Chromosomal evolution, environmental heterogeneity, and migration drive spatial patterns of species richness in Calochortus (Liliaceae).

We used nuclear genomic data and statistical models to evaluate the ecological and evolutionary processes shaping spatial variation in species richness in Calochortus (Liliaceae, 74 spp.). Calochortus occupies diverse habitats in the western United States and Mexico and has a center of diversity in the California Floristic Province, marked by multiple orogenies, winter rainfall, and highly divergent climates and substrates (including serpentine). We used sequences of 294 low-copy nuclear loci to produce a time-calibrated phylogeny, estimate historical biogeography, and test hypotheses regarding drivers of present-day spatial patterns in species number. Speciation and species coexistence require reproductive isolation and ecological divergence, so we examined the roles of chromosome number, environmental heterogeneity, and migration in shaping local species richness. Six major clades-inhabiting different geographic/climatic areas, and often marked by different base chromosome numbers (n = 6 to 10)-began diverging from each other ~10.3 Mya. As predicted, local species number increased significantly with local heterogeneity in chromosome number, elevation, soil characteristics, and serpentine presence. Species richness is greatest in the Transverse/Peninsular Ranges where clades with different chromosome numbers overlap, topographic complexity provides diverse conditions over short distances, and several physiographic provinces meet allowing immigration by several clades. Recently diverged sister-species pairs generally have peri-patric distributions, and maximum geographic overlap between species increases over the first million years since divergence, suggesting that chromosomal evolution, genetic divergence leading to gametic isolation or hybrid inviability/sterility, and/or ecological divergence over small spatial scales may permit species co-occurrence.

Unearthing Modes of Climatic Adaptation in Underground Storage Organs Across Liliales.

Testing adaptive hypotheses about how continuous traits evolve in association with developmentally structured discrete traits, while accounting for the confounding influence of other, hidden, evolutionary forces, remains a challenge in evolutionary biology. For example, geophytes are herbaceous plants-with underground buds-that use underground storage organs (USOs) to survive extended periods of unfavorable conditions. Such plants have evolved multiple times independently across all major vascular plant lineages. Even within closely related lineages, however, geophytes show impressive variation in the morphological modifications and structures (i.e.,"types" of USOs) that allow them to survive underground. Despite the developmental and structural complexity of USOs, the prevailing hypothesis is that they represent convergent evolutionary "solutions" to a common ecological problem, though some recent research has drawn this conclusion into question. We extend existing phylogenetic comparative methods to test for links between the hierarchical discrete morphological traits associated with USOs and adaptation to environmental variables, using a phylogeny of 621 species in Liliales. We found that plants with different USO types do not differ in climatic niche more than expected by chance, with the exception of root morphology, where modified roots are associated with lower temperature seasonality. These findings suggest that root tubers may reflect adaptations to different climatic conditions than those represented by other types of USOs. Thus, the tissue type and developmental origin of the USO structure may influence the way it mediates ecological relationships, which draws into question the appropriateness of ascribing broad ecological patterns uniformly across geophytic taxa. This work provides a new framework for testing adaptive hypotheses and for linking ecological patterns across morphologically varying taxa while accounting for developmental (non-independent) relationships in morphological data. [Climatic niche evolution; geophytes; imperfect correspondence; macroevolution.].

Orthology-based analysis helps map evolutionary diversification and predict substrate class use of BAHD acyltransferases.

Large enzyme families catalyze metabolic diversification by virtue of their ability to use diverse chemical scaffolds. How enzyme families attain such functional diversity is not clear. Furthermore, duplication and promiscuity in such enzyme families limits their functional prediction, which has produced a burgeoning set of incompletely annotated genes in plant genomes. Here, we address these challenges using BAHD acyltransferases as a model. This fast-evolving family expanded drastically in land plants, increasing from one to five copies in algae to approximately 100 copies in diploid angiosperm genomes. Compilation of >160 published activities helped visualize the chemical space occupied by this family and define eight different classes based on structural similarities between acceptor substrates. Using orthologous groups (OGs) across 52 sequenced plant genomes, we developed a method to predict BAHD acceptor substrate class utilization as well as origins of individual BAHD OGs in plant evolution. This method was validated using six novel and 28 previously characterized enzymes and helped improve putative substrate class predictions for BAHDs in the tomato genome. Our results also revealed that while cuticular wax and lignin biosynthetic activities were more ancient, anthocyanin acylation activity was fixed in BAHDs later near the origin of angiosperms. The OG-based analysis enabled identification of signature motifs in anthocyanin-acylating BAHDs, whose importance was validated via molecular dynamic simulations, site-directed mutagenesis and kinetic assays. Our results not only describe how BAHDs contributed to evolution of multiple chemical phenotypes in the plant world but also propose a biocuration-enabled approach for improved functional annotation of plant enzyme families.

