long intergenic spacers associated with rDNAcopies (Gorokhova et al., 2002). These changes acted tocompensate for selection to produce lower fecundity, andhighlights how rapidly life‐history traits can respond to selectionvia alterations to the machinery responsible for organismalgrowth (Hairston et al., 2005; Weider et al., 2005).Overall, the empirical evidence supports the notion that trade‐

offs in life‐history traits associatedwith the allocation of resourcesinto growth are mediated in large part by cellular investment intothe machinery of protein translation, and, thus, this system oughtto be a major focus when investigating the evolution of growth‐related trade‐offs. Furthermore, by understanding what ismediating a trade‐off at the molecular level, it is possible toavoid performing experimental evolution studies with keynutrients being at levels that are not rate‐limiting and therebyabolishing the trade‐offs that are the subject of study. Futureintegration of stoichiometry and evolutionary biology promises tobe rewarding (Kay et al., 2005), although further progress in thisdirection may require the development of a formal metabolictheory (Sousa et al., 2008).The trade‐off between growth and differentiation is one of

fundamental importance to developing embryos, determing boththeir rate of development and their eventual body size (Fig. 3).Such a trade‐off emerges because as cells become terminallydifferentiated they often lose the ability to proliferate, therebylimiting the continued growth of the organism (Loomis, '32). It hasbeen proposed that this trade‐off underlies the difference between

precocial and altricial reproductive strategies in animals (RicklefsandWeremiuk, '77); precocial offspring aremore highly developedthan altricial offspring, and, unlike the latter, can open their eyesand move unaided shortly after birth. However, altricial speciestypically have much higher post‐embryonic growth rates relativeto precocial species, in agreement with the existence of a trade‐off.Tissue types that are expected to contribute to this trade‐off arethose in which terminally differentiated cells no longer divide, andin animals, tissues with this characteristic are limited to skeletalmuscle, nerves, and bone (Ricklefs et al., '94), althoughmany othertissue types will also exhibit reduced proliferative capacities asthey differentiate (Ricklefs et al., '98; Arendt, 2000).While increased growth and body size tend to correlate with

higher fecundity and competitiveness across species (Kingsolverand Huey, 2008), there may be ecological constraints that preventa large body size from being attained, such as a need to developrapidly. Under these circumstances, maternal investments mayplay a major role in off‐setting the costs of rapid differentiation,thereby enabling higher levels of growth than would be otherwisepossible. This can be achieved through the deposition of energy‐rich yolk in the egg (Storm and Angilletta, 2007), behavioraleffects such as the incubation of eggs (Martin and Schwabl, 2008),andmaternal contributions to the ribosome pool of early embryos.Hence, maternal effects are crucial for understanding theadaptation of early embryos to their ecological circumstances(Kalinka and Tomancak, 2012), and comparativemolecular studiesare helping to shed light on the essential transition from maternalcontrol to zygotic genome activation (Heyn et al., 2014). Progressin understanding the importance of maternal investment can bemade at first using relatively simple experimental approaches. In a

Figure 3. Schematic illustrating the growth‐differentiation trade‐off. Open circles represent dividing, totipotent cells, and circles withpatterns represent terminally differentiated cells belonging to specific tissue types. On the left, rapid development, illustrated by a shorterperiod of time until all tissue types have been produced, results in fewer cells than on the right where development is slower. The arrowheadon the right indicates that one potential consequence of slower development and the production of more cells is that there is a greater chancethat new cell types emerge during the ontogenetic period.



recent study, Landberg (2014) surgically removed yolk from theeggs of a salamander species, which resulted in early hatching,and reduced development and body size indicating that maternalinvestments impact both differentiation and growth.Rates of differentiation of somatic and germline cells can also

impact animal morphology both in the short‐ and long‐term. Inparticular, the timing of germline determination can havesignificant evolutionary consequences for individual species. Ifthe germline is determined relatively late during embryogenesisthen there will be several mitotic divisons preceding theirspecification during which time there is scope for competitionbetween soma cells for access to the future germline, and, thus,species with late germline specification are “open” to bothdeleterious and rare beneficial modifications of their ontogeny(Buss, '87). Furthermore, Buss argues that early and late germlinespecification correspond to progenesis and hypermorphosis(exaggerated somatic growth) in Gould's ('77) terminology.However, it is unclear to what extent the timing of germlinedetermination correlates with age at sexualmaturity, as is requiredby the scheme proposed by Buss, and it would be valuable to knowwhether such a relationship holds, and, if so, why. In general,discovering the major molecular processes and pathways that areinvolved in modulating the rate of development will enhance ourunderstanding of how growth and differentiation trade‐offsevolve and how these might be related to heterochronicmorphological evolution.

SUMMARYAttempts to understand the evolution of morphological differ-ences between species by starting with the morphology andworking backwards will ultimately be misleading. Evolutionarytrajectories are oblique and goalless, and species adapt to theimmediate demands of their environment via life‐history traitsthat are closely related to fitness. Changes in traits such as the ageat sexual maturity and the rate of development can produceseveral correlated changes in the morphology of an organism,especially as body size usually scales allometrically with shape.For these reasons, we advocate an approach to understandingmorphological evolution that places emphasis on deciphering therole of ecological variables in driving changes in life‐history traits,and on uncovering the relationship between these changes andlonger‐term heterochronic alterations in organismal ontogenies.We describe different experimental strategies for making

progress in this direction. First, pairs of species known to havediverged recently, and that have distinct life‐history strategies andmorphologies, represent ideal systems for comparative studies atboth the ecological and molecular levels. As an example of such asystem, we discussed the swamp and marsh rabbits inhabitingneighboring regions of the southern United States. Second,experimental evolution studies can be employed to selectpopulations under different ecological constraints followed byanalysis of the resulting changes in both life‐history and

morphology. Long‐term field experiments may be the best wayto reproduce realistic ecological pressures, and the peculiarities ofisland evolution can illuminate the best approaches to producerapid evolutionary changes over short periods of time.Finally, we discuss the importance of understanding how trade‐

offs in energy allocation are mediated at the molecular level. Thetrade‐off between growth and differentiation is of particularinterest as it strongly influences the rate of development, bodysize, and the use of alternative reproductive strategies, and itinvolves the fundamental transition from the mother to thezygote. By marshaling tools from diverse disciplines, such aspopulation genomics, ecological stoichiometry, and life‐historytheory, it will be possible to build a solid foundation forunderstanding how animal morphology is moulded within anecological context.

ACKNOWLEDGMENTThe author gratefully acknowledge Iva Kelava for making all threefigures, and providing helpful comments on the manuscript.

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Towards an ecological understanding of morphological evolution