Tae-Soo Jang

Evolutionary Consequences, Constraints and Potential of Polyploidy in Plants

Polyploidy, the possession of more than 2 complete genomes, is a major force in plant evolution known to affect the genetic and genomic constitution and the phenotype of an organism, which will have consequences for its ecology and geography as well as for lineage diversification and speciation. In this review, we discuss phylogenetic patterns in the incidence of polyploidy including possible underlying causes, the role of polyploidy for diversification, the effects of polyploidy on geographical and ecological patterns, and putative underlying mechanisms as well as chromosome evolution and evolution of repetitive DNA following polyploidization. Spurred by technological advances, a lot has been learned about these aspects both in model and increasingly also in nonmodel species. Despite this enormous progress, long-standing questions about polyploidy still cannot be unambiguously answered, due to frequently idiosyncratic outcomes and insufficient integration of different organizational levels (from genes to ecology), but likely this will change in the near future. See also the sister article focusing on animals by Choleva and Janko in this themed issue.

Chromosomal diversification and karyotype evolution of diploids in the cytologically diverse genus Prospero(Hyacinthaceae)

BackgroundProspero (Hyacinthaceae) provides a unique system to assess the impact of genome rearrangements on plant diversification and evolution. The genus exhibits remarkable chromosomal variation but very little morphological differentiation. Basic numbers of x = 4, 5, 6 and 7, extensive polyploidy, and numerous polymorphic chromosome variants were described, but only three species are commonly recognized: P. obtusifolium, P. hanburyi, and P. autumnale s.l., the latter comprising four diploid cytotypes. The relationship between evolutionary patterns and chromosomal variation in diploids, the basic modules of the extensive cytological diversity, is presented.ResultsEvolutionary inferences were derived from fluorescence in situ hybridization (FISH) with 5S and 35S rDNA, genome size estimations, and phylogenetic analyses of internal transcribed spacer (ITS) of 35S rDNA of 49 diploids in the three species and all cytotypes of P. autumnale s.l. All species and cytotypes possess a single 35S rDNA locus, interstitial except in P. hanburyi where it is sub-terminal, and one or two 5S rDNA loci (occasionally a third in P. obtusifolium) at fixed locations. The localization of the two rDNA types is unique for each species and cytotype. Phylogenetic data in the P. autumnale complex enable tracing of the evolution of rDNA loci, genome size, and direction of chromosomal fusions: mixed descending dysploidy of x = 7 to x = 6 and independently to x = 5, rather than successive descending dysploidy, is proposed.ConclusionsAll diploid cytotypes are recovered as well-defined evolutionary lineages. The cytogenetic and phylogenetic approaches have provided excellent phylogenetic markers to infer the direction of chromosomal change in Prospero. Evolution in Prospero, especially in the P. autumnale complex, has been driven by differentiation of an ancestral karyotype largely unaccompanied by morphological change. These new results provide a framework for detailed analyses of various types of chromosomal rearrangements and karyotypic variation in polyploids.

Differential amplification of satellite PaB6 in chromosomally hypervariable Prospero autumnale complex (Hyacinthaceae)

Background and Aims Chromosomal evolution, including numerical and structural changes, is a major force in plant diversification and speciation. This study addresses genomic changes associated with the extensive chromosomal variation of the Mediterranean Prospero autumnale complex (Hyacinthaceae), which includes four diploid cytotypes each with a unique combination of chromosome number (x = 5, 6, 7), rDNA loci and genome size. Methods A new satellite repeat PaB6 has previously been identified, and monomers were reconstructed from next-generation sequencing (NGS) data of P. autumnale cytotype B6B6 (2n = 12). Monomers of all other Prospero cytotypes and species were sequenced to check for lineage-specific mutations. Copy number, restriction patterns and methylation levels of PaB6 were analysed using Southern blotting. PaB6 was localized on chromosomes using fluorescence in situ hybridization (FISH). Key Results The monomer of PaB6 is 249 bp long, contains several intact and truncated vertebrate-type telomeric repeats and is highly methylated. PaB6 is exceptional because of its high copy number and unprecedented variation among diploid cytotypes, ranging from 104 to 106 copies per 1C. PaB6 is always located in pericentromeric regions of several to all chromosomes. Additionally, two lineages of cytotype B7B7 (x = 7), possessing either a single or duplicated 5S rDNA locus, differ in PaB6 copy number; the ancestral condition of a single locus is associated with higher PaB6 copy numbers. Conclusions Although present in all Prospero species, PaB6 has undergone differential amplification only in chromosomally variable P. autumnale, particularly in cytotypes B6B6 and B5B5. These arose via independent chromosomal fusions from x = 7 to x = 6 and 5, respectively, accompanied by genome size increases. The copy numbers of satellite DNA PaB6 are among the highest in angiosperms, and changes of PaB6 are exceptionally dynamic in this group of closely related cytotypes of a single species. The evolution of the PaB6 copy numbers is discussed, and it is suggested that PaB6 represents a recent and highly dynamic system originating from a small pool of ancestral repeats.

