M ORE IS BETTER: THE USES OF DEVELOPMENTAL GENETIC DATA

American Journal of Botany 96(1): 83–95. 2009. MORE IS BETTER: THE USES OF DEVELOPMENTAL GENETIC DATA TO RECONSTRUCT PERIANTH EVOLUTION1 Lena C. Hil...
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American Journal of Botany 96(1): 83–95. 2009.

MORE IS BETTER: THE USES OF DEVELOPMENTAL GENETIC DATA TO RECONSTRUCT PERIANTH EVOLUTION1

Lena C. Hileman2,5 and Vivian F. Irish3,4 2Department of Ecology and Evolutionary Biology, University of Kansas, Lawrence Kansas 66045 USA; 3Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven Connecticut 06520 USA; and 4Department of Ecology and Evolutionary Biology, Yale University, New Haven Connecticut 06520 USA

The origin and evolution of the perianth remains enigmatic. While it seems likely that an undifferentiated perianth consisting of tepals arose early in angiosperm evolution, it is unclear when and how differentiated perianths consisting of distinct organs, such as petals and sepals, arose. Phylogenetic reconstructions of ancestral perianth states across angiosperms have traditionally relied on morphological data from extant species, but these analyses often produce equivocal results. Here we describe the use of developmental genetic data as an additional strategy to infer the ancestral perianth character state for different angiosperm clades. By assessing functional data in combination with expression data in a maximum likelihood framework, we provide a novel approach for investigating the evolutionary history of the perianth. Results of this analysis provide new insights into perianth evolution and provide a proof of concept for using this strategy to explore the incorporation of developmental genetic data in character state reconstructions. As the assumptions outlined here are tested and more genetic data are generated, we hope that ancestral state reconstructions based on multiple lines of evidence will converge. Key words: ancestral state reconstruction; angiosperm; APETALA3; AP3; development; evolution; maximum likelihood; perianth.

The evolution of flowers is thought to underlie the extensive radiation of angiosperms through enhancement of efficient interactions with animal pollinators to facilitate reproductive success (Regal, 1977). Flowers are generally composed of a perianth of sterile organs surrounding the reproductive organs— the pollen-bearing stamens and ovule-bearing carpels, with the perianth often differentiated into protective outer organs (sepals) and showy inner organs (petals). Flower morphology has diversified through evolutionary modifications of these floral organs, with some of the most striking changes in form due to modifications in perianth morphology. For example, perianth organization has transitioned from spiral to whorled phyllotaxy, distinct perianth organ types have evolved multiple times, and there have been a number of transitions from radial to bilateral perianth symmetry. Across flowering plants, the perianth may remain undifferentiated as tepals, or may be highly differentiated into outer sepals and inner petals, or may have evolved derived morphologies such as the palea, lemma, and lodicules of grass flowers (Endress, 2006; Ronse De Craene, 2007). Transitions between an undifferentiated (unipartite) and a differentiated (bipartite) perianth appear to have occurred multiple times during angiosperm evolution (Fig. 1) (Zanis et al., 2003; Soltis et al., 2005; Endress, 2006; Ronse De Craene, 2007, 2008). These morphological data have been used to reconstruct ancestral character states for key nodes in the angiosperm phylogeny (e.g., the ancestor of core eudicots, all eudicots, or of all angiosperms). However, these analyses have not led to a fully satisfying answer as to the likely course of perianth evolution in specific cases; for example, did the ances1

tors of eudicots and core eudicots develop a differentiated or undifferentiated perianth (Fig. 1) (Zanis et al., 2003; Soltis et al., 2005; Endress, 2006; Drea et al., 2007; Ronse De Craene, 2007, 2008)? Furthermore, it has been hypothesized that the diversity in differentiated perianths may reflect unique underlying developmental programs, with distinct petals being derived in some cases from bract-like organs (bracteopetals) or in other cases evolving as modified sterile stamens (andropetals, Hiepko, 1965; Takhtajan, 1991; Endress, 1994). Because diverse perianth architectures likely reflect differences in developmental programs, we suggest that the incorporation of developmental genetic data into such analyses is a valuable and independent means of refining such reconstructions of ancestral character states. Over the past few years, we have seen an unprecedented number of studies that have examined the expression and/or function of APETALA3/DEFICIENS (AP3/DEF, hereafter referred to as AP3) homologs in diverse angiosperm taxa. In Arabidopsis thaliana, AP3 is required to specify petal and stamen identity, and loss of AP3 activity results in homeotic transformations of stamens to carpels and petals to sepals, resulting in an undifferentiated perianth (Jack et al., 1992). Functional analyses of AP3 homologs have now been carried out in diverse taxa including snapdragon (Antirrhinum majus, Sommer et al., 1990; Schwarz-Sommer et al., 1992), flowering tobacco (Nicotiana benthamiana, Liu et al., 2004), tomato (Solanum lycopersicum, de Martino et al., 2006), petunia (Petunia hybrida, Rijpkema et al., 2006), poppy (Papaver somniferum, Drea et al., 2007), rice (Oryza sativa, Prasad and Vijayraghavan, 2003; Xiao et al., 2003), and maize (Zea mays, Ambrose et al., 2000; Whipple et al., 2004). Given this boom in availability of functional data, as well as detailed gene expression data, we are now in a position to begin to reconstruct ancestral AP3 protein function (and patterns of gene expression) across much of angiosperm diversity. Determining ancestral function and expression of developmental regulators will allow for the extrapolation of ancestral morphology at key nodes in angiosperm phylogeny.

Manuscript received 19 February 2008; revision accepted 8 July 2008.

The authors thank P. Cartwright and J. Preston for helpful discussions and two anonymous reviewers for their comments. This work was supported by NSF (IOB-0616025) to L.C.H. and NSF (IOB-0516789) to V.F.I. 5 Author for correspondence (e-mail: [email protected]) doi:10.3732/ajb.0800066

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how the petal developmental module characterized in Arabidopsis has diversified across flowering plants to specify unique perianth organ identities. In Arabidopsis, AP3 expression commences at petal primordium initiation and continues to late stages of petal development (Jack et al., 1992). AP3 expression is necessary throughout petal development because loss of AP3 function at various times during petal differentiation results in loss of petal cell identity (Bowman et al., 1989; Carpenter and Coen, 1990; Zachgo et al., 1995). AP3 expression is initiated by the combined activities of LEAFY (LFY), a plant-specific transcription factor, in conjunction with UNUSUAL FLORAL ORGANS (UFO), a component of an SCF E3 ubiquitin ligase complex, which acts as a transcriptional cofactor of LFY to activate AP3 expression (Fig. 2) (Lee et al., 1997; Chae et al., 2008). AP3 is required to coordinate intercellular signaling necessary for normal petal morphogenesis and presumably carries this out by regulating the expression of downstream genes required for cell–cell signaling (Jenik and Irish, 2001). A number of genes have been identified as transcriptional targets of AP3 action in Arabidopsis; these include AP3 itself, PI, NAC-LIKE ACTIVATED BY AP3/PI (NAP), APETALA1 (AP1), GATA, NITRATE-INDUCIBLE, CARBON METABOLISM-INVOLVED (GNC) and GNC-LIKE (GNL; Fig. 2). AP3 has been shown to positively regulate its own expression, by binding to consensus CArG box sequences in its promoter (Hill

Fig. 1. Ancestral character state reconstruction for evolutionary transitions between a unipartite and bipartite perianth based on morphology, generated under the parsimony criterion as implemented in Mesquite version 2.01 (Maddison and Maddison 2007). Ancestral reconstructions only at major nodes are shown (base of angiosperms, magnoliid dicots, monocots + eudicots, eudicots, and core eudicots). Phylogeny and character states taken largely from Soltis et al. (2005), with the positions of Caryophyllids, Ceratophyllum, Chloranthus, and monocots according to Moore et al. (2007). The ancestral condition for angiosperms was likely an undifferentiated perianth, but the number and direction of change in perianth morphology is equivocal, with uncertainties in ancestral state reconstruction at the base of the eudicots, and eudicots + monocots.