Putative adaptive loci show parallel clinal variation in a California-endemic wildflower.

As the global climate crisis continues, predictions concerning how wild populations will respond to changing climate conditions are informed by an understanding of how populations have responded and/or adapted to climate variables in the past. Changes in the local biotic and abiotic environment can drive differences in phenology, physiology, morphology and demography between populations leading to local adaptation, yet the molecular basis of adaptive evolution in wild non-model organisms is poorly understood. We leverage comparisons between two lineages of Calochortus venustus occurring along parallel transects that allow us to identify loci under selection and measure clinal variation in allele frequencies as evidence of population-specific responses to selection along climatic gradients. We identify targets of selection by distinguishing loci that are outliers to population structure and by using genotype-environment associations across transects to detect loci under selection from each of nine climatic variables. Despite gene flow between individuals of different floral phenotypes and between populations, we find evidence of ecological specialization at the molecular level, including genes associated with key plant functions linked to plant adaptation to California's Mediterranean climate. Single-nucleotide polymorphisms (SNPs) present in both transects show similar trends in allelic similarity across latitudes indicating parallel adaptation to northern climates. Comparisons between eastern and western populations across latitudes indicate divergent genetic evolution between transects, suggesting local adaptation to either coastal or inland habitats. Our study is among the first to show repeated allelic variation across climatic clines in a non-model organism.

Comparative transcriptomics of a monocotyledonous geophyte reveals shared molecular mechanisms of underground storage organ formation.

Many species from across the vascular plant tree-of-life have modified standard plant tissues into tubers, bulbs, corms, and other underground storage organs (USOs), unique innovations which allow these plants to retreat underground. Our ability to understand the developmental and evolutionary forces that shape these morphologies is limited by a lack of studies on certain USOs and plant clades. We take a comparative transcriptomics approach to characterizing the molecular mechanisms of tuberous root formation in Bomarea multiflora (Alstroemeriaceae) and compare these mechanisms to those identified in other USOs across diverse plant lineages; B. multiflora fills a key gap in our understanding of USO molecular development as the first monocot with tuberous roots to be the focus of this kind of research. We sequenced transcriptomes from the growing tip of four tissue types (aerial shoot, rhizome, fibrous root, and root tuber) of three individuals of B. multiflora. We identified differentially expressed isoforms between tuberous and non-tuberous roots and tested the expression of a priori candidate genes implicated in underground storage in other taxa. We identify 271 genes that are differentially expressed in root tubers versus non-tuberous roots, including genes implicated in cell wall modification, defense response, and starch biosynthesis. We also identify a phosphatidylethanolamine-binding protein, which has been implicated in tuberization signalling in other taxa and, through gene-tree analysis, place this copy in a phylogenetic context. These findings suggest that some similar molecular processes underlie the formation of USOs across flowering plants despite the long evolutionary distances among taxa and non-homologous morphologies (e.g., bulbs vs. tubers). (Plant development, tuberous roots, comparative transcriptomics, geophytes).

A natural diversity screen in Arabidopsis thaliana reveals determinants for HopZ1a recognition in the ZAR1-ZED1 immune complex.

