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Genetic map-based studies of reticulate evolution in plants
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Genetic map-based studies of reticulate evolution in plants Loren H. Rieseberg and Richard D. Noyes Botanists have long been interested in the role of hybridization in plant evolution. There are several pathways by which hybrid lineages can be stabilized (apomixis, allopolyploidy and recombinational speciation), and technological advances in genetic mapping have enhanced our understanding of these pathways. Important recent discoveries include a simple genetic basis of apomixis, the dynamic nature of nascent allopolyploid genomes and the critical role of fertility selection in recombinational speciation.
arwin1 viewed speciation as a strictly divergent process in which a new species diverged over time from an ancestral form. Although he recognized the existence of hybrids, they were not viewed as evidence for reticulate evolution (Box 1), because many hybrids were almost completely sterile (reticlate evolution refers to the grafting together of formerly separate branches on an evolutionary tree). Moreover, it had been shown that latergeneration hybrids typically revert back to the parental forms2. Near the time of Darwin’s publication, however, botanists were finding evidence that in some instances hybrids were constant and fertile and did not segregate back towards the parental species2. Later evidence indicated that the constancy of these hybrids resulted from either apomixis or allopolyploidy3,4. In the early part of this century, it was shown that hybrid reproduction could also be stabilized at the diploid level through ‘recombinational speciation’. Since then, considerable evidence has accumulated concerning the mechanistic basis of reticulate evolution in plants, and now the application of genetic mapping is having a major impact5,6. Mapping data contribute in two ways. First, the kinds of genomic changes that accompany or facilitate hybrid speciation can be inferred by tracking the linear order, number and distribution of ancestral species markers in hybrid and allopolyploid genomes. Second, mapped markers provide a powerful means of dissecting the genetic basis of complex traits that facilitate hybrid reproduction (e.g. apomixis) or that have been duplicated during allopolyploidy. Here we review several mechanisms by which fertile, stable lineages can arise via hybridization, emphasizing the contributions of genetic linkage mapping data to our understanding of these mechanisms.
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Apomixis
Apomixis includes diverse developmental pathways by which a plant reproduces asexually via seed7,8. The most widespread form of apomixis is gametophytic apomixis, in which plants produce unreduced gametophytes via apospory or diplospory, and embryos via diploid parthenogenesis or apogamety. Most gametophytic apomicts are polyploid, of hybrid origin, and display facultative rather than obligate apomixis9. In the absence of recombination (and excepting mutation), hybrid genotypes are perpetuated indefinitely by apomixis. Early on, apomixis was considered to be a byproduct of hybridization and polyploidy and to be complex genetically10. Later work based on crosses, however, suggested monogenic inheritance and either dominant or ploidy-dependent expression7,11. A possible one-locus genetic mechanism for apomixis has been proposed12, according to which asexual seed formation results from a mutation in the gene controlling meiotic divisions in the ovule. In this model, elimination of meiotic divisions is thought to advance megagametophyte development and pleiotropically trigger parthenogenetic development. Similarly, it has been proposed that apomixis 254
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results from a complex conflict between sets of asynchronous developmental genes13. Traditionally, apomicts have been considered to be evolutionary dead ends10. However, it has been well demonstrated that, through pollen and occasional reduced ovules, apomicts can participate in crosses both with other agamosperms and with sexual plants, and might act as intermediaries in gene flow between otherwise reproductively isolated taxa14. It has even been suggested that apomictic taxa might represent an early stage in the formation of sexual polyploids characterized by atypical (polysporic or polyembryonic) developmental schemes13. Current interest in apomixis is motivated principally by the goal of introducing the trait into agricultural crops in order to fix desirable heterozygous genotypes and to facilitate propagation15,16. Thus, unlike studies of polyploid and recombinational speciation, the aim of genetic map-based studies of apomicts is to identify apomixis-linked markers to facilitate crossing programs and to use knowledge of genomic architecture to understand the inheritance and molecular biology of apomixis. The best-studied example involves hybrids synthesized to introgress apomixis into maize from apomictic Tripsacum spp. Several markers that are consistently associated with diplospory in segregating progeny of a maize– Tripsacum cross have been identified17. Comparisons with a known linkage map for maize showed that all of the relevant markers mapped to the distal end of maize chromosome 6, suggesting the presence of a single locus controlling