In this paper, we provide an explicit phylogenetic framework for integrating such developmental genetic data into our understanding of perianth evolution. This analysis should serve as a model for the incorporation of developmental genetic data, together with morphological data, in inferring ancestral states for a wide range of characters. AP3 specifies identity of second whorl perianth organs in Arabidopsis— The model plant species Arabidopsis thaliana develops flowers with a bipartite perianth consisting of outer whorl sepals and inner whorl petals. Mutations in the MADSbox genes AP3 and PISTILLATA (PI) result in the homeotic transformation of petals into sepals; therefore, the products of these genes are necessary for the specification of a bipartite perianth (Bowman et al., 1989; Jack et al., 1992; Goto and Meyerowitz, 1994). Recent Arabidopsis developmental genetic studies have led to a synthetic understanding of the genetic module necessary for proper petal development. In addition, comparative AP3 functional studies are beginning to shed light on the extent to which homologs of AP3 are necessary for the specification of distinct perianth organ identities in diverse angiosperms. Ultimately, the aim of such studies is to determine

Fig. 2. Known genetic interactions in the Arabidopsis petal developmental module. Lines ending with arrows indicate positive regulation of target genes; lines ending bluntly indicate negative regulation of target genes. See text for descriptions of genes.

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et al., 1998; Tilly et al., 1998; Honma and Goto, 2000; Lamb et al., 2002). AP3 negatively regulates the expression of the MADS box transcription factor AP1, which is also required for petal development (Irish and Sussex, 1990; Mandel et al., 1992; Sundstrom et al., 2006). AP1, in turn, positively regulates the expression of UFO, resulting in a feedback control loop that presumably acts to maintain appropriate levels of AP3 expression during petal development (Ng and Yanofsky, 2001). In addition, AP3 promotes the expression of NAP, a putative NAC family transcription factor that has been implicated in regulating the transition from cell division to cell expansion during later phases of petal growth (Sablowski and Meyerowitz, 1998; Guo and Gan, 2006). In contrast, AP3 represses the expression of GNC and GNL, two GATA transcription factors required to promote chlorophyll biosynthesis, providing one explanation as to how chlorophyll production is downregulated during the development of Arabidopsis petals (Mara and Irish, 2008). Comparative analyses of AP3 function during perianth development— The identification of downstream targets of AP3 action in Arabidopsis is beginning to illuminate the hierarchy of events governing the development of petals in this species. To what extent are such processes conserved in specifying petal or perianth organ development in other angiosperms whose perianth structures may have evolved independently of Arabidopsis? To start to address this issue, a number of researchers have focused on characterizing the roles of AP3 homologs in diverse angiosperm taxa. In addition to Arabidopsis, the function of AP3 lineage genes has been examined in the core eudicot species snapdragon (Sommer et al., 1990; Schwarz-Sommer et al., 1992), flowering tobacco (Liu et al., 2004), tomato (de Martino et al., 2006), and petunia (Rijpkema et al., 2006), the noncore eudicot species opium poppy (Drea et al., 2007), and the monocot species rice (Prasad and Vijayraghavan, 2003; Xiao et al., 2003) and maize (Ambrose et al., 2000; Whipple et al., 2004). Near the base of the core eudicots, a gene duplication event in the AP3 lineage gave rise to the paralogous gene lineages AP3 and TM6 (Fig. 3) (Kramer et al., 1998). Therefore, many core eudicot species possess paralogous AP3 lineage and TM6 lineage genes in their genome. For example, AP3 lineage orthologs are AP3, DEF, TAP3, PhDEF (also known as GREEN PETAL), and NbDEF in Arabidopsis, snapdragon, tomato, petunia and flowering tobacco, respectively (Fig. 3), whereas TM6 lineage orthologs are TM6, PhTM6, and NbTM6 in tomato, petunia and flowering tobacco, respectively (Fig. 3). Interestingly, Arabidopsis and snapdragon, our two most well-developed models for studying flower development, have independently lost the TM6 paralog (Lamb and Irish, 2003; Vandenbussche et al., 2004) (Fig. 3). The retention of both AP3 lineage and TM6 lineage genes in many core eudicots may be explained by either the subfunctionalization or neofunctionalization of these genes following their origin via gene duplication (Ohno, 1970; Force et al., 1999, 2005). Under a neofunctionalization model, both AP3 and TM6 paralogs are retained because one paralog has evolved a novel function, for example, a novel role in specifying perianth development. Under a subfunctionalization model, both AP3 and TM6 paralogs are retained because some aspect(s) of ancestral function is retained by one gene copy, for example, a role in specifying perianth development, while another aspect(s) of ancestral function is retained by the other paralog, for example, a role in specifying stamen identity during flower development. Reconstructing ancestral function is critical to test these alternative hypotheses, as is a critical assessment of

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the function of AP3 and TM6 lineage gene products in core eudicots. In snapdragon, loss of DEF function results in similar homeotic transformations to those seen in Arabidopsis; petals are transformed into sepals and stamens into carpels (Table 1) (Sommer et al., 1990; Schwarz-Sommer et al., 1992). Therefore, in both snapdragon and Arabidopsis, the AP3 homolog is necessary for establishing a bipartite perianth. The solanaceous species, tomato, petunia, and flowering tobacco retain both AP3 lineage and TM6 lineage genes in their genomes (Liu et al., 2004; Vandenbussche et al., 2004; de Martino et al., 2006). In tomato, loss of TAP3 function also results in the homeotic transformation of petals into sepals and stamens into carpels. On the other hand, loss of tomato TM6 function only results in homeotic transformation of stamens into carpels and has little effect on perianth development (Table 1) (de Martino et al., 2006). The phenotypes produced by loss of PhDEF and PhTM6 function in petunia are distinct from those of tomato, but also support the idea that the AP3 and TM6 paralogs have diverged in terms of their function. In contrast to the situation in tomato (TAP3), mutations in petunia PhDEF result in homeotic transformations of petals to sepals, but have little effect on stamen development. Loss of petunia PhTM6 function alone does not result in any obvious phenotypic abnormalities, but coordinated downregulation of both petunia PhDEF and PhTM6 results in the transformation of stamens to carpels and petals to sepals. These data show that both PhDEF and PhTM6 are required for specification of stamen identity, but PhDEF alone is required for proper petal specification (Table 1) (Rijpkema et al., 2006). Therefore, although their functions are distinct, in both tomato and petunia the AP3 paralog is necessary for the development of a bipartite perianth, while the TM6 paralog has no obvious role in specifying perianth identity. The poppy genome also contains two AP3 paralogs, PapsAP3-1, and PapsAP3-2 (Fig. 3). However, the duplication that gave rise to these paralogous gene lineages is independent of the AP3/TM6 duplication, having occurred early in the radiation of the noncore eudicot lineage Papaveraceae (Drea et al., 2007). In poppy, PapsAP3-1, but not PapsAP3-2, is necessary for the specification of a bipartite perianth. Loss of PapsAP3-1 function leads to the homeotic transformation of petals into sepals, with stamens being largely unaffected. In contrast, loss of PapsAP3-2 function results in the homeotic transformation of stamens into carpels with no effect on poppy perianth development (Table 1) (Drea et al., 2007). Maize appears to possess a single AP3 homolog, Silky1, which is necessary for the specification of inner whorl perianth organs (i.e., a bipartite perianth, Ambrose et al., 2000). Loss of Silky1 function results in the homeotic transformation of lodicules into palea/lemma-like organs, as well as the transformation of stamens to carpels, resulting in an abundance of silks (Ambrose et al., 2000; Whipple et al., 2004) (Table 1). Very similar phenotypes are found in rice plants lacking AP3 homolog function (Nagasawa et al., 2003; Xiao et al., 2003). These functional analyses of AP3 homologs in widely divergent angiosperm species serve to highlight several important conclusions about the roles of these genes in bipartite perianth specification. Notably, these studies make clear that AP3 homologs do not always have a role in the specification of perianth identity. Interestingly, the AP3 homologs in poppy, petunia, and tomato, which do not have demonstrable functions in petal specification, are nonetheless expressed in developing petal primordia (Fig. 5 and Table 1) (de Martino et al., 2006; Rijpkema