Pathogen pressure on hosts can lead to the evolution of genes regulating the innate immune response. By characterizing naturally occurring polymorphisms in immune receptors, we can better understand the molecular determinants of pathogen recognition. ZAR1 is an ancient Arabidopsis thaliana NLR (Nucleotide-binding [NB] Leucine-rich-repeat [LRR] Receptor) that recognizes multiple secreted effector proteins from the pathogenic bacteria Pseudomonas syringae and Xanthomonas campestris through its interaction with receptor-like cytoplasmic kinases (RLCKs). ZAR1 was first identified for its role in recognizing P. syringae effector HopZ1a, through its interaction with the RLCK ZED1. To identify additional determinants of HopZ1a recognition, we performed a computational screen for ecotypes from the 1001 Genomes project that were likely to lack HopZ1a recognition, and tested ~300 ecotypes. We identified ecotypes containing polymorphisms in ZAR1 and ZED1. Using our previously established Nicotiana benthamiana transient assay and Arabidopsis ecotypes, we tested for the effect of naturally occurring polymorphisms on ZAR1 interactions and the immune response. We identified key residues in the NB or LRR domain of ZAR1 that impact the interaction with ZED1. We demonstrate that natural diversity combined with functional assays can help define the molecular determinants and interactions necessary to regulate immune induction in response to pathogens.

Get the shovel: morphological and evolutionary complexities of belowground organs in geophytes.

Herbaceous plants collectively known as geophytes, which regrow from belowground buds, are distributed around the globe and throughout the land plant tree of life. The geophytic habit is an evolutionarily and ecologically important growth form in plants, permitting novel life history strategies, enabling the occupation of more seasonal climates, mediating interactions between plants and their water and nutrient resources, and influencing macroevolutionary patterns by enabling differential diversification and adaptation. These taxa are excellent study systems for understanding how convergence on a similar growth habit (i.e., geophytism) can occur via different morphological and developmental mechanisms. Despite the importance of belowground organs for characterizing whole-plant morphological diversity, the morphology and evolution of these organs have been vastly understudied with most research focusing on only a few crop systems. Here, we clarify the terminology commonly used (and sometimes misused) to describe geophytes and their underground organs and highlight key evolutionary patterns of the belowground morphology of geophytic plants. Additionally, we advocate for increasing resources for geophyte research and implementing standardized ontological definitions of geophytic organs to improve our understanding of the factors controlling, promoting, and maintaining geophyte diversity.

Revisiting floral fusion: the evolution and molecular basis of a developmental innovation.

Throughout the evolution of the angiosperm flower, developmental innovations have enabled the modification or elaboration of novel floral organs enabling subsequent diversification and expansion into new niches, for example the formation of novel pollinator relationships. One such developmental innovation is the fusion of various floral organs to form complex structures. Multiple types of floral fusion exist; each type may be the result of different developmental processes and is likely to have evolved multiple times independently across the angiosperm tree of life. The development of fused organs is thought to be mediated by the NAM/CUC3 subfamily of NAC transcription factors, which mediate boundary formation during meristematic development. The goal of this review is to (i) introduce the development of fused floral organs as a key 'developmental innovation', facilitated by a change in the expression of NAM/CUC3 transcription factors; (ii) provide a comprehensive overview of floral fusion phenotypes amongst the angiosperms, defining well-known fusion phenotypes and applying them to a systematic context; and (iii) summarize the current molecular knowledge of this phenomenon, highlighting the evolution of the NAM/CUC3 subfamily of transcription factors implicated in the development of fused organs. The need for a network-based analysis of fusion is discussed, and a gene regulatory network responsible for directing fusion is proposed to guide future research in this area.

Phylogeny of Paullinia L. (Paullinieae: Sapindaceae), a diverse genus of lianas with dynamic fruit evolution.

