Origin and genome evolution of polyploids in the genus Melampodium

Polyploidy is a prominent feature of plants contributing to their diversification and speciation. Although polyploidy has impacted nearly all plants, many questions are still open regarding the spatial and temporal conditions facilitating polyploidization, the evolutionary success of polyploids, mechanisms and trends shaping polyploid genome evolution. A suitable system for addressing aspects of temporal and spatial origins and genomic evolution of polyploids is provided by series Sericea of the genus Melampodium (Asteraceae). This species complex, centered in Mexico, includes five exclusively allopolyploid taxa (tetraploids and hexaploids). Hypotheses on their origins have recently been inferred from molecular, chromosomal and morphological data. Based on genome size and rDNA loci evolution, initial hypotheses on their genomic evolution have also been proposed. Recent methodological advances, especially low-cost next generation sequencing (NGS) applied both to coding and repetitive genome fractions and implemented by molecular, bioinformatic, and cytogenetic techniques offer new tools to further advance our understanding of the evolutionary trajectories of this polyploid complex in particular and of polyploids in general. In the proposed project a combination of methods and techniques will be used to analyze in detail the origin and evolution of allotetraploids and allohexaploids of Melampodium series Sericea. Phylogenetic and phylogeographic approaches will be employed to elucidate the exact evolutionary origin and history of this complex, and will be complemented with in-depth analyzes of genome evolution following polyploidization. The comparative investigation of this polyploid complex will address significant evolutionary questions concerning the impact of polyploid origin and evolutionary history on type and frequencies of genomic changes contributing to speciation. Specifically, the proposal addresses the following questions: How do the origins of allopolyploids relate to the phylogeographic history of their lower-ploid ancestors? Have the allopolyploids had multiple origins and were these allopolyploidizations temporally and/or spatially clustered? Does the mode of polyploid formation influence the establishment of wide distribution areas in allopolyploids? Do allopolyploids show niche shifts with respect to their ancestral diploid taxa? Do allopolyploids respond differently to climatic oscillations than their lower-ploid ancestors? The outcome of the phylogenetic analyses will enable better interpretation of the genomic changes in diploids and polyploids: Which repeats contribute to significant changes of genome size in diploid species and what is their dynamics over the evolutionary time? Do evolutionary genome dynamics in polyploids differ from those in their lower-ploid ancestors, and are the changes reflected in chromosomal restructuring? Is there parental genome bias/dominance in polyploids? Are there differences in genome dynamics between species of different ploidy levels (tetraploid versus hexaploid) and/or of different ages? Does polyploid evolution repeat itself?

The genus Melampodium (Asteraceae) comprises 40 species distributed mostly throughout Mexico. The presence of several base chromosome numbers (x = 9, 10, 11, 12, 14) and high incidence of polyploidy (whole genome duplications) makes it an ideal model for studying genome evolution following polyploidization. Half of the species in the genus (20 spp.) belong to section Melampodium (x = 10), although this base number was not inferred as most likely ancestral base chromosome number. Despite all 14 diploid species in this section having the same chromosome number, they differ nearly 4.5-fold in the size of the genome. Reconstruction of ancestral diploid genome size in the section revealed two independent instances of genome size increase during the evolution of this group, both caused by increase of copy numbers of the same type of retrotransposons (Ty1-copia superfamily), and one case of genome downsizing resulting from proportional reduction across most repeat types. Overall profiles of repetitive DNA fraction in all species of sect. Melampodium have been obtained from analyses of several millions of DNA fragments sequenced using Next Generation Sequencing Illumina platform and clustered using bioinformatics pipeline RepeatExplorer. Repeat-based phylogeny reconstruction largely reflected the current phylogenetic hypotheses on species relationships. A model-based analysis revealed that various types/lineages of the repeats evolved at different rates and time scales in the analysed genomes. Nearly 40% of all Melampodium species are polyploids, with series Sericea of section Melampodium composed exclusively of five allopolyploids. Allopolyploid origin of these species has been confirmed using novel formamide-free genomic in situ hybridization (GISH), a method that allows more effective mapping of DNAs of all parental genomes in polyploids. The polyploids were shown to originate recurrently and to be relatively young (max. 1.4 My). The age of allopolyploids was established using a novel approach implemented in the program BEAST that can be applied for dating or the origin of any allopolyploid species. Comparative analyses of repeat landscapes in the genomes of parental species and allopolyploids revealed relatively low levels of post-polyploidization change accompanying diploidization. The older allotetraploids experienced more changes than derivative allohexaploids. The degree of similarity of parental species and age of allopolyploids were shown to influence the extent of genomic changes. Despite low levels of large-scale repeat changes and limited interaction between parental subgenomes, genomes of allopolyploids underwent some chromosomal restructuring. Two newly identified parent-specific tandem repeats have spread to the other parental subgenome in allopolyploids suggesting restructuring despite genome size additivity. Phylogeographic and phylogenetic analyses of the allopolyploids and their parental species have been performed using low-copy nuclear and plastid DNA markers, and high-throughput RAD sequencing data and indicated recurrent formation of allopolyploids from widespread parental taxa.