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Fig. 3. Phylogeny of AP3 lineage genes used in this study for ancestral character state reconstruction of AP3 homolog function. Locations of gene duplication events indicated by black circles. Gene relationships and branch lengths optimized under maximum likelihood using the GTR+Γ model of molecular evolution as implemented in GARLI version 0.951 (Zwickl, 2006). The inferred losses of TM6 lineage genes in Arabidopsis and Antirrhinum majus are indicated with dashed lines.

et al., 2006; Drea et al., 2007). Furthermore, the observations that a number of AP3 homologs in a variety of distantly related angiosperms do in fact condition bipartite perianth identity, suggests that the expression of these genes in the perianth was a necessary prerequisite to the evolution of distinct perianth organ identities (Whipple et al., 2007). Presumably the evolution of distinct petal types relies not only on AP3, but the recruitment of an AP3-dependent module of genetic interactions, to specify perianth organ identity. Reconstructing ancestral states from developmental genetic data—Developmental genetic data can be viewed as additional information that may help untangle homologies between morphological traits (Abouheif, 1997; Baum and Whitlock, 1999; Brigandt, 2003; Jaramillo and Kramer, 2007). For example, in the case of the perianth, it remains unclear how many times, and in which flowering plant lineages, evolutionary transitions between a unipartite and bipartite perianth have occurred (Fig. 1 and see Zanis et al., 2003; Soltis et al., 2005; Endress, 2006; Drea et al., 2007; Ronse De Craene, 2007, 2008). If we reconstruct ancestral character states for AP3 function, we might find, for instance, that the AP3 lineage gene in the ancestor of eudicots is inferred to have functioned in specifying distinct perianth organ

identities. If this ancestral function were inferred, we would extrapolate that the eudicot ancestor developed flowers with a bipartite perianth and that unipartite perianths, and perhaps bipartite perianths, have been independently derived within the eudicots. It may seem that comparative AP3 functional data will not provide additional insight into our understanding of perianth evolution beyond what we can learn from morphology. One might assume that AP3 function will, in all cases, be tightly associated with the development of a bipartite perianth. However, as discussed, a number of AP3 homologs do not function in specifying perianth identity. However, these results are complicated by the fact that these AP3 lineage genes derive from duplication events (Fig. 3), and in all cases, a paralogous AP3 gene is necessary for the specification of a bipartite perianth. Given this history of gene duplications in the AP3 lineage (Fig. 3) (Kramer et al., 1998; Kramer etal., 2003; Stellari et al., 2004; Aagaard et al., 2005; Drea et al., 2007), there are additional reasons to assess the functional fate of AP3 paralogs in a strictly phylogenetic context. After AP3 duplication events, resulting paralogs may often be immediately lost through the accumulation of deleterious mutations. On the other hand, both paralogs may be maintained through subfunctionalization or the evolution of novel function (neofunctionalization) of at

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AP3 lineage genes that have been characterized using functional and/or in situ mRNA hybridization expression analyses.

Taxonomic lineage

AP3-lineage gene

Loss of function homeotic phenotype

Core eudicots

Monocots

Arabidopsis thaliana AP3 Antirrhinum majus DEF Solanum lycopersicum TAP3 Solanum lycopersicum TM6 Petunia hybrida PhDEF Petunia hybrida PhTM6 Rumex acetosa RaD1 Papaver somniferum PapsAP3–1 Papaver somniferum PapsAP3–2 Aquilegia vulgaris AqvAP3–1 Aquilegia vulgaris AqvAP3–2 Aquilegia vulgaris AqvAP3–3 Zea mays Silky1

Magnoliid dicots

Streptochaeta angustifolia SaAP3 Joinvillea ascendens JaAP3 Elegia elephas EeAP3a Elegia elephas EeAP3b Elaeis guineensis EgDEF1 Asparagus officinalis AoDEF Aristolochia manshuriensis ArmAP3

Petals to sepals, stamens to carpels Petals to sepals, stamens to carpels Petals to sepals, stamens to carpels Stamens to carpels Petals to sepals Stamens to carpels (in def background) Not available Petals to sepals Stamens to carpels Not available Not available Not available Lodicules to palea/lemma-like organs, stamens to carpels Not available Not available Not available Not available Not available Not available Not available

Noncore eudicots

Loss of function character state

Expression character state

1 1 1 0 1 0 n/a 1 0 n/a n/a n/a 1

2 2 2 1 2 1 0 2 1 1 1 2 2

n/a n/a n/a n/a n/a n/a n/a

2 1 2 2 1 2 1

Notes: Loss of function character state assignments: 0, no homeotic transformation of perianth organs; 1, homeotic transformation of perianth organs (e.g., petals to sepals, lodicules to palea/lemma-like organs). Expression data character state assignments: 0, no expression of AP3-lineage gene in perianth organs; 1, AP3-lineage gene expression present in perianth organs, but weak, patchy, or transient; 2, AP3-lineage gene expression ubiquitous in perianth organs from early through later stages of development. n/a, not applicable.

least one paralog (Ohno, 1970; Force et al., 1999, 2005). For distinguishing between subfunctionalization and neofunctionalization hypotheses, it is critical to confidently reconstruct ancestral AP3 functional states predating such duplication events. For example, if AP3 in the ancestor of eudicots is inferred to have functioned only in specifying stamen development, then derived AP3 paralogs that specify petal development would have been independently recruited into that role. On the other hand, if AP3 in the ancestor of eudicots in inferred to have functioned in specifying both stamen and petal identity, then derived AP3 paralogs that specify petal development would simply have retained that ancestral subfunction. To place developmental genetic data into a comparative phylogenetic framework, we reconstructed the ancestral states for protein function or gene expression on our best estimate of the AP3 gene phylogeny using standard likelihood methods (Schluter et al., 1997; Pagel, 1999). Given a tree and defined character distribution for terminals on that tree, likelihood methods will reconstruct the history of ancestral states for that character so as to maximize the likelihood of evolving the observed terminal character states (Maddison and Maddison, 2006). When looking at character evolution of protein function or gene expression, a likelihood approach is preferable to a parsimony approach because it enables branch lengths in the gene tree to be taken into account (i.e., there is more opportunity for character state changes along longer branches than shorter branches). In addition, a likelihood framework provides an estimate of the degree of uncertainty in the ancestral state reconstructions (e.g., Schluter et al., 1997). Although floral developmental genetic data have previously been placed in a phylogenetic framework (for example, Kim et al., 2005; Zahn et al., 2005; Rasmussen et al., 2009, pp. 96–109 in this issue), the analyses presented here are the first that take a maximum likelihood approach and are the first to explicitly focus on the growing amounts of functional data available for AP3 lineage genes across a diversity of angiosperms.