Paullinia L. is a genus of c. 220 mostly Neotropical forest-dwelling lianas that display a wide diversity of fruit morphologies. Paullinia resembles other members of the Paullinieae tribe in being a climber with stipulate compound leaves and paired inflorescence tendrils. However, it is distinct in having capsular fruits with woody, coriaceous, or crustaceous pericarps. While consistent in this basic plan, the pericarps of Paullinia fruits are otherwise highly variable-in some species they are winged, whereas in others they are without wings or covered with spines. With the exception of the water-dispersed indehiscent spiny fruits of some members of Paullinia sect. Castanella, all species are dehiscent, opening their capsules while they are still attached to the branch, to reveal arillate animal-dispersed seeds. Here we present a molecular phylogeny of Paullinia derived from 11 molecular markers, including nine newly-developed single-copy nuclear markers amplified by microfluidics PCR. This is the first broadly sampled molecular phylogeny for the genus. Paullinia is supported as monophyletic and is sister to Cardiospermum L., which together are sister to Serjania Mill + Urvillea Kunth. We apply this novel phylogenetic hypothesis to test previous infrageneric classifications and to infer that unwinged fruits represent the ancestral condition, from which there were repeated evolutionary transitions and reversals. However, because the seeds of both winged and unwinged fruits are dispersed by animals, we conclude that the repeated transitions in fruit morphology may relate to visual display strategies to attract animal dispersers, and do not represent transitions to wind dispersal.

Pleistocene diversification in an ancient lineage: a role for glacial cycles in the evolutionary history of Dioon Lindl. (Zamiaceae).

PREMISE OF THE STUDY: Recent estimates of crown ages for cycad genera (Late Miocene) challenge us to consider what processes have produced the extant diversity of this ancient group in such relatively little time. Pleistocene climate change has driven major shifts in species distributions in Mexico and may have led to speciation in the genus Dioon by forcing populations to migrate up in elevation, thereby becoming separated by topography. METHODS: We inferred orthologs from transcriptomes of five species and sequenced these in 42 individuals representing all Dioon species. From these data and published plastid sequences, we inferred dated species trees and lineage-specific diversification rates. KEY RESULTS: Analyses of 84 newly sequenced nuclear orthologs and published plastid data confirm four major clades within Dioon, all of Pleistocene age. Gene tree analysis, divergence dates, and an increase in diversification rate support very recent and rapid divergence of extant taxa. CONCLUSIONS: This study confirms the Pleistocene age of Dioon species and implicates Pleistocene climate change and established topography in lineage spitting. These results add to our understanding of the cycads as evolutionarily dynamic lineages, not relicts or evolutionary dead ends. We also find that well-supported secondary calibration points can be reliable in the absence of fossils. Our hypothesis of lineage splitting mediated by habitat shifts may be applicable to other taxa that are restricted to elevation specific ecotones.

Building the monocot tree of death: Progress and challenges emerging from the macrofossil-rich Zingiberales.

PREMISE OF THE STUDY: Inclusion of fossils in phylogenetic analyses is necessary in order to construct a comprehensive "tree of death" and elucidate evolutionary history of taxa; however, such incorporation of fossils in phylogenetic reconstruction is dependent on the availability and interpretation of extensive morphological data. Here, the Zingiberales, whose familial relationships have been difficult to resolve with high support, are used as a case study to illustrate the importance of including fossil taxa in systematic studies. METHODS: Eight fossil taxa and 43 extant Zingiberales were coded for 39 morphological seed characters, and these data were concatenated with previously published molecular sequence data for analysis in the program MrBayes. KEY RESULTS: Ensete oregonense is confirmed to be part of Musaceae, and the other seven fossils group with Zingiberaceae. There is strong support for Spirematospermum friedrichii, Spirematospermum sp. 'Goth', S. wetzleri, and Striatornata sanantoniensis in crown Zingiberaceae while "Musa" cardiosperma, Spirematospermum chandlerae, and Tricostatocarpon silvapinedae are best considered stem Zingiberaceae. Inclusion of fossils explains how different topologies from morphological and molecular data sets is due to shared plesiomorphic characters shared by Musaceae, Zingiberaceae, and Costaceae, and most of the fossils. CONCLUSIONS: Inclusion of eight fossil taxa expands the Zingiberales tree and helps explain the difficulty in resolving relationships. Inclusion of fossils was possible in part due to a large morphological data set built using nondestructive microcomputed tomography data. Collaboration between paleo- and neobotanists and technology such as microcomputed tomography will help to build the tree of death and ultimately improve our understanding of the evolutionary history of monocots.

Monocot plastid phylogenomics, timeline, net rates of species diversification, the power of multi-gene analyses, and a functional model for the origin of monocots.