diplosporous gametophyte development. Additional cosegregating markers corresponding to the maize 6L linkage group have been identified by subsequent work18. Hybrid aneuploid and translocation lines, in combination with marker data, showed that all of the markers, and thus the putative genes for diplospory, occur on the long arm of Tripsacum chromosome 16. Introgression of this single fragment correlates with diplospory in maize hybrids. High-density marker maps of this region are being constructed and may well reveal details of the architecture and function of the factors controlling apomixis in plants. The combination of highly detailed genome-wide genetic linkage maps and genomic synteny facilitate the identification of apomixis markers in diverse grass species. For instance, initial screenings in Brachiaria with evenly dispersed markers of known locations in the maize genome revealed a single marker segregating with apospory19. Concentrated analysis of markers belonging to the same maize linkage group were subsequently used to construct a genetic map of the region (Fig. 1). Two molecular markers that were consistently associated with apospory in hybrid progeny from several crosses between apomictic and sexual species of pearl millet (Pennisetum spp.) have also been identified20. Because the markers were not observed to segregate separately in sexual taxa, these data were interpreted as evidence that the aposporous phenotype resulted from the introgression of a
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Box 1. Glossary of terms
Brachiaria sp.
Maize chromosome 5
opc4
Allopolyploid: A polyploid in which one or more sets of chromosomes come from different species or widely different strains. Aneuploid: Individual or population that differs in the number of individual chromosomes. Apogamety: The presence of an apomictic pathway in which a cell of an unreduced megagametophyte other than the egg gives rise to the embryo. Apomixis: Asexual reproduction via seed. Apospory: In apomixis, the formation of an unreduced megagametophyte via mitotic divisions of the nucellus or other vegetative cells of the ovule. Clade: All of the organisms descended from a particular common ancestor. Diploid parthenogenesis: Apomictic pathway in which an egg cell of an unreduced gametophyte develops into the embryo. Diplospory: In apomixis, the formation of an unreduced megagametophyte via mitotic divisions of the megasporocyte. Disomic inheritance: Random segregation of alleles at two homologous loci. Gametophytic apomixis: The presence of apomictic pathways that include the development of an unreduced megagametophyte via apospory or diplospory. Homeologous chromosomes: Homologous chromosomes derived from different parental species in an allopolyploid. Reticulate evolution: The fusing of previously separated branches on an evolutionary tree. Synteny: Conserved gene order. Tetrasomic inheritance: Random segregation of alleles at four homologous loci.
single chromosome fragment and possibly a single ‘apomixis locus’ to which both markers were linked. Surveys of these markers across 19 sexual and apomictic Pennisetum spp.21 revealed that one of them, UGT197, is consistently associated with apomictic taxa in all but one section of the genus surveyed. This study is significant because its demonstration of the fidelity of markers linked to apomictic development across species suggests that phylogenetic methods may potentially be used to test hypotheses that apomixis genes move among and across clades via introgressive hybridization. Because research has been so keenly focused on the chromosome fragment specific for apomixis, general knowledge of the structure and recombinational behavior of the genomes of apomictic species is in its infancy. However, in a recent study of the tetraploid apomict Paspalum simplex, analysis of heterozygous RFLP profiles in first generation (F1) progeny resulting from crosses with sexual plants revealed patterns consistent with tetrasomic inheritance, thus providing evidence for an autotetraploid origin of the apomictic lineage22. Studies like this are just a first step in developing genome maps of apomicts. Construction of such detailed genetic maps would be highly informative about the nature of recombination (rates and distribution) in apomictic lineages. Furthermore, comparative studies of detailed linkage maps among apomicts and putative sexual progenitors could well help to identify the origin and architecture of apomictic genomes and be used to study genome evolution under relaxed recombination. Although many of the studies discussed provide evidence that gametophytic apomixis is associated with a single chromosome fragment, the number of genes at the locus or the specific function of the genes has not yet been determined, and the interplay between heterozygosity, polyploidy and expression of the apomictic phenotypes is not well understood. However, with ever more detailed genetic maps in hand, the cloning of apomixis-associated
csu134 csu149 Apomixisrelated locus
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umc72 csu137 csu134
umc72 umc90
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Fig. 1. Comparison of the chromosomal region related to apomixis in Brachiaria and the homeologous fragment of maize chromosome 5. Molecular markers mapped in both taxa are shown. Broken lines indicate homologous markers. Modified from Ref. 19.
