Phylogeny of extant and fossil Juglandaceae inferred from the integration of molecular and morphological data sets

It is widely acknowledged that integrating fossils into data sets of extant taxa is imperative for proper placement of fossils, resolution of relationships, and a better understanding of character evolution. The importance of this process has been further magnified because of the crucial role of fossils in dating divergence times. Outstanding issues remain, including appropriate methods to place fossils in phylogenetic trees, the importance of molecules versus morphology in these analyses, as well as the impact of potentially large amounts of missing data for fossil taxa. In this study we used the angiosperm clade Juglandaceae as a model for investigating methods of integrating fossils into a phylogenetic framework of extant taxa. The clade has a rich fossil record relative to low extant diversity, as well as a robust molecular phylogeny and morphological database for extant taxa. After combining fossil organ genera into composite and terminal taxa, our objectives were to (1) compare multiple methods for the integration of the fossils and extant taxa (including total evidence, molecular scaffolds, and molecular matrix representation with parsimony [MRP]); (2) explore the impact of missing data (incomplete taxa and characters) and the evidence for placing fossils on the topology; (3) simulate the phylogenetic effect of missing data by creating "artificial fossils"; and (4) place fossils and compare the impact of single and multiple fossil constraints in estimating the age of clades. Despite large and variable amounts of missing data, each of the methods provided reasonable placement of both fossils and simulated "artificial fossils" in the phylogeny previously inferred only from extant taxa. Our results clearly show that the amount of missing data in any given taxon is not by itself an operational guideline for excluding fossils from analysis. Three fossil taxa (Cruciptera simsonii, Paleoplatycarya wingii, and Platycarya americana) were placed within crown clades containing living taxa for which relationships previously had been suggested based on morphology, whereas Polyptera manningii, a mosaic taxon with equivocal affinities, was placed firmly as sister to two modern crown clades. The position of Paleooreomunnea stoneana was ambiguous with total evidence but conclusive with DNA scaffolds and MRP. There was less disturbance of relationships among extant taxa using a total evidence approach, and the DNA scaffold approach did not provide improved resolution or internal support for clades compared to total evidence, whereas weighted MRP retained comparable levels of support but lost crown clade resolution. Multiple internal minimum age constraints generally provided reasonable age estimates, but the use of single constraints provided by extinct genera tended to underestimate clade ages.     

Molecular phylogeny, biogeography, and systematics of Dicerandra (Lamiaceae), a genus endemic to the southeastern United States

Dicerandra, an endemic mint of the southeastern United States, comprises nine species, all of which are threatened or endangered and restricted to sandhill vegetation and a mosaic of scrub habitats. Molecular phylogenetic analyses of Dicerandra based on data from the nuclear and plastid genomes for all 13 taxa of the genus, identified two strongly supported clades, corresponding to the four annual and to the five perennial species of Dicerandra. However, the nuclear and plastid trees were incongruent in their placement of two perennial taxa, D. cornutissima and D. immaculata var. savannarum, perhaps due to ancient hybridization or to lineage sorting. Based on these analyses, the widespread D. linearifolia is not monophyletic, with populations of D. linearifolia var. linearifolia falling into either western or eastern clades. The western clade, comprising populations of D. linearifolia var. linearifolia and var. robustior, occurs in an area drained by rivers flowing toward the Gulf of Mexico, whereas the eastern clade, comprising populations of D. linearifolia var. linearifolia, D. densiflora, D. odoratissima, and D. radfordiana (i.e., all the annual species), occupies a region drained by rivers flowing to the Atlantic Ocean. Although this pattern of genetic differentiation between populations from these two river drainages has been documented in several animal species, it has not previously been reported for plants. A revised subgeneric classification is presented to reflect the annual and perennial clades.     

A physical map for the Amborella trichopoda genome sheds light on the evolution of angiosperm genome structure

Background

Recent phylogenetic analyses have identified Amborella trichopoda, an understory tree species endemic to the forests of New Caledonia, as sister to a clade including all other known flowering plant species. The Amborella genome is a unique reference for understanding the evolution of angiosperm genomes because it can serve as an outgroup to root comparative analyses. A physical map, BAC end sequences and sample shotgun sequences provide a first view of the 870 Mbp Amborella genome.

Results

Analysis of Amborella BAC ends sequenced from each contig suggests that the density of long terminal repeat retrotransposons is negatively correlated with that of protein coding genes. Syntenic, presumably ancestral, gene blocks were identified in comparisons of the Amborella BAC contigs and the sequenced Arabidopsis thaliana, Populus trichocarpa, Vitis vinifera and Oryza sativa genomes. Parsimony mapping of the loss of synteny corroborates previous analyses suggesting that the rate of structural change has been more rapid on lineages leading to Arabidopsis and Oryza compared with lineages leading to Populus and Vitis. The gamma paleohexiploidy event identified in the Arabidopsis, Populus and Vitis genomes is shown to have occurred after the divergence of all other known angiosperms from the lineage leading to Amborella.

Conclusions

When placed in the context of a physical map, BAC end sequences representing just 5.4% of the Amborella genome have facilitated reconstruction of gene blocks that existed in the last common ancestor of all flowering plants. The Amborella genome is an invaluable reference for inferences concerning the ancestral angiosperm and subsequent genome evolution.     