PREMISE OF THE STUDY: We present the first plastome phylogeny encompassing all 77 monocot families, estimate branch support, and infer monocot-wide divergence times and rates of species diversification. METHODS: We conducted maximum likelihood analyses of phylogeny and BAMM studies of diversification rates based on 77 plastid genes across 545 monocots and 22 outgroups. We quantified how branch support and ascertainment vary with gene number, branch length, and branch depth. KEY RESULTS: Phylogenomic analyses shift the placement of 16 families in relation to earlier studies based on four plastid genes, add seven families, date the divergence between monocots and eudicots+Ceratophyllum at 136 Mya, successfully place all mycoheterotrophic taxa examined, and support recognizing Taccaceae and Thismiaceae as separate families and Arecales and Dasypogonales as separate orders. Only 45% of interfamilial divergences occurred after the Cretaceous. Net species diversification underwent four large-scale accelerations in PACMAD-BOP Poaceae, Asparagales sister to Doryanthaceae, Orchidoideae-Epidendroideae, and Araceae sister to Lemnoideae, each associated with specific ecological/morphological shifts. Branch ascertainment and support across monocots increase with gene number and branch length, and decrease with relative branch depth. Analysis of entire plastomes in Zingiberales quantifies the importance of non-coding regions in identifying and supporting short, deep branches. CONCLUSIONS: We provide the first resolved, well-supported monocot phylogeny and timeline spanning all families, and quantify the significant contribution of plastome-scale data to resolving short, deep branches. We outline a new functional model for the evolution of monocots and their diagnostic morphological traits from submersed aquatic ancestors, supported by convergent evolution of many of these traits in aquatic Hydatellaceae (Nymphaeales).

The phylogeny of Heliconia (Heliconiaceae) and the evolution of floral presentation.

Heliconia (Heliconiaceae, order Zingiberales) is among the showiest plants of the Neotropical rainforest and represent a spectacular co-evolutionary radiation with hummingbirds. Despite the attractiveness and ecological importance of many Heliconia, the genus has been the subject of limited molecular phylogenetic studies. We sample seven markers from the plastid and nuclear genomes for 202 samples of Heliconia. This represents ca. 75% of accepted species and includes coverage of all taxonomic subgenera and sections. We date this phylogeny using fossils associated with other families in the Zingiberales; in particular we review and evaluate the Eocene fossil Ensete oregonense. We use this dated phylogenetic framework to evaluate the evolution of two components of flower orientation that are hypothesized to be important for modulating pollinator discrimination and pollen placement: resupination and erect versus pendant inflorescence habit. Our phylogenetic results suggest that the monophyletic Melanesian subgenus Heliconiopsis and a small clade of Ecuadorian species are together the sister group to the rest of Heliconia. Extant diversity of Heliconia originated in the Late Eocene (39Ma) with rapid diversification through the Early Miocene, making it the oldest known clade of hummingbird-pollinated plants. Most described subgenera and sections are not monophyletic, though closely related groups of species, often defined by shared geography, mirror earlier morphological cladistic analyses. Evaluation of changes in resupination and inflorescence habit suggests that these characters are more homoplasious than expected, and this largely explains the non-monophyly of previously circumscribed subgenera, which were based on these characters. We also find strong evidence for the correlated evolution of resupination and inflorescence habit. The correlated model suggests that the most recent common ancestor of all extant Heliconia had resupinate flowers and erect inflorescences. Finally, we note our nearly complete species sampling and dated phylogeny allow for an assessment of taxonomic history in terms of phylogenetic diversity. We find approximately half of the currently recognized species, corresponding to half of the phylogenetic diversity, have been described since 1975, highlighting the continued importance of basic taxonomic research and conservation initiatives to preserve both described and undiscovered species of Heliconia.

Speciation dynamics and biogeography of Neotropical spiral gingers (Costaceae).