chromosomal fragments for use in transformation studies appears to be imminent. This avenue of research, perhaps in combination with comparative transcription studies, may reveal the mechanistic basis of apomixis and permit the manipulation of the trait for agronomic purposes. Allopolyploidy
Allopolyploidy is the most frequent solution to the problems of hybrid sterility and segregation10. In its simplest form, genome duplication in hybrids leads to the formation of meiotically normal and gametically fertile allopolyploids. Genome duplication also leads to instantaneous reproductive isolation between the new allopolyploid species and its parents. The situation is more complex when the polyploid hybrids are derived from more closely related genomes. In the absence of substantial chromosomal or genic differentiation (i.e. autopolyploidy), homeologous chromosome pairing and tetrasomic inheritance can be observed23. This may or may not involve multivalent formation at meiosis or a reduction in fertility. It also is possible that, within a polyploid hybrid, some sets of chromosomes will be well-differentiated and exhibit disomic inheritance, whereas other sets will show little differentiation and display tetrasomic inheritance10. This ‘segmental allopolyploid’ behavior may be common in early-generation synthetic polyploids, but tends to be absent in established polyploids24. In some instances, segmental allopolyploids appear quickly to assume the behavior of autopolyploids because of occasional homeologous pairing24. This leads to a feedback effect: recombination between homeologs increases their similarity, July 1998, Vol. 3, No. 7
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The utility of genetic mapping for studying allopolyploid speciation was first demonC strated in maize28. Maize is considered to D be an ancient polyploid, and RFLP markers H D1 were used to determine the number and loC cation of duplicated sequences in its genD ome28. Although approximately 30% of the E maize loci examined were duplicated, they H2 were distributed in a complex manner not consistent with simple allopolyploidy. It was B F therefore suggested that either the dupliA B G cated sequences were produced by segmental duplication in the absence of polyploidy G C E or that numerous translocations and inverD2 sions had occurred in maize following polyploidization28. Follow-up studies of RFLP B variation in maize suggested that a much F higher proportion of loci (>70%) are dupliH E A cated33, providing further support for the A polyploidy argument. Moreover, analyses of coalescence times for duplicated sequences F in the maize genome revealed two distinctive groups of sequences, which correspond roughly to 20.5 and 11.4 million years ago35. E G This suggests that the diploid progenitors of maize diverged approximately 20.5 million years ago, and the allotetraploid event ocFig. 2. Genetic map of Brassica nigra showing the distribution of eight triplicated sets of chromosomal segments (A–H). The triplicated segments, which carry homologous sets of curred nine to ten million years later. loci, are indicated by letters (A–H) and color. In two instances, a segment has been broken Genetic mapping of other crop plants has up and now occurs in more than one place in the genome (i.e. D1/D2 and H1/H2). revealed several additional examples of anHomeologous triplicated segments differ in length, suggesting that recombination rates cient polyploidy30,34,36. For example, an RFLP may not be conserved following karyotypic evolution. Modified from Ref. 34. map of ‘diploid’ Brassica nigra (n = 8, where n is the haploid chromosome number) reveals that its genome is extensively duplicated which in turn increases the frequency of pairing, and so forth. (Fig. 2) and appears to consist of three complete, but rearranged, Alternatively, rapid divergence between genomes may lead to a copies of a common ancestral genome34. Comparison of the B. cessation of homeologous recombination and stable allopolyploid nigra map with that of two other ‘diploid’ Brassica spp., B. rapa behavior25. (n = 10) and B. oleracea (n = 9), indicates that the three are reTheoretical studies have identified several factors that favor the markably similar in genome content, although some differences in establishment of polyploids in nature26,27. These include differen- gene order are apparent. The simplest explanation for this pattern tial niche preference, a selfing mating system, high fecundity and is that Brassica descended from an ancestral hexaploid approxian iteroparous life history (repeated opportunities for reproduc- mately ten million years ago. However, the comparative mapping tion). Niche separation and selfing enhance the probability of suc- data also suggest considerable