Reconstructing ancestral AP3 function— We coded functional character states for AP3 lineage genes in Arabidopsis, snapdragon, tomato, petunia, poppy, and maize as 0 or 1 (Table 1), with 1 being gene products that, when silenced, result in homeotic transformation of perianth organs, and 0 being gene products that, when silenced, do not result in homeotic transformation of perianth organs. For the data in this analysis, we define homeotic transformation of the perianth as either the transformation of petals to sepal-like organs (eudicots) or the transformation of lodicules to palea/lemma-like organs (maize). Therefore, for the purposes of this analysis, we are equating homeotic transformation of perianth organs resulting from AP3 loss of function, with the development of an undifferentiated perianth in a species that otherwise would develop a bipartite perianth. Ancestral character states for AP3 function were optimized using maximum likelihood (Schluter et al., 1997) as implemented in the program Mesquite version 2.01 (Maddison and Maddison, 2006, 2007) allowing for a single rate of change between character states (model Mk1). A likelihood ratio test rejected a more complex likelihood model of evolution that allows for different forward and backward rates of change between states (model AssymMK) in favor of the Mk1 model. Ancestral character state assignment at a given node in the phylogeny was considered to be significant over alternative states at that node if the difference in –log likelihood was equal to or greater than 2 log units (Edwards, 1972). The AP3 gene phylogeny used for ancestral state reconstruction was generated, and branch lengths optimized, using the maximum likelihood criterion under the GTR+Γ model of molecular evolution (Yang, 1994a, b) as implemented in the program GARLI version 0.951 (Zwickl, 2006). The maximum likelihood ancestral character state reconstruction for AP3 functional data are equivocal as to whether AP3 in the ancestor of core eudicots, eudicots, or eudicots + monocots functioned in specifying a bipartite perianth (nodes C, B, and A, respectively, Fig. 4 and Table 2). Although the most likely re-

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construction suggests that AP3 in the ancestor of monocots + eudicots functioned in specifying perianth organ identity (proportional likelihood of 0.662, node A, Fig. 4; −log likelihood of 5.54, Table 2), this bias toward a role in perianth development is not significant. If this trend toward AP3 functioning in perianth organ identity specification in the ancestor or monocots + eu-

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dicots holds up as more data are amassed, it would suggest that a bipartite perianth was present before the diversification of monocots + eudicots, with multiple subsequent evolutionary transitions toward a unipartite perianth. Although we are now seeing an emergence of functional data for AP3 lineage genes and the accumulation of such data allows us to test hypotheses

Fig. 4. Phylogeny of AP3 lineage genes that have been functionally characterized. At each node, a pie chart shows the proportional likelihood that the ancestor had a particular character state. Black represents the proportional likelihood that the AP3 homolog in the ancestor functioned in the specification of distinct perianth organ identities (bipartite perianth); white represents the proportional likelihood that the AP3 homolog in the ancestor did not function in specifying distinct perianth organ identities (unipartite perianth). Pie charts labeled A–C indicate the ancestor of eudicots + monocots, eudicots and core eudicots, respectively. Gene relationships and branch lengths optimized under maximum likelihood using the GTR+Γ model of molecular evolution as implemented in GARLI version 0.951 (Zwickl, 2006). Numbers above branches indicate node support based on 500 maximum likelihood bootstrap replicates. Asterisks indicate nodes at which the character state with the largest proportional likelihood is significantly preferred over the alternative character state.

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Table 2.

Ancestral state (0–2) reconstruction for AP3 homolog function and expression at key nodes in the angiosperm phylogeny. −Log likelihood of ancestral states

Node

Functional data (Fig. 4)

Core eudicots (node C)

state 0: 5.85 state 1: 5.79

Eudicots (node B)

state 0: 5.82 state 1: 5.82

Monocots (node D)

n/a

Monocots + eudicots (node A)

state 0: 6.21 state 1: 5.54

Expression data (Fig. 6)

state 0: 22.37 state 1: 19.71* state 2: 20.24* state 0: 23.18 state 1: 19.58* state 2: 20.43* state 0: 23.10 state 1: 19.94* state 2: 19.90* state 0: 22.74 state 1: 19.60* state 2: 20.42*

Notes: Nodes A–D as labeled in Figs. 4 and 6. A particular ancestral state reconstruction is considered significantly preferred over an alternative state at a node when the difference in –log likelihood values is equal to or greater than 2 log units. Significance under this criterion is indicated with an asterisk. Functional data character state assignments: 0, no homeotic transformation of perianth organs; 1, homeotic transformation of perianth organs (e.g., petals to sepals, lodicules to palea/lemma-like organs). Expression data character state assignments: 0, no expression of AP3-lineage gene in perianth organs; 1, AP3-lineage gene expression present in perianth organs, but weak, patchy, or transient; 2, AP3-lineage gene expression ubiquitous in perianth organs from early through later stages of development.

of ancestral function in a comparative phylogenetic framework, it is clear that missing data are likely limiting our ability to confidently reconstruct ancestral AP3 functional states (Cunningham et al., 1998; Losos, 1999; Case et al., 2008). Reconstructing ancestral AP3 gene expression— The role of a gene in developmental patterning is most clearly revealed by functional genetic analyses; however, such analyses can be difficult or time consuming. As a proxy, expression patterns can be used to infer the function of a gene, although clearly expression does not always reflect gene function. For instance, there are multiple examples illustrating that even though AP3 homologs may be expressed in the perianth, they do not necessarily function in the specification of perianth organ identity (de Martino et al., 2006; Rijpkema et al., 2006; Drea et al., 2007). Nonetheless, there are considerably more expression data than functional analyses available for AP3 homologs, prompting us to explore the utility of these data in assessing hypotheses of ancestral gene function. To investigate the relationship between gene function and gene expression, we first looked at examples where both expression and functional data are available for AP3 lineage genes (Arabidopsis, snapdragon, tomato, petunia, poppy, maize). Based on these analyses, we conclude that there is strong evidence that patterns of expression are good predictors of function. Ubiquitous expression of AP3 homologs from relatively early through relatively late stages of perianth development cooccurs with a functional role in the specification of perianth organ identity (Fig. 5 and Table 1). On the other hand, patchy, weak, or transient expression of AP3 homologs during perianth development co-occurs with a lack of homeotic function during perianth development. We examined in situ patterns of RNA hybridization for AP3 homologs across a diversity of monocots and eudicots includ-

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ing Arabidopsis (Jack et al., 1992, 1994), snapdragon (SchwarzSommer et al., 1992), tomato (de Martino et al., 2006), petunia (Halfter et al., 1994; Vandenbussche et al., 2004), sorrel (Rumex acetosa, Ainsworth et al., 1995), opium poppy (Drea et al., 2007), columbine (Aquilegia vulgaris, Kramer et al., 2007), maize (Ambrose et al., 2000), Streptochaeta angustifolia, Joinvillea ascendens, Elegia elephas (Whipple et al., 2007), oil palm (Elaeis guineensis, Adam et al., 2007), Asparagus officinalis (Park et al., 2003), and Aristolochia manshuriensis (Jaramillo and Kramer, 2004) (Fig. 5 and Table 1), and coded the observed pattern of AP3 expression during perianth development as 0: absent, 1: present, but weak, patchy or transient, or 2: present and ubiquitous from relatively early through later stages of flower development (Figs. 5, 6, and Table 1). Ancestral character states for AP3 gene expression were optimized using the maximum likelihood criterion as described. The maximum likelihood ancestral character state reconstruction for AP3 gene expression suggests that weak, patchy, or transient expression occurred in the perianth of the eudicot + monocot (node A), eudicot (node B) and core eudicot ancestor (node C, proportional likelihoods = 0.676, 0.688 and 0.604, respectively, Fig. 6; −log likelihoods = 19.60, 19.58, and 19.71 respectively, Table 2). For all three of these ancestral character state inferences, the alternative, that the ancestor ubiquitously expressed AP3 in the perianth, cannot be rejected (Table 2). It remains to be seen whether this trend toward weak, patchy, or transient expression of AP3 in the ancestors of these major angiosperm lineages holds up to further sampling and analysis. If so, it would suggest that before the radiation of these lineages, AP3 did not play a specific role in establishing discrete identities of perianth organs. We might further extrapolate that these ancestors developed flowers with an undifferentiated perianth and that AP3 function was recruited independently during the radiation of monocots and eudicots to specify distinct organ identities such as petals or lodicules. Interestingly, the results of our analyses based on AP3 functional and expression data are not in complete agreement. Although neither analysis provides a statistically significant result, the functional data suggest that the activity of AP3 in ancestral species was necessary for the specification of a bipartite perianth, whereas the expression data suggest that AP3 in ancestral species only functioned in specifying the identity of stamens (Figs. 4 and 6, Table 2). We consider functional data to be optimal for determining the ancestral roles of AP3 homologs during angiosperm evolution. However, we acknowledge that there are certainly tradeoffs in the choice of data. Ultimately, there will be limits in our ability to assay AP3 function in diverse nonmodel angiosperm species (although, see next section New tools to advance plant developmental evolutionary studies). On the other hand, there will be few factors limiting the accumulation of detailed expression data (i.e., in situ RNA hybridization studies) in diverse species, but there are clear difficulties associated with the accurate interpretation of AP3 function from patterns of gene expression. Detailed AP3 expression data representing all major angiosperm lineages would greatly benefit this type of ancestral state reconstruction, for instance, from a diversity of magnoliid dicots, monocots, and eudicots. However, of utmost interest would be expression data from early diverging core eudicot lineages and noncore eudicots that are more closely related to core eudicots than Ranunculales. For instance, there can be found a diversity of perianth phenotypes among the Berberidopsidales, Buxaceae, Trochodendraceae, Proteales, and Sabiaceae (Stevens, 2001). Increased sampling