Species can arise via the divisive effects of allopatry as well as due to ecological and/or reproductive character displacement within sympatric populations. Two separate lineages of Costaceae are native to the Neotropics; an early-diverging clade endemic to South America (consisting of ca. 16 species in the genera Monocostus, Dimerocostus and Chamaecostus); and the Neotropical Costus clade (ca. 50 species), a diverse assemblage of understory herbs comprising nearly half of total familial species richness. We use a robust dated molecular phylogeny containing most of currently known species to inform macroevolutionary reconstructions, enabling us to examine the context of speciation in Neotropical lineages. Analyses of speciation rate revealed a significant variation among clades, with a rate shift at the most recent common ancestor of the Neotropical Costus clade. There is an overall predominance of allopatric speciation in the South American clade, as most species display little range overlap. In contrast, sympatry is much higher within the Neotropical Costus clade, independent of node age. Our results show that speciation dynamics during the history of Costaceae is strongly heterogeneous, and we suggest that the Costus radiation in the Neotropics arose at varied geographic contexts.

Evolution of petaloidy in the zingiberales: An assessment of the relationship between ultrastructure and gene expression patterns.

BACKGROUND: The development of petal-like organs has occurred repetitively throughout angiosperm evolution. Despite homoplasy, it is possible that common underlying molecular mechanisms are repeatedly recruited to drive the development of petaloid organs. In Zingiberales, infertile, petal-like structures replace fertile stamens, resulting in petaloidy in androecial whorls. Assuming that androecial petaloidy is a shared derived characteristic, we expect to find common ultrastructure and molecular mechanisms underlying androecial petaloidy across Zingiberales. RESULTS: We show that petaloidy in Zingiberales is associated with tightly packed, protruding epidermal cells. Expression patterns for candidate genes involved in petal identity differ between the petaloid organs of Costaceae v. Cannaceae, despite similar macro- and microscopic organization. For all candidate gene families analyzed, our data suggest at least one Zingiberales-specific duplication event. CONCLUSIONS: Our data suggest that the patterns of B-class gene expression across the Zingiberales do not correlate with the occurrence of petaloidy, indicating that androecial petaloidy might have evolved independently of B-class gene expression in some lineages. It is possible that gene duplication may play a role in the diversity of petaloid structures found throughout the Zingiberales. It is likely that Zingiberales petaloidy may also result from the deployment of genes other than those involved in specification of petal identity. Developmental Dynamics 244:1121-1132, 2015. © 2015 Wiley Periodicals, Inc.

Adaptation in flower form: a comparative evodevo approach.

Evolutionary developmental biology (evodevo) attempts to explain how the process of organismal development evolves, utilizing a comparative approach to investigate changes in developmental pathways and processes that occur during the evolution of a given lineage. Evolutionary genetics uses a population approach to understand how organismal changes in form or function are linked to underlying genetics, focusing on changes in gene and genotype frequencies within populations and the fixation of genotypic variation into traits that define species or evoke speciation events. Microevolutionary processes, including mutation, genetic drift, natural selection and gene flow, can provide the foundation for macroevolutionary patterns observed as morphological evolution and adaptation. The temporal element linking microevolutionary processes to macroevolutionary patterns is development: an organism's genotype is converted to phenotype by ontogenetic processes. Because selection acts upon the phenotype, the connection between evolutionary genetics and developmental evolution becomes essential to understanding adaptive evolution in organismal form and function. Here, we discuss how developmental genetic studies focused on key developmental processes could be linked within a comparative framework to study the developmental genetics of adaptive evolution, providing examples from research on two key processes of plant evodevo - floral symmetry and organ fusion - and their role in the adaptation of floral form.

Seed morphology and anatomy and its utility in recognizing subfamilies and tribes of Zingiberaceae.

