Oriane Hidalgo

The cardueae (Compositae) revisited : Insights from its, trnL-trnF, and matK nuclear and chloroplast DNA analysis

Abstract The new outline of relationships in basal branches of the family Compositae Giseke confirms that the sister group to the tribe Cardueae Cass. are not Mutisieae Cass., but rather a group of African genera now classified as the tribe Tarchonantheae Kostel. This change implies that the monophyly of the Cardueae must be reassessed on a molecular basis. Moreover, new collections in recent years allow us to extend our sampling to 70 of the 74 genera of the tribe. We performed a new molecular study of the tribe using one nuclear region (ITS) and two chloroplastic markers (trnL-trnF and matK) in addition to a more appropriate outgroup. Our results confirm that the Cardueae is a natural group but indicate some changes in subtribal delineation: the subtribe Cardopatiinae Less. is recognized and some genera are moved to other subtribes (Myopordon Boiss., Nikitinia Iljin, Syreitschikovia Pavlov, and the Xeranthemum L. group). A recapitulation of a number of interesting questions that remain unresolved in the classification of some large genera is presented.

Genome size in Echinops L. and related genera (Asteraceae, Cardueae): karyological, ecological and phylogenetic implications.

Abstract Genome size was assessed by flow cytometry in 33 species belonging to seven genera of the tribe Cardueae (Asteraceae), which can be grouped in three taxonomic complexes. 2C nuclear DNA content ranged from 1.49 to 16.98 pg, which is more than elevenfold variation. Genome size correlated well with some karyological traits. Nuclear DNA amount variations also have systematic and evolutionary implications and/or are linked to adaptations to ecological conditions.

Extensive ribosomal DNA (18S-5.8S-26S and 5S) colocalization in the North American endemic sagebrushes (subgenus Tridentatae, Artemisia, Asteraceae) revealed by FISH

Chromomycin A3 banding and fluorescent in situ hybridization (FISH) have been performed for six Artemisia species with special emphasis on subgenus Tridentatae. Morphometrical data on karyotype characters were calculated and idiograms with the position of GC-rich regions and 18S-5.8S-26S and 5S sites of ribosomal DNA were constructed. These sites were all colocalized. To our knowledge, this is the first time in the large family Asteraceae, indeed in angiosperms in general, that colocalization of the two rDNA regions studied is found at every single marked locus. In addition, transcriptionally active nucleolar organizer regions were detected after silver nitrate staining. Tridentatae is a cytogenetically homogeneous subgenus, which suggests that evolution of these species has not been coupled with important karyotypic reorganization. However, a few species are taxonomically difficult and show substantial differences. A loss of rDNA loci has been detected in a tetraploid taxon with respect to the diploids studied. These data provide clarifying insight into interspecific relationships between the studied taxa and overall evolutionary and systematic relationships of the Tridentatae.

Recent updates and developments to plant genome size databases

Two plant genome size databases have been recently updated and/or extended: the Plant DNA C-values database (http://data.kew.org/cvalues), and GSAD, the Genome Size in Asteraceae database (http://www.asteraceaegenomesize.com). While the first provides information on nuclear DNA contents across land plants and some algal groups, the second is focused on one of the largest and most economically important angiosperm families, Asteraceae. Genome size data have numerous applications: they can be used in comparative studies on genome evolution, or as a tool to appraise the cost of whole-genome sequencing programs. The growing interest in genome size and increasing rate of data accumulation has necessitated the continued update of these databases. Currently, the Plant DNA C-values database (Release 6.0, Dec. 2012) contains data for 8510 species, while GSAD has 1219 species (Release 2.0, June 2013), representing increases of 17 and 51%, respectively, in the number of species with genome size data, compared with previous releases. Here we provide overviews of the most recent releases of each database, and outline new features of GSAD. The latter include (i) a tool to visually compare genome size data between species, (ii) the option to export data and (iii) a webpage containing information about flow cytometry protocols.