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of AP3 expression data in these lineages would likely provide better resolution of ancestral states at the eudicot and core eudicot nodes. For drawing more accurate conclusions about AP3 function from expression data, a key contribution will be testing a major assumption of our analyses—that weak, patchy, and transient expression of AP3 correlates with a lack of organ identity specification in the perianth. The assumption that lack of ubiquitous AP3 expression in perianth organs is correlated with a lack of function in the specification of a bipartite perianth is based on comparisons of AP3 function and expression in poppy, tomato, and petunia. In all three species, there is a history of AP3 lineage gene duplication (Fig. 3), possibly affecting the observed co-occurrence. In other words, the observed co-occurrence may be complicated by the fact that these AP3 lineage genes have paralogs that are necessary for the specification of a bipartite perianth. It is worth noting, however, that functional data from Arabidopsis and snapdragon support our assumption that AP3 expression is required from early through late stages of perianth development to specify petal identity (Bowman et al., 1989; Zachgo et al., 1997). Ultimately, we hope that as more AP3 functional and expression data are generated and as assumptions outlined here are tested, ancestral state reconstructions based on either type of data will converge. New tools to advance plant developmental evolutionary studies— The initial discovery that AP3 lineage genes are homeotic regulators of petal and stamen identity in Arabidopsis and snapdragon resulted from traditional forward genetic approaches in these model species (Sommer et al., 1990; Jack et al., 1992). Although subsequent comparative studies of AP3 function have, to some extent, relied on forward genetics (e.g., petunia and maize, van der Krol et al., 1993; Whipple et al., 2004), it is mainly through the application of reverse genetic approaches that we have seen the field move so rapidly in recent years, with the functional characterization of AP3 homologs from flowering tobacco (Liu et al., 2004), tomato (de Martino et al., 2006), petunia (Rijpkema et al., 2006), poppy (Drea et al., 2007) and rice (Lee et al., 2003; Xiao et al., 2003). In species where stable transformation has proven to be an efficient tool for exogenous DNA delivery, for example, tomato (Fillatti et al., 1987) and petunia (Horsch et al., 1985), a reverse genetic approach can be taken by introducing constructs into plant genomes that express antisense or hairpin RNAs and elicit silencing of endogenous gene expression by RNA interference (RNAi) (Cogoni and Macino, 2000; Watson et al., 2005). The great efforts toward developing new plant model species are often associated with the development of stable transformation protocols. It is therefore promising that these efforts may allow us to apply stable reverse genetic strategies in a diversity of angiosperm species in the near future. However, even with the development of new model species, stable transformation may be limited by the difficulties associated with developing protocols that quickly and efficiently lead to large populations of transgenic plants. In addition, emerging

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model species may not always be the most appropriate for specific comparative developmental studies. Therefore, the ability to undertake functional analyses in nonmodel species is, and will continue to be, of great utility in the field of plant developmental evolution. On the forefront of nonmodel system reverse genetic approaches is the application of virus-induced gene silencing (VIGS) (Ruiz et al., 1998) in a number of plant species. VIGS technology takes advantage of homology-dependent viral defense mechanisms in plants. Viruses have been engineered that can target specific plant genes of interest for silencing (Burch-Smith et al., 2004; Robertson, 2004; Watson et al., 2005), with silencing likely occurring by similar mechanisms as RNAi (Robertson, 2004). Generally, viruses known to infect certain plant species have been genetically modified to induce VIGS in those, or closely related, plants. For example, Tobacco rattle virus-based (TRVbased) VIGS is used to silence genes in the close relatives tobacco (Ratcliff et al., 2001; Burch-Smith et al., 2004), tomato (Liu et al., 2002), and pepper, potato, and petunia (Ryu et al., 2004). Barley stripe mosaic virus-based VIGS has been used in barley (Oikawa et al., 2007) and wheat (Holzberg et al., 2002), while Cucumber mosaic virus has been modified to induce VIGS in soybean (Nagamatsu et al., 2007). Because of the wide host range of TRV (Robinson and Harrison, 1989), TRV-based VIGS was suggested to be a promising tool for gene silencing in species that are distant relatives of tobacco and tomato (Hileman et al., 2005). The first successful application of this kind was in the noncore eudicot species, opium poppy (Hileman et al., 2005), followed soon after by successful TRV-based VIGS in additional noncore eudicots—columbine (Gould and Kramer, 2007) and California poppy (Eschscholzia californica, Wege et al., 2007), as well as in Arabidopsis (Burch-Smith et al., 2006). Given the development of tools for stable transformation in emerging model species, in combination with new applications for VIGS in diverse nonmodel species, we are entering an era of plant developmental evolutionary biology that will allow us to critically dissect developmental genetic pathways in diverse angiosperm species. Moving beyond AP3: Exploring perianth developmental genetic modules— The AP3-dependent developmental pathway is an important prerequisite for the development of a bipartite perianth. What, then, is the genetic basis for the specification of diverse perianth phenotypes? To understand the evolution of distinct perianth phenotypes, we ultimately need to understand the developmental genetic modules that specify these organs. In Arabidopsis, a considerable amount of research has uncovered many of the AP3-dependent regulatory pathways involved in petal specification (Fig. 2, for further review, see Irish, 2008). Auto- and cross-regulatory feedback loops necessary for maintaining appropriate AP3 expression are important aspects of this developmental module in Arabidopsis (Fig. 2). Some of these autoregulatory loop controls may be conserved across core eudicots because similar direct autoregulatory feedback control of AP3 expression by AP3 protein appears to be con-



Fig. 5. Expression of AP3 lineage genes in a diversity of angiosperm species at relatively early and late stages of flower development. Representational drawings summarize published images from Jack et al. (1992); Schwarz-Sommer et al. (1992); Halfter et al. (1994); Ainsworth et al. (1995); Ambrose et al. (2000); Park et al. (2003); Jaramillo and Kramer (2004); Vandenbussche et al. (2004); de Martino et al. (2006); Adam et al. (2007); Drea et al. (2007); Kramer et al. (2007); Whipple et al. (2007). Strong expression (relative to expression in stamens) is indicated by dark red, weak expression (relative to expression in stamens) is indicated in pink. Arrowheads indicate AP3 homolog expression, strong or weak, in developing perianth organs, or in the floral meristem where perianth organs are expected to initiate.

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Fig. 6. Phylogeny of AP3 lineage genes that have been characterized by in situ mRNA hybridization expression analyses. At each node, a pie chart shows the proportional likelihood that the ancestor had a particular character state. White represents the proportional likelihood that AP3 in the ancestor was not expressed in perianth organs. Red represents the proportional likelihood that AP3 in the ancestor exhibited variable or weak expression, which co-occurs with AP3 gene products that do not function in specifying distinct perianth organ identities (unipartite perianth). Black represents the proportional likelihood that expression of AP3 in the ancestor was strong and ubiquitous in perianth organs from relatively early through later stages of flower development; this pattern of expression co-occurs with AP3 gene products that function in the specification of distinct perianth organ identities (bipartite perianth). Pie charts labeled A–D indicate the ancestor of eudicots + monocots, eudicots, core eudicots, and monocots, respectively. Gene relationships and branch lengths optimized under maximum likelihood using the GTR + Γ model of molecular evolution as implemented in GARLI v0.951 (Zwickl, 2006). Numbers above branches indicate node support based on 500 maximum likelihood bootstrap replicates. Asterisks indicate nodes at which a single character state with the largest proportional likelihood is significantly preferred over alternative character states.