Triparental origin of triploid onion, Allium × cornutum (Clementi ex Visiani, 1842), as evidenced by molecular, phylogenetic and cytogenetic analyses

BackgroundReconstruction of the parental origins of cultivated plants from wild relatives, especially after long periods of domestication, is not a trivial task. However, recent advances in molecular phylogenetics, among other approaches, have proved to be very informative in analyses of the origin and evolution of polyploid genomes. An established minor garden crop, triploid onion Allium × cornutum (Clementi ex Visiani, 1842) (2n = 3x = 24), is widespread in southeastern Asia and Europe. Our previous cytogenetic analyses confirmed its highly heterozygous karyotype and indicated its possible complex triparental genome origin. Allium cepa L. and Allium roylei Stearn were suggested as two putative parental species of A. × cornutum, whereas the third parental species remained hitherto unknown.ResultsHere we report the phylogenetic analyses of the internal transcribed spacers ITS1-5.8S-ITS2 of 35S rDNA and the non-transcribed spacer (NTS) region of 5S rDNA of A. × cornutum and its relatives of the section Cepa. Both ITS and NTS sequence data revealed intra-individual variation in triploid onion, and these data clustered into the three main clades, each with high sequence homology to one of three other species of section Cepa: A. cepa, A. roylei, and unexpectedly, the wild Asian species Allium pskemense B. Fedtsh. Allium pskemense is therefore inferred to be the third, so far unknown, putative parental species of triploid onion Allium × cornutum. The 35S and 5S rRNA genes were found to be localised on somatic chromosomes of A. × cornutum and its putative parental species by double fluorescent in situ hybridisation (FISH). The localisation of 35S and 5S rDNA in A. × cornutum chromosomes corresponded to their respective positions in the three putative parental species, A. cepa, A. pskemense, and A. roylei. GISH (genomic in situ hybridisation) using DNA of the three putative parental diploids corroborated the results of the phylogenetic study.ConclusionsThe combined molecular, phylogenetic and cytogenetic data obtained in this study provided evidence for a unique triparental origin of triploid onion A. × cornutum with three putative parental species, A. cepa, A. pskemense, and A. roylei.

Formamide-Free Genomic in situ Hybridization Allows Unambiguous Discrimination of Highly Similar Parental Genomes in Diploid Hybrids and Allopolyploids.

Polyploidy and hybridization play an important role in plant diversification and speciation. The application of genomic in situ hybridization (GISH) allows the identification of parental genomes in hybrids, thus elucidating their origins and allowing for analysis of their genomic evolution. The performance of GISH depends on the similarity of the parental genomes and on the age of hybrids. Here, we present the formamide-free GISH (ff-GISH) protocol applied to diploid and polyploid hybrids of monocots (Prospero, Hyacinthaceae) and dicots (Melampodium, Asteraceae) differing in similarity of the parental genomes and in chromosome and genome sizes. The efficiency of the new protocol is compared to the standard GISH protocol. As a result, ff-GISH allowed efficient labeling and discrimination of the parental chromosome sets in diploid and allopolyploid hybrids in Prospero autumnale species complex. In contrast, the standard GISH protocol failed to differentiate the parental genomes due to high levels of similar repetitive DNA. Likewise, an unambiguous identification of parental genomes in allotetraploid Melampodium nayaritense (Asteraceae) was possible after ff-GISH, whereas the standard GISH hybridization performance was suboptimal. The modified method is simple and non-toxic and allows the discrimination of very similar parental genomes in hybrids. This method lends itself to modifications and improvements and can also be used for FISH.

Structural polymorphisms and distinct genomic composition suggest recurrent origin and ongoing evolution of B chromosomes in the Prospero autumnale complex (Hyacinthaceae).

Summary Supernumerary B chromosomes (Bs) are genomic parasitic components, originating from the A complement via chromosomal rearrangements, which follow their own evolutionary trajectories. They often contain repetitive DNAs, some shared with regular chromosomes and some newly evolved. Genomic composition, origin and evolution of Bs have been analysed in the chromosomally variable Prospero autumnale complex. Two rDNAs and a satellite DNA (PaB6) from regular chromosomes were mapped to Bs of 26 plants from three diploid cytotypes, their hybrids and polyploid derivatives. In homoploid diploid hybrids, genomic in situ hybridization (GISH) allowed B painting with the parental DNAs. Bs were structurally variable and highly enriched in 5S rDNA and satDNA PaB6, and rarely in 35S rDNA. Eleven combinations of rDNA and PaB6 localization were observed. The quantities of PaB6 in Bs and regular chromosomes were not correlated, suggesting amplification mechanisms other than recombination. PaB6 and 5S rDNA amounts increased with increasing ploidy level. GISH revealed two independent origins of Bs. The structural variation, repeat content, repeat‐type fluctuations and differing genomic affinities of Bs in different cytotypes suggest that they represent young proto‐B chromosomes. Bs in P. autumnale probably form recurrently as by‐products of the extensive genome restructuring within this chromosomally variable species complex.

rDNA Loci Evolution in the Genus Glechoma (Lamiaceae).