AUTOPOLYPLOIDY IN TOLMIEA MENZIESII (SAXIFRAGACEAE)

Tolmiea menziesii comprises diploid (2n = 14) and tetraploid (2n = 28) cytotypes that differ in geographic distribution. Chromosome counts now available indicate that the diploid and tetraploid cytotypes occupy the southern and northern portions, respectively, of the range of Tolmiea. Available data strongly suggest that Tolmiea represents an example of autopolyploidy. The genus is monotypic and very distinct in both floral and vegetative morphology among genera of tribe Saxifrageae. Infraspecific taxa have not been recognized in T. menziesii, and the two cytotypes appear to be indistinguishable morphologically. Karyotypic, flavonoid chemical, and preliminary allozymic data are all in agreement with the contention that the tetraploid cytotype is of autopolyploidal origin.     

ESTIMATES OF INTRAGAMETOPHYTIC SELFING AND INTERPOPULATIONAL GENE FLOW IN HOMOSPOROUS FERNS

Relatively little information is available on mating systems and interpopulational gene flow in species of homosporous pteridophytes. Because of the proximity of antheridia and archegonia on the same thallus, it has long been maintained that intragametophytic selling is the predominant mode of reproduction in natural populations of homosporous ferns and other homosporous plants. Furthermore, quantitative estimates of interpopulational gene flow via spore dispersal are lacking. In this paper, we examine five species of homosporous ferns (Botrychium virginianum, Polystichum munitum, P. imbricans, Blechnum spicant, and Dryopteris expansa) and present estimates of 1) rates of intragametophytic selling, 2) levels of interpopulational gene flow, and 3) interpopulational genetic differentiation (F_ST). Our data demonstrate that mating systems vary among species of ferns, just as they do among species of seed plants. The data also suggest that levels of interpopulational gene flow are generally high. The F_ST values indicate little genetic divergence among populations for all species except Dryopteris expansa, which exhibits significant levels of interpopulational genetic differentiation. Patterns of genetic diversity in the five species examined are related to the mating system and rate of interpopulational gene flow in each species. The F_ST values for all species except Botrychium virginianum are in close agreement with those predicted for an island model of population structure.     

GENETIC VARIATION AND POPULATION STRUCTURE IN THE FERN BLECHNUM SPICANT (BLECHNACEAE) FROM WESTERN NORTH AMERICA

Population genetic structure in the homosporous fern Blechnum spicant was analyzed in six populations from western North America. Each population was divided into approximately 10 m by 10 m subpopulations, and genetic variation within and among subpopulations was compared using enzyme electrophoresis and F statistics. These analyses indicated that there was no evidence of genetic structure in four of the six populations examined. However, significant genetic heterogeneity among subpopulations was observed for the other two populations. The genetic structure of these populations may be attributable, in part, to family structure resulting from high rates of intragametophytic selling and/or spatial patchiness in the distribution of individuals due to limited habitat availability in these areas. Outcrossing populations of B. spicant generally lack genetic structure, whereas the most highly inbreeding population maintains significant genetic structure. The information obtained in this investigation of population genetic structure in Blechnum spicant is consistent with data for angiosperms and gymnosperms. It appears that the outcrossing mating system and effective mechanism of spore dispersal in B. spicant may account for the general lack of genetic structure within populations of this species.     

Expression of floral MADS-box genes in basal angiosperms: implications for the evolution of floral regulators

The ABC model of floral organ identity is based on studies of Arabidopsis and Antirrhinum, both of which are highly derived eudicots. Most of the genes required for the ABC functions in Arabidopsis and Antirrhinum are members of the MADS-box gene family, and their orthologs are present in all major angiosperm lineages. Although the eudicots comprise 75% of all angiosperms, most of the diversity in arrangement and number of floral parts is actually found among basal angiosperm lineages, for which little is known about the genes that control floral development. To investigate the conservation and divergence of expression patterns of floral MADS-box genes in basal angiosperms relative to eudicot model systems, we isolated several floral MADS-box genes and examined their expression patterns in representative species, including Amborella (Amborellaceae), Nuphar (Nymphaeaceae) and Illicium (Austrobaileyales), the successive sister groups to all other extant angiosperms, plus Magnolia and Asimina, members of the large magnoliid clade. Our results from multiple methods (relative-quantitative RT-PCR, real-time PCR and RNA in situ hybridization) revealed that expression patterns of floral MADS-box genes in basal angiosperms are broader than those of their counterparts in eudicots and monocots. In particular, (i) AP1 homologs are generally expressed in all floral organs and leaves, (ii) AP3/PI homologs are generally expressed in all floral organs and (iii) AG homologs are expressed in stamens and carpels of most basal angiosperms, in agreement with the expectations of the ABC model; however, an AG homolog is also expressed in the tepals of Illicium. The broader range of strong expression of AP3/PI homologs is inferred to be the ancestral pattern for all angiosperms and is also consistent with the gradual morphological intergradations often observed between adjacent floral organs in basal angiosperms.