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served in snapdragon and petunia (Schwarz-Sommer et al., 1992; Halfter et al., 1994). However, this aspect of autoregulation may not be conserved outside of core eudicots, because opium poppy AP3 genes do not appear to be subject to the same regulatory processes (Drea et al., 2007). Furthermore, AP3 protein homodimerization, which may be important for regulating specific downstream components in the petal developmental pathway, appears to have been lost early in core eudicot evolution (Winter et al., 2002). Together, these observations suggest that while an AP3-dependent pathways may be necessary for specifying different perianth morphologies, it is likely that the postulated independent evolution of different kinds of perianth organs (Hiepko, 1965; Takhtajan, 1991) have relied on considerable regulatory remodeling of this pathway. One might expect that in species hypothesized to have bracteopetals—most core eudicots, Gunnerales, Proteales, and some Ranunculales, the underlying petal developmental module would be quite distinct from species hypothesized to develop andropetals—some rosids, Caryophyllales, and some Ranunculales (Ronse De Craene, 2007). Ultimately, the evolution of such developmental genetic modules should be explored in the type of strict phylogenetic framework outlined in this paper, with ancestral states being reconstructed for characters including protein function, gene expression, auto- and cross-regulation, and protein–protein interactions. Summary— Here we have presented a phylogenetic framework for integrating developmental genetic data into our understanding of morphological character evolution. We have focused specifically on the expression and function of AP3 and its role in perianth development in a diversity of angiosperms as an exemplar for such studies. As additional AP3 functional and expression data are amassed, it will help resolve the questions posed here. We are hopeful that, as more data are gathered, evidence from gene expression, protein function, and morphology will converge on a well-supported answer to the question of the evolution of differentiated perianth morphology across angiosperms. Key taxa for future sampling should include both monocots and eudicots that have a diversity of perianth phenotypes (Soltis et al., 2005). Functional tools are being developed that hold promise for such future studies. These tools will be invaluable for moving beyond studies of AP3 to consideration of the evolution of the entire petal module. A new approach is needed to understand how and when a bipartite perianth has evolved and the genetic basis for the diversity of perianth organs found in angiosperms. LITERATURE CITED Aagaard, J. E., R. G. Olmstead, J. H. Willis, and P. C. Phillips. 2005. Duplication of floral regulatory genes in the Lamiales. American Journal of Botany 92: 1284–1293. Abouheif, E. 1997. Developmental genetics and homology: A hierarchical approach. Trends in Ecology & Evolution 12: 405–408. Adam, H., S. Jouannic, Y. Orieux, F. Morcillo, F. Richaud, Y. Duval, and J. W. Tregear. 2007. Functional characterization of MADS box genes involved in the determination of oil palm flower structure. Journal of Experimental Botany 58: 1245–1259. Ainsworth, C., S. Crossley, V. Buchanan-Wollaston, M. Thangavelu, and J. Parker. 1995. Male and female flowers of the dioecious plant sorrel show different patterns of MADS box gene expression. Plant Cell 7: 1583–1598. Ambrose, B. A., D. R. Lerner, P. Ciceri, C. M. Padilla, M. F. Yanofsky, and R. J. Schmidt. 2000. Molecular and genetic analy-

93

ses of the Silky1 gene reveal conservation in floral organ specification between eudicots and monocots. Molecular Cell 5: 569–579. Baum, D. A., and B. A. Whitlock. 1999. Plant development: Genetic clues to petal evolution. Current Biology 9: R525–R527. Bowman, J. L., D. R. Smyth, and E. M. Meyerowitz. 1989. Genes directing flower development in Arabidopsis. Plant Cell 1: 37–52. Brigandt, I. 2003. Homology in comparative, molecular, and evolutionary developmental biology: The radiation of a concept. Journal of Experimental Zoology, B, Molecular and Developmental Evolution 299B: 9–17. Burch-Smith, T. M., J. C. Anderson, G. B. Martin, and S. P. DineshKumar. 2004. Applications and advantages of virus-induced gene silencing for gene function studies in plants. Plant Journal 39: 734–746. Burch-Smith, T. M., M. Schiff, Y. L. Liu, and S. P. Dinesh-Kumar. 2006. Efficient virus-induced gene silencing in Arabidopsis. Plant Physiology 142: 21–27. Carpenter, R., and E. S. Coen. 1990. Floral homeotic mutations produced by transposon-mutagenesis in Antirrhinum majus. Genes & Development 4: 1483–1493. Case, A. L., S. W. Graham, T. D. Macfarlane, and S. C. H. Barrett. 2008. A phylogenetic study of evolutionary transitions in sexual systems in Australasian Wurmbea (Colchicaceae). International Journal of Plant Sciences 169: 141–156. Chae, E., Q. K.-G. Tan, T. A. Hill, and V. F. Irish. 2008. An Arabidopsis F-box protein acts as a transcriptional co-factor to regulate floral development. Development 135: 1235–1245. Cogoni, C., and G. Macino. 2000. Post-transcriptional gene silencing across kingdoms. Current Opinion in Genetics & Development 10: 638–643. Cunningham, C. W., K. E. Omland, and T. H. Oakley. 1998. Reconstructing ancestral character states: A critical reappraisal. Trends in Ecology & Evolution 13: 361–366. de Martino, G., I. Pan, E. Emmanuel, A. Levy, and V. F. Irish. 2006. Functional analyses of two tomato APETALA3 genes demonstrate diversification in their roles in regulating floral development. Plant Cell 18: 1833–1845. Drea, S., L. C. Hileman, G. de Martino, and V. F. Irish. 2007. Functional analyses of genetic pathways controlling petal specification in poppy. Development 134: 4157–4166. Edwards, A. F. 1972. Likelihood. Cambridge University Press, Cambridge, UK. Endress, P. K. 1994. Floral structure and evolution of primitive angiosperms: Recent advances. Plant Systematics and Evolution 192: 79–97. Endress, P. K. 2006. Angiosperm floral evolution: Morphological developmental framework. Advances in Botanical Research 44: 1–61. Fillatti, J., J. Kiser, B. Rose, and L. Comai. 1987. Efficient transformation of tomato and the introduction and expression of a gene for herbicide tolerance. In D. J. Nevins and R. A. Jones [eds.], Tomato biotechnology: Proceedings of a symposium held at the University of California, Davis, California, August 20-22, 1986, 199–210. A.R. Liss, New York, New York, USA. Force, A., W. A. Cresko, F. B. Pickett, S. R. Proulx, C. Amemiya, and M. Lynch. 2005. The origin of subfunctions and modular gene regulation. Genetics 170: 433–446. Force, A., M. Lynch, F. B. Pickett, A. Amores, Y. L. Yan, and J. Postlethwait. 1999. Preservation of duplicate genes by complementary, degenerative mutations. Genetics 151: 1531–1545. Goto, K., and E. M. Meyerowitz. 1994. Function and regulation of the Arabidopsis floral homeotic gene PISTILLATA. Genes & Development 8: 1548–1560. Gould, B. A., and E. M. Kramer. 2007. Virus-induced gene silencing as a tool for functional analyses in the emerging model plant Aquilegia (columbine, Ranunculaceae). BMC Plant Methods 3: 6. Guo, Y. F., and S. S. Gan. 2006. AtNAP, a NAC family transcription factor, has an important role in leaf senescence. Plant Journal 46: 601–612. Halfter, U., N. Ali, J. Stockhaus, L. Ren, and N.-H. Chua. 1994. Ectopic expression of a single homeotic gene, the Petunia gene