PREMISE OF THE STUDY: Recent phylogenetic analyses based on molecular data suggested that the monocot family Zingiberaceae be separated into four subfamilies and four tribes. Robust morphological characters to support these clades are lacking. Seeds were analyzed in a phylogenetic context to test independently the circumscription of clades and to better understand evolution of seed characters within Zingiberaceae. METHODS: Seventy-five species from three of the four subfamilies were analyzed using synchrotron based x-ray tomographic microscopy (SRXTM) and scored for 39 morphoanatomical characters. KEY RESULTS: Zingiberaceae seeds are some of the most structurally complex seeds in angiosperms. No single seed character was found to distinguish each subfamily, but combinations of characters were found to differentiate between the subfamilies. Recognition of the tribes based on seeds was possible for Globbeae, but not for Alpinieae, Riedelieae, or Zingibereae, due to considerable variation. CONCLUSIONS: SRXTM is an excellent, nondestructive tool to capture morphoanatomical variation of seeds and allows for the study of taxa with limited material available. Alpinioideae, Siphonochiloideae, Tamijioideae, and Zingiberoideae are well supported based on both molecular and morphological data, including multiple seed characters. Globbeae are well supported as a distinctive tribe within the Zingiberoideae, but no other tribe could be differentiated using seeds due to considerable homoplasy when compared with currently accepted relationships based on molecular data. Novel seed characters suggest tribal affinities for two currently unplaced Zingiberaceae taxa: Siliquamomum may be related to Riedelieae and Monolophus to Zingibereae, but further work is needed before formal revision of the family.

Molecular evolution and patterns of duplication in the SEP/AGL6-like lineage of the Zingiberales: a proposed mechanism for floral diversification.

The diversity of floral forms in the plant order Zingiberales has evolved through alterations in floral organ morphology. One striking alteration is the shift from fertile, filamentous stamens to sterile, laminar (petaloid) organs in the stamen whorls, attributed to specific pollination syndromes. Here, we examine the role of the SEPALLATA (SEP) genes, known to be important in regulatory networks underlying floral development and organ identity, in the evolution of development of the diverse floral organs phenotypes in the Zingiberales. Phylogenetic analyses show that the SEP-like genes have undergone several duplication events giving rise to multiple copies. Selection tests on the SEP-like genes indicate that the two copies of SEP3 have mostly evolved under balancing selection, probably due to strong functional restrictions as a result of their critical role in floral organ specification. In contrast, the two LOFSEP copies have undergone differential positive selection, indicating neofunctionalization. Reverse transcriptase-polymerase chain reaction, gene expression from RNA-seq data, and in situ hybridization analyses show that the recovered genes have differential expression patterns across the various whorls and organ types found in the Zingiberales. Our data also suggest that AGL6, sister to the SEP-like genes, may play an important role in stamen morphology in the Zingiberales. Thus, the SEP-like genes are likely to be involved in some of the unique morphogenetic patterns of floral organ development found among this diverse order of tropical monocots. This work contributes to a growing body of knowledge focused on understanding the role of gene duplications and the evolution of entire gene networks in the evolution of flower development.

Origin and diversification of the Milla Clade (Brodiaeoideae, Asparagaceae): a Neotropical group of six geophytic genera.

The Milla clade currently comprises six genera of geophytic plants distributed from Arizona to Guatemala. Three genera (Behria, Jaimehintonia and Petronymphe) are monotypic while the remaining genera (Bessera, Dandya and Milla) contain from two to ten (Milla) species. Parsimony, Maximum Likelihood and Bayesian Inference analyses were conducted with plastid and nuclear DNA sequences from a total of 181 plants belonging to 15 species in all six genera. Molecular dating was performed under a relaxed clock model. We examined the phylogenetic relationships of the genera and species, estimated origin-divergence times for the clade and genera and determined the ancestral distribution area of the clade by optimizing ancestral areas given current biogeographic distributions. The phylogenetic results suggest that final decisions on limits of the six genera in the Milla clade will have to be established until further taxonomic work is completed for Milla, in particular for the group of populations included under the name M. biflora. The later genus is rendered polyphyletic by other genera of the family. The origin of the Milla clade is estimated at 15.8Ma. Ancestral area of the clade most likely was located in the California Floristic Province and dispersal occurred most likely to the Chihuahuan-Coahuila Plateaus and the Trans-Mexican Volcanic Belt and from there to Baja California and the Sierra Madre del Sur. Two hypotheses that need further testing are proposed to explain complex relationships of genera and polyphyly of Milla, one in relation to fragmentation of populations and pollinator shifts and another suggesting that populations remained in refugia in the Trans-Mexican Volcanic Belt.