karyotypic evolution since the cessful matings during the early stages of establishment. Random polyploidization event: a minimum of 24 chromosomal rearrangemating would result in a preponderance of mating with the diploid ments are required to explain observed differences in gene order34. parental species and subsequent production of sterile triploid seed. In addition to providing evidence of duplicated genes and genStochastic events due to a small number of polyploid colonizers omes, mapping data can be used to examine the evolutionary fate of decrease the chance of establishment, but this barrier is minimized duplicated sequences. An example of this application comes from by high fecundity and iteroparity. In fact, Roderiguez27 concludes studies of ancient polyploid soybean, in which quantitative trait loci that ‘the establishment of polyploids in higher plants is not an un- for seed protein and oil occurred in concordant positions across likely event’, a conclusion that is supported by the high frequency homeologous chromosomes30. Thus, genes affecting these traits apof polyploidy in plants and recent evidence for the multiple origins pear to have retained similar functions following polyploidization. of many polyploid species, even including those originating in Evidence for ancient polyploidy is not limited to plants. For this century23. example, accumulating data hint at an ancient polyploid origin for Mapping data indicate that considerable genetic change accom- most vertebrates. Many single invertebrate genes correspond to as panies allopolyploid speciation. Homeologous genomes in allo- many as four equally related vertebrate genes on different chromopolyploid species appear to be highly rearranged relative to each somes37. Jawless hagfish and lampreys appear to be allotetraploids, other28–30 and to their diploid progenitors31 (although see Ref. 32). whereas jawed vertebrates from fish to humans are likely to be alloMany of these changes appear to occur rapidly following poly- octoploids37. Likewise, analyses of duplicate genes suggest that ploidization6, suggesting that they are an important component of yeast represents a degenerate tetraploid that arose roughly 100 milpolyploid speciation. Mapping data have also served to confirm lion years ago38. Although most of the duplicated genes have apsuspected examples of ancient polyploidy33 and to indicate some un- parently since been deleted, and gene order is highly rearranged, expected and even more ancient polyploid events34. the evidence for allotetraploidy is quite strong. For 50 out of 55 H1
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trends in plant science reviews duplicate regions, gene orientation with respect to the centromere is the same in both copies38. Additionally, if this pattern arose by successive duplication rather than allotetraploidy, at least some triplicated regions should be expected: none has been reported.
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The distribution of duplicated sequences has Q R 199 B6 also been studied in several neopolyploid 313 taxa. In tetraploid cotton, for example, map443 ping of duplicate loci allowed identification of 11 of the 13 expected homeologous pairs 227 rp1 between the A and D subgenomes29. Two of nf73 these pairs are collinear; four differ by at least one inversion; two differ by at least two inversions; and at least one pair shows a Synthesized H. anomalus hybrids translocation. To determine whether the ex239 tensive chromosomal reorganization between Synthesized H. anomalus hybrids the A and D genomes occurred before or after 103 polyploidization, the genomes of the A and D diploid progenitors have also been mapped (C.L. Brubaker et al., unpublished). Comparison of the diploid and polyploid genH. petiolaris markers or S linkage blocks 295 omes revealed that the A and D genomes of the diploid species were most similar in A15 gene order, and the A and D subgenomes of H. annuus markers or the tetraploid were most divergent. This linkage blocks suggests that rates of structural evolution increased following polyploidization. Experimental study has also revealed evi40 cM dence for rapid genomic evolution following 471 6 polyploidization . The most detailed studies come from synthetic Brassica allotetraploids generated by interspecific hybridizations between ‘diploid’ Brassica spp.: B. rapa (A genome); B. nigra (B genome); and B. oleracea Synthesized H. anomalus hybrids (C genome). The crosses were: A ⫻ B; B ⫻ A; C ⫻ A; and A ⫻ C (Ref. 6). The AB and Fig. 3. Genomic composition of ancient and experimental hybrid lineages for three selected BA synthetic tetraploids have the same nulinkage groups. Letters at the left of each linkage group designate linkage blocks in Helianthus clear genome