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green petal, is sufficient to convert sepals to petaloid organs. EMBO Journal 13: 1443–1449. Hiepko, P. 1965. Vergleichend-morphologische und entwicklungsgeschichtliche Untersuchungen über das Perianth bei den Polycarpicae. Botanische Jarhbücher für Systematik. Pflanzengeschichte und Pflanzengeographie 84: 360–363. Hileman, L. C., S. Drea, G. de Martino, A. Litt, and V. F. Irish. 2005. Virus-induced gene silencing is an effective tool for assaying gene function in the basal eudicot species Papaver somniferum (opium poppy). Plant Journal 44: 334–341. Hill, T. A., C. D. Day, S. C. Zondlo, A. G. Thackeray, and V. F. Irish. 1998. Discrete spatial and temporal cis-acting elements regulate transcription of the Arabidopsis floral homeotic gene APETALA3. Development 125: 1711–1721. Holzberg, S., P. Brosio, C. Gross, and G. P. Pogue. 2002. Barley stripe mosaic virus-induced gene silencing in a monocot plant. Plant Journal 30: 315–327. Honma, T., and K. Goto. 2000. The Arabidopsis floral homeotic gene PISTILLATA is regulated by discrete cis-elements responsive to induction and maintenance signals. Development 127: 2021–2030. Horsch, R. B., J. E. Fry, N. L. Hoffmann, D. Eichholtz, S. G. Rogers, and R. T. Fraley. 1985. A simple and general method for transferring genes into plants. Science 227: 1229–1231. Irish, V. F. 2008. The Arabidopsis petal: A model for plant organogenesis. Trends in Plant Science 13: 430–436. Irish, V. F., and I. M. Sussex. 1990. Function of the apetala-1 gene during Arabidopsis floral development. Plant Cell 2: 741–753. Jack, T., L. L. Brockman, and E. M. Meyerowitz. 1992. The homeotic gene APETALA3 of Arabidopsis thaliana encodes a MADS box and is expressed in petals and stamens. Cell 68: 683–697. Jack, T., G. L. Fox, and E. M. Meyerowitz. 1994. Arabidopsis homeotic gene APETALA3 ectopic expression: Transcriptional and posttranscriptional regulation determine floral organ identity. Cell 76: 703–716. Jaramillo, M. A., and E. M. Kramer. 2004. APETALA3 and PISTILLATA homologs exhibit novel expression patterns in the unique perianth of Aristolochia (Aristolochiaceae). Evolution & Development 6: 449–458. Jaramillo, M. A., and E. M. Kramer. 2007. The role of developmental genetics in understanding homology and morphological evolution in plants. International Journal of Plant Sciences 168: 61–72. Jenik, P. D., and V. F. Irish. 2001. The Arabidopsis floral homeotic gene APETALA3 differentially regulates intercellular signaling required for petal and stamen development. Development 128: 13–23. Kim, S., J. Koh, M. J. Yoo, H. Z. Kong, Y. Hu, H. Ma, P. S. Soltis, and D. E. Soltis. 2005. Expression of floral MADS-box genes in basal angiosperms: Implications for the evolution of floral regulators. Plant Journal 43: 724–744. Kramer, E. M., V. S. Di Stilio, and P. M. Schluter. 2003. Complex patterns of gene duplication in the APETALA3 and PISTILLATA lineages of the Ranunculaceae. International Journal of Plant Sciences 164: 1–11. Kramer, E. M., R. L. Dorit, and V. F. Irish. 1998. Molecular evolution of genes controlling petal and stamen development: Duplication and divergence within the APETALA3 and PISTILLATA MADS-box gene lineages. Genetics 149: 765–783. Kramer, E. M., L. Holappa, B. Gould, M. A. Jaramillo, D. Setnikov, and P. M. Santiago. 2007. Elaboration of B gene function to include the identity of novel floral organs in the lower eudicot Aquilegia. Plant Cell 19: 750–766. Lamb, R. S., T. A. Hill, Q. K. G. Tan, and V. F. Irish. 2002. Regulation of APETALA3 floral homeotic gene expression by meristem identity genes. Development 129: 2079–2086. Lamb, R. S., and V. F. Irish. 2003. Functional divergence within the APETALA3/PISTILLATA floral homeotic gene lineages. Proceedings of the National Academy of Sciences, USA 100: 6558–6563. Lee, I., D. S. Wolfe, O. Nilsson, and D. Weigel. 1997. A LEAFY coregulator encoded by UNUSUAL FLORAL ORGANS. Current Biology 7: 95–104.

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Lee, S., J. S. Jeon, K. An, Y. H. Moon, S. Lee, Y. Y. Chung, and G. An. 2003. Alteration of floral organ identity in rice through ectopic expression of OsMADS16. Planta 217: 904–911. Liu, Y., N. Nakayama, M. Schiff, A. Litt, V. F. Irish, and S. P. DineshKumar. 2004. Virus induced gene silencing of a DEFICIENS ortholog in Nicotiana benthamiana. Plant Molecular Biology 54: 701–711. Liu, Y. L., M. Schiff, and S. P. Dinesh-Kumar. 2002. Virus-induced gene silencing in tomato. Plant Journal 31: 777–786. Losos, J. B. 1999. Uncertainty in the reconstruction of ancestral character states and limitations on the use of phylogenetic comparative methods. Animal Behaviour 58: 1319–1324. Maddison, W. P., and D. R. Maddison. 2006. StochChar: A package of Mesquite modules for stochastic models of character evolution, version 1.1. Website http://mesquiteproject.org. Maddison, W. P., and D. R. Maddison. 2007. Mesquite: A modular system for evolutionary analysis, version 2.01. Website http://mesquiteproject.org. Mandel, M. A., C. Gustafson-Brown, B. Savidge, and M. F. Yanofsky. 1992. Molecular characterization of the Arabidopsis floral homeotic gene APETALA1. Nature 360: 273–277. Mara, C. D., and V. F. Irish. 2008. Two GATA transcription factors are downstream effectors of floral homeotic gene action in Arabidopsis. Plant Physiology. 147: 707–718. Moore, M. J., C. D. Bell, P. S. Soltis, and D. E. Soltis. 2007. Using plastid genome-scale data to resolve enigmatic relationships among basal angiosperms. Proceedings of the National Academy of Sciences, USA 104: 19363–19368. Nagamatsu, A., C. Masuta, M. Senda, H. Matsuura, A. Kasai, S. Hong, K. Kitamura, J. Abe, and A. Kanazawa. 2007. Functional analysis of soybean genes involved in flavonoid biosynthesis by virusinduced gene silencing. Plant Biotechnology Journal 5: 778–790. Nagasawa, N., M. Miyoshi, Y. Sano, H. Satoh, H. Hirano, H. Sakai, and Y. Nagato. 2003. SUPERWOMAN1 and DROOPING LEAF genes control floral organ identity in rice. Development 130: 705–718. Ng, M., and M. F. Yanofsky. 2001. Activation of the Arabidopsis B class homeotic genes by APETALA1. Plant Cell 13: 739–753. Ohno, S. 1970. Evolution by gene duplication. Springer-Verlag, Heidelberg, Germany. Oikawa, A., A. Rahman, T. Yamashita, H. Taira, and S. Kidou. 2007. Virus-induced gene silencing of P23k in barley leaf reveals morphological changes involved in secondary wall formation. Journal of Experimental Botany 58: 2617–2625. Pagel, M. 1999. The maximum likelihood approach to reconstructing ancestral character states of discrete characters on phylogenies. Systematic Biology 48: 612–622. Park, J. H., Y. Ishikawa, R. Yoshida, A. Kanno, and T. Kameya. 2003. Expression of AODEF, a B-functional MADS-box gene, in stamens and inner tepals of the dioecious species Asparagus officinalis L. Plant Molecular Biology 51: 867–875. Prasad, K., and U. Vijayraghavan. 2003. Double-stranded RNA interference of a rice PI/GLO paralog, OsMADS2, uncovers its second-whorlspecific function in floral organ patterning. Genetics 165: 2301–2305. Rasmussen, D. A., E. M. Kramer, and E. A. Zimmer. 2009. One size fits all? Molecular evidence for a commonly inherited petal identity program in Ranunculales. American Journal of Botany 96: 96–109. Ratcliff, F., A. M. Martin-Hernandez, and D. C. Baulcombe. 2001. Tobacco rattle virus as a vector for analysis of gene function by silencing. Plant Journal 25: 237–245. Regal, P. J. 1977. Ecology and evolution of flowering plant dominance. Science 196: 622–629. Rijpkema, A. S., S. Royaert, J. Zethof, G. van der Weerden, T. Gerats, and M. Vandenbussche. 2006. Analysis of the Petunia TM6 MADS box gene reveals functional divergence within the DEF/ AP3 lineage. Plant Cell 18: 1819–1832. Robertson, D. 2004. VIGS vectors for gene silencing: Many targets, many tools. Annual Review of Plant Biology 55: 495–519.