Glechoma L. (Lamiaceae) is distributed in eastern Asia and Europe. Understanding chromosome evolution in Glechoma has been strongly hampered by its small chromosomes, constant karyotype and polyploidy. Here phylogenetic patterns and chromosomal variation in Glechoma species are considered, using genome sizes, chromosome mapping of 5S and 35S rDNAs by fluorescence in situ hybridisation (FISH), and phylogenetic analyses of internal transcribed spacers (nrITS) of 35S rDNA and 5S rDNA NTS sequences. Species and populations of Glechoma are tetraploid (2n = 36) with base chromosome number of x = 9. Four chromosomes carry pericentric 5S rDNA sites in their short arms in all the species. Two to four of these chromosomes also carry 35S rDNA in subterminal regions of the same arms. Two to four other chromosomes have 35S rDNA sites, all located subterminally within short arms; one individual possessed additional weak pericentric 35S rDNA signals on three other chromosomes. Five types of rDNA locus distribution have been defined on the basis of 35S rDNA variation, but none is species-specific, and most species have more than one type. Glechoma hederacea has four types. Genome size in Glechoma ranges from 0.80 to 0.94 pg (1C), with low levels of intrapopulational variation in all species. Phylogenetic analyses of ITS and NTS sequences distinguish three main clades coinciding with geographical distribution: European (G. hederacea–G. hirsuta), Chinese and Korean (G. longituba), and Japanese (G. grandis). The paper presents the first comparative cytogenetic analyses of Glechoma species including karyotype structure, rDNA location and number, and genome size interpreted in a phylogenetic context. The observed variation suggests that the genus is still in genomic flux. Genome size, but not rDNA loci number and distribution, provides a character for species delimitation which allows better inferences of interspecific relationships to be made, in the absence of well-defined morphological differentiation.

Multiple Origins and Nested Cycles of Hybridization Result in High Tetraploid Diversity in the Monocot Prospero

Polyploidy is a major driving force in angiosperm evolution, but our understanding of establishment and early diversification processes following allo- vs. auto-polyploidy is limited. An excellent system to address such questions is the monocot plant Prospero autumnale, as it comprises several genomically and chromosomally distinct diploid cytotypes and their auto- and allotetraploid derivatives. To infer origins and evolutionary trajectories of the tetraploids, we use genome size data, in situ hybridization with parental genomic DNAs and specific probes (satDNA, rDNAs), as well as molecular-phylogenetic analyses. Thus, we demonstrate that an astounding range of allotetraploid lineages has been formed recurrently by chromosomal re-patterning, interactions of chromosomally variable parental genomes and nested cycles of extensive hybridization, whereas autotetraploids have originated at least twice and are cytologically stable. During the recurrent formation and establishment across wide geographic areas hybridization in some populations could have inhibited lineage diversification and nascent speciation of such a hybrid swarm. However, cytotypes that became fixed in populations enhanced the potential for species diversification, possibly exploiting the extended allelic base, and fixed heterozygosity that polyploidy confers. The time required for polyploid cytotype fixation may in part reflect the lag phase reported for polyploids between their formation and species diversification.

Euchromatic Supernumerary Chromosomal Segments—Remnants of Ongoing Karyotype Restructuring in the Prospero autumnale Complex?

Supernumerary chromosomal segments (SCSs) represent additional chromosomal material that, unlike B chromosomes, is attached to the standard chromosome complement. The Prospero autumnale complex (Hyacinthaceae) is polymorphic for euchromatic large terminal SCSs located on the short arm of chromosome 1 in diploid cytotypes AA and B7B7, and tetraploid AAB7B7 and B6B6B7B7, in addition to on the short arm of chromosome 4 in polyploid B7B7B7B7 and B7B7B7B7B7B7 cytotypes. The genomic composition and evolutionary relationships among these SCSs have been assessed using fluorescence in situ hybridisation (FISH) with 5S and 35S ribosomal DNAs (rDNAs), satellite DNA PaB6, and a vertebrate-type telomeric repeat TTAGGG. Neither of the rDNA repeats were detected in SCSs, but most contained PaB6 and telomeric repeats, although these never spanned whole SCSs. Genomic in situ hybridisation (GISH) using A, B6, and B7 diploid genomic parental DNAs as probes revealed the consistently higher genomic affinity of SCSs in diploid hybrid B6B7 and allopolyploids AAB7B7 and B6B6B7B7 to genomic DNA of the B7 diploid cytotype. GISH results suggest a possible early origin of SCSs, especially that on chromosome 1, as by-products of the extensive genome restructuring within a putative ancestral P. autumnale B7 genome, predating the complex diversification at the diploid level and perhaps linked to B-chromosome evolution.

Contents Vol. 146, 2015

Jacqueline Smith Division of Genetics and Genomics Roslin Institute, Roslin Midlothian EH25 9PS (UK) Tel. (+44) 131 527 4200 Fax (+44) 131 440 0434 E-mail: jacqueline.smith@bbsrc.ac.uk Plant cytogenetics and genomics Andreas Houben Institute of Plant Genetics and Crop Plant, Research (IPK) Corrents-Str. 3 Gatersleben, D–06466 (Germany) Tel. (+1) 785 532 2364; Fax (+1) 785 532 5692 E-mail: houben@ipk-gatersleben.de