Biology, Genome Evolution, Biotechnological Issues and Research Including Applied Perspectives in Artemisia (Asteraceae)

Abstract Artemisia is one of the largest genera of the family Asteraceae or Compositae, itself the biggest flowering plant family. It comprises around 600 taxa at specific and subspecific levels, present in all continents but Antarctica, mostly distributed in the Northern Hemisphere, with no more than 25 taxa in the Southern Hemisphere. The genus displays a huge ecological plasticity, with species occurring from sea level to high mountains and from arid zones to wetlands. Some species are cosmopolitan, including landscape-dominating plants over large areas, and others are endemics with a quite restricted distribution area. Many species of the genus have economic uses at both folk and industrial levels, and some of them are widely cultivated and submitted to breeding programmes as crops. In this review, we will set out the state of art of Artemisia systematics and phylogeny, as well as all the biological aspects of the genus, with particular attention paid to those of genome organization, and of applied questions related to its useful taxa. In the first part of this chapter, we will review all the systematic points in the genus and in some closely related genera that constitute, with the core genus Artemisia, a pool with controversial structuring. Besides, the infrageneric classification will be addressed. All these questions will be treated in the light of recent molecular phylogenetic studies, which have had an important impact on its systematics and taxonomy. A second part will be devoted to genome organization and evolution in Artemisia, with special attention to cytogenetic data, including genome size, and genetic variability. These points are relevant for understanding the evolutionary pathways in the genus and for applied purposes. The third and fourth parts of the chapter will review, respectively, the uses of Artemisia species in different domains and the biotechnological issues linked to their productivity. Finally, the perspectives of the knowledge and applied aspects of the genus will be addressed.

Genome size variation and evolution in the family Asteraceae

The nuclear DNA content data available in “A genome size database in the Asteraceae” (GSAD: www.asteraceaegenomesize.com) have been analyzed, together with other parameters (i.e. ecological, karyological, cytogenetic), in order to establish hypotheses on the systematic, phylogenetic and evolutionary aspects of genome size in one of the largest angiosperm families. The novelty of this work is a comprehensive analysis of the whole family with the following aims: (1) to update the knowledge of genome size values in the Asteraceae; (2) to infer evolutionary trends of genome size, compared with other plant groups; and (3) to detect gaps in this field in the family and outline further research priorities. The analysis of this dataset shows that most Asteraceae genomes (57.23%) range from very small (1C ⩽ 1.4 pg) to small (1C ⩽ 3.5 pg). Gains and losses of DNA occur throughout the phylogeny of the family but although ancestral values for the basal nodes remain mostly equivocal, often small and very small ancestral genome sizes are reconstructed. Most genome size data (96.74%) are concentrated in five tribes, which broadly reflect their species richness. The relationships between genome size and other cytogenetic and ecological features have been analyzed and discussed, highlighting several general patterns. Further studies are needed to fill the gaps in genome size knowledge in the Asteraceae and more detailed research in some groups could provide information about mechanisms regulating genome expansions and contractions.

Genome evolution of ferns: evidence for relative stasis of genome size across the fern phylogeny

The genome evolution of ferns has been considered to be relatively static compared with angiosperms. In this study, we analyse genome size data and chromosome numbers in a phylogenetic framework to explore three hypotheses: the correlation of genome size and chromosome number, the origin of modern ferns from ancestors with high chromosome numbers, and the occurrence of several whole-genome duplications during the evolution of ferns. To achieve this, we generated new genome size data, increasing the percentage of fern species with genome sizes estimated to 2.8% of extant diversity, and ensuring a comprehensive phylogenetic coverage including at least three species from each fern order. Genome size was correlated with chromosome number across all ferns despite some substantial variation in both traits. We observed a trend towards conservation of the amount of DNA per chromosome, although Osmundaceae and Psilotaceae have substantially larger chromosomes. Reconstruction of the ancestral genome traits suggested that the earliest ferns were already characterized by possessing high chromosome numbers and that the earliest divergences in ferns were correlated with substantial karyological changes. Evidence for repeated whole-genome duplications was found across the phylogeny. Fern genomes tend to evolve slowly, albeit genome rearrangements occur in some clades.

GSAD: a genome size in the Asteraceae database.

THE Asteraceae are one of the largest families of angiosperms, comprising 24,000 to 30,000 species in over 1,600 to 2,000 genera (1 and references therein). It has a worldwide distribution, with the exception of Antarctica and includes many economically important species which are used, for example, as foods, medicines, and ornamentals. Asteraceae species are the target of many evolutionary studies and more recently they have also become the focus of new genome sequencing programs. New model species for evolutionary-developmental (evo-devo) research have been selected within the Asteraceae such as Gerbera, Helianthus, and Senecio, whereas Tragopogon is the focus of intensive studies on polyploidization mechanisms (2). The first evo-devo studies in the Asteraceae have been very promising despite complications arising from the genetic and epigenetic changes associated with polyploidy which is very frequent in the family. The term ‘‘C-value’’ was coined by Swift (3) to define the gametic nuclear DNA content (genome size) expressed in picograms. Nowadays, genome size research covers a large and diverse range of biological fields and extends across all plant groups. For example, studies have been carried out on genome size nomenclature (4), to improve methodological aspects (5) and to find possible explanations of how and why genome size changes occur in plants (6). Data on nuclear DNA amounts are interesting not only per se but are also of practical use. For instance, the success of techniques such as AFLPs and nuclear microsatellites are influenced by genome size, while the choice of a species for possible genome sequencing or evo-devo project is also determined, in part, by genome size. Interest in genome size has increased over the years and this has led to the development of several related databases (e.g., for plants 7–9). Following on from our own research studies on genome size in the Asteraceae family and given that the family is one of the most intensely studied from many aspects, we have developed a genome size database focused specifically on the Asteraceae (which we have named the ‘‘Genome size in the Asteraceae database’’, GSAD). It is hoped that this will become a significant tool for comparative research and for future genome size studies.