January 2009]

Hileman and Irish—Perianth evolution and development

Robinson, D. J., and B. D. Harrison. 1989. Tobacco rattle virus. CMI/ AAB Descriptions of Plant Viruses no. 346, Commonwealth Mycological Institute, Kew, UK. Ronse De Craene, L. P. 2007. Are petals sterile stamens or bracts? The origin and evolution of petals in the core eudicots. Annals of Botany 100: 621–630. Ronse De Craene, L. P. 2008. Homology and evolution of petals in the core eudicots. Systematic Botany 33: 301–325. Ruiz, M. T., O. Voinnet, and D. C. Baulcombe. 1998. Initiation and maintenance of virus-induced gene silencing. Plant Cell 10: 937–946. Ryu, C. M., A. Anand, L. Kang, and K. S. Mysore. 2004. Agrodrench: A novel and effective agroinoculation method for virus-induced gene silencing in roots and diverse solanaceous species. Plant Journal 40: 322–331. Sablowski, R. W. M., and E. M. Meyerowitz. 1998. A homolog of NO APICAL MERISTEM is an immediate target of the floral homeotic genes APETALA3/PISTILLATA. Cell 92: 93–103. Schluter, D., T. Price, A. O. Mooers, and D. Ludwig. 1997. Likelihood of ancestor states in adaptive radiation. Evolution 51: 1699–1711. Schwarz-Sommer, Z., I. Hue, P. Huijser, P. J. Flor, R. Hansen, F. Tetens, W. E. Lönnig, et al. 1992. Characterization of the Antirrhinum floral homeotic MADS-box gene deficiens: Evidence for DNA-binding and autoregulation of its persistent expression throughout flower development. EMBO Journal 11: 251–263. Soltis, D. E., P. E. Soltis, P. K. Endress, and M. W. Chase. 2005. Phylogeny and evolution of angiosperms. Sinauer, Sunderland, Massachusetts, USA. Sommer, H., J. Beltrán, P. Huijser, H. Pape, W. Lönnig, H. Saedler, and Z. Schwarz-Sommer. 1990. Deficiens, a homeotic gene involved in the control of flower morphogenesis in Antirrhinum majus: The protein shows homology to transcription factors. EMBO Journal 9: 605–613. Stellari, G. M., M. A. Jaramillo, and E. M. Kramer. 2004. Evolution of the APETALA3 and PISTILLATA lineages of MADS-box-containing genes in the basal angiosperms. Molecular Biology and Evolution 21: 506–519. Stevens, P. F. 2001 [onward]. Angiosperm Phylogeny Website, version 8, June 2007 [more or less continuously updated]. Website http://www. mobot.org/MOBOT/research/APweb. Sundstrom, J. F., N. Nakayama, K. Glimelius, andV. F. Irish. 2006. Direct regulation of the floral homeotic APETALA1 gene by APETALA3 and PISTILLATA in Arabidopsis. Plant Journal 46: 593–600. Takhtajan, A. 1991. Evolutionary trends in flowering plants. Columbia University Press, New York, New York, USA. Tilly, J. J., D. W. Allen, and T. Jack. 1998. The CArG boxes in the promoter of the Arabidopsis floral organ identity gene APETALA3 mediate diverse regulatory effects. Development 125: 1647–1657. van der Krol, A. R., A. Brunelle, S. Tsuchimoto, and N.-H. Chua. 1993. Functional analysis of petunia floral homeotic MADS box gene pMADS1. Genes & Development 7: 1214–1228. Vandenbussche, M., J. Zethof, S. Royaert, K. Weterings, and T. Gerats. 2004. The duplicated B-class heterodimer model: Whorl-specific

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effects and complex genetic interactions in Petunia hybrida flower development. Plant Cell 16: 741–754. Watson, J. M., A. F. Fusaro, M. B. Wang, and P. M. Waterhouse. 2005. RNA silencing platforms in plants. FEBS Letters 579: 5982–5987. Wege, S., A. Scholz, S. Gleissberg, and A. Becker. 2007. Highly efficient virus-induced gene silencing (VIGS) in California poppy (Eschscholzia californica): An evaluation of VIGS as a strategy to obtain functional data from non-model plants. Annals of Botany 100: 641–649. Whipple, C. J., P. Ciceri, C. M. Padilla, B. A. Ambrose, S. L. Bandong, and R. J. Schmidt. 2004. Conservation of B-class floral homeotic gene function between maize and Arabidopsis. Development 131: 6083–6091. Whipple, C. J., M. J. Zanis, E. A. Kellogg, and R. J. Schmidt. 2007. Conservation of B class gene expression in the second whorl of a basal grass and outgroups links the origin of lodicules and petals. Proceedings of the National Academy of Sciences, USA 104: 1081–1086. Winter, K. U., C. Weiser, K. Kaufmann, A. Bohne, C. Kirchner, A. Kanno, H. Saedler, and G. Theissen. 2002. Evolution of class B floral homeotic proteins: Obligate heterodimerization originated from homodimerization. Molecular Biology and Evolution 19: 587–596. Xiao, H., Y. Wang, D. F. Liu, W. M. Wang, X. B. Li, X. F. Zhao, J. C. Xu, W. X. Zhai, and L. H. Zhu. 2003. Functional analysis of the rice AP3 homologue OsMADS16 by RNA interference. Plant Molecular Biology 52: 957–966. Yang, Z. 1994a. Estimating the pattern of nucleotide substitution. Journal of Molecular Evolution 39: 105–111. Yang, Z. 1994b. Maximum likelihood phylogenetic estimation from DNA sequences with variable rates over sites: approximate methods. Journal of Molecular Evolution 39: 306–314. Zachgo, S., H. Saedler, and Z. Schwarz-Sommer. 1997. Pollenspecific expression of DEFH125, a MADS-box transcription factor in Antirrhinum with unusual features. Plant Journal 11: 1043–1050. Zachgo, S., E. D. Silva, P. Motte, W. Trobner, H. Saedler, and Z. Schwarz-Sommer. 1995. Functional-analysis of the Antirrhinum floral homeotic DEFICIENS gene in-vivo and in-vitro by using a temperature-sensitive mutant. Development 121: 2861–2875. Zahn, L. M., J. Leebens-Mack, C. W. dePamphilis, H. Ma, and G. Theissen. 2005. To B or not to B a flower: The role of DEFICIENS and GLOBOSA orthologs in the evolution of the angiosperms. Journal of Heredity 96: 225–240. Zanis, M. J., P. S. Soltis, Y. L. Qiu, E. Zimmer, and D. E. Soltis. 2003. Phylogenetic analyses and perianth evolution in basal angiosperms. Annals of the Missouri Botanical Garden 90: 129–150. Zwickl, D. J. 2006. Genetic algorithm approaches for the phylogenetic analysis of large biological datasets under the maximum likelihood criterion. Ph.D. dissertation, University of Texas at Austin, Austin, Texas, USA.

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