Assessing duplication and loss of APETALA1/FRUITFULL homologs in Ranunculales

Gene duplication and loss provide raw material for evolutionary change within organismal lineages as functional diversification of gene copies provide a mechanism for phenotypic variation. Here we focus on the APETALA1/FRUITFULL MADS-box gene lineage evolution. AP1/FUL genes are angiosperm-specific and have undergone several duplications. By far the most significant one is the core-eudicot duplication resulting in the euAP1 and euFUL clades. Functional characterization of several euAP1 and euFUL genes has shown that both function in proper floral meristem identity, and axillary meristem repression. Independently, euAP1 genes function in floral meristem and sepal identity, whereas euFUL genes control phase transition, cauline leaf growth, compound leaf morphogenesis and fruit development. Significant functional variation has been detected in the function of pre-duplication basal-eudicot FUL-like genes, but the underlying mechanisms for change have not been identified. FUL-like genes in the Papaveraceae encode all functions reported for euAP1 and euFUL genes, whereas FUL-like genes in Aquilegia (Ranunculaceae) function in inflorescence development and leaf complexity, but not in flower or fruit development. Here we isolated FUL-like genes across the Ranunculales and used phylogenetic approaches to analyze their evolutionary history. We identified an early duplication resulting in the RanFL1 and RanFL2 clades. RanFL1 genes were present in all the families sampled and are mostly under strong negative selection in the MADS, I and K domains. RanFL2 genes were only identified from Eupteleaceae, Papaveraceae s.l., Menispermaceae and Ranunculaceae and show relaxed purifying selection at the I and K domains. We discuss how asymmetric sequence diversification, new motifs, differences in codon substitutions and likely protein-protein interactions resulting from this Ranunculiid-specific duplication can help explain the functional differences among basal-eudicot FUL-like genes.

Virus-induced gene silencing (VIGS) in Cysticapnos vesicaria, a zygomorphic-flowered Papaveraceae (Ranunculales, basal eudicots)

BACKGROUND AND AIMS Studies of evolutionary diversification in the basal eudicot family Papaveraceae, such as the transition from actinomorphy to zygomorphy, are hampered by the lack of comparative functional studies. So far, gene silencing methods are only available in the actinomorphic species Eschscholzia californica and Papaver somniferum. This study addresses the amenability of Cysticapnos vesicaria, a derived fumitory with zygomorphic flowers, to virus-induced gene silencing (VIGS), and describes vegetative and reproductive traits in this species. METHODS VIGS-mediated downregulation of the C. vesicaria PHYTOENE DESATURASE gene (CvPDS) and of the FLORICAULA gene CvFLO was carried out using Agrobacterium tumefaciens transfer of Tobacco rattle virus (TRV)-based vectors. Wild-type and vector-treated plants were characterized using reverse transcription-PCR (RT-PCR), in situ hybridization, and macroscopic and scanning electron microscopic imaging. KEY RESULTS Cysticapnos vesicaria germinates rapidly, can be grown at high density, has a short life cycle and is self-compatible. Inoculation of C. vesicaria with a CvPDS-VIGS vector resulted in strong photobleaching of green parts and reduction of endogenous CvPDS transcript levels. Gene silencing persisted during inflorescence development until fruit set. Inoculation of plants with CvFLO-VIGS affected floral phyllotaxis, symmetry and floral organ identities. CONCLUSIONS The high penetrance, severity and stability of pTRV-mediated silencing, including the induction of meristem-related phenotypes, make C. vesicaria a very promising new focus species for evolutionary-developmental (evo-devo) studies in the Papaveraceae. This now enables comparative studies of flower symmetry, inflorescence determinacy and other traits that diversified in the Papaveraceae.