complement as the natural neoanomalus and indicate homology to linkages previously mapped in the parental species, tetraploid, B. juncea (n = 18), and the AC H. annuus and H. petiolaris5 (the broken line indicates the break-point between the R and and CA synthetic tetraploids have the same S blocks of that linkage group). The distribution of parental chromosomal blocks in the synthesized hybrids (left linkage group) and H. anomalus (right linkage group) is indicated by nuclear genome complement as B. napus colored bars. Regions harboring recombination points are indicated by a color scale, with (n = 19). RFLP comparisons of F2 and F5 the intensity of shading indicating the likelihood that a particular region was derived from plants revealed evidence for rapid genome one parent or the other. Modified from Ref. 5. change. On average, 9.6% of restriction fragments were dissimilar between F2 and F5 plants from the AB line, 8.2% from the BA line, 4.1% from the AC line, and 3.7% from the CA line. Notably, provide a source of genetic variation for adaptive evolution and dithe paternal subgenome in the AB and BA tetraploids showed versification within polyploid lineages. Additionally, if the kinds greater differentiation from its diploid parent than did the mater- of genomic changes observed here are arbitrary, it seems likely that nal subgenome, but no significant differences in subgenome evo- polyploid lineages independently derived from the same parental lution were observed in the AC and CA tetraploids. This suggests material can rapidly diverge and potentially achieve reproductive that cytoplasmic–nuclear interactions influenced genomic change isolation. This could be explored by testing the compatibility of in the AB and BA tetraploids, but not in the AC and CA tetra- polyploid populations that molecular evidence has shown to be ploids. Presumably, the lack of significant cytoplasmic effects in independently derived from the same parental species. the AC and CA tetraploids reflects higher levels of cytonuclear compatibility, because the A and C subgenomes are more closely Recombinational speciation related phylogenetically than are the A and B subgenomes. The most widely accepted model for diploid or homoploid hybrid As stressed by Song et al.6, these data have significant impli- speciation is the recombinational model described by Grant39. cations for our understanding of polyploid evolution. They demon- According to this model, the sorting of chromosomal rearrangestrate that much of the genomic change found in natural polyploids ments in later-generation hybrids could, by chance, lead to the probably accompanies polyploid formation. These changes possibly formation of new populations that are homozygous for a novel July 1998, Vol. 3, No. 7
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trends in plant science reviews In comparison to allopolyploidy, researchers have expended much less effort towards understanding the mechanistic basis of recombinational speciation. The only relevant studies concern the origin of a wild hybrid sunflower species, Helianthus anomalus5,42.
100
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Fig. 4. Mean pollen fertility in three synthetic hybrid lineages between Helianthus annuus and H. petiolaris. Lineage pedigrees are as follows (generation number in parentheses): lineage I, P(0)F1(1)-BC1(2)-BC2(3)-F2(4)-F3(5); lineage II, P(0)-F1(1)-F2(2)BC1(3)-BC2(4)-F3(5); and lineage III, P(0)-F1(1)-F2(2)-F3(3)BC1(4)-BC2(5). Abbreviations: P, parental; F, sibmating; BC, backcrossing. All backcrosses were in the direction of H. annuus. For each line, 100 pollen grains from each of 20 plants per generation were tested by viability staining. Standard error bars are omitted for clarity.
combination of rearrangements. The new hybrid population would be fertile, stable and have the same ploidy as its parents, yet would be at least partially isolated from both parental species by a chromosomal sterility barrier. Although this model focused on chromosomal rearrangements, it is clear that the sorting of genic sterility factors could generate similar results. Thus, current models incorporate both genic and chromosomal sterility factors40. Other factors that appear to play a critical role in recombinational speciation include: strong natural selection for the most fertile or viable hybrid segregants5,41; rapid chromosomal evolution42; and the availability of habitats suitable for the establishment of hybrid neospecies40. The feasibility of the recombinational model has been explored experimentally via crossing studies40. These studies validated the recombinational model by demonstrating that fertile and viable hybrid lineages can be obtained after only a small number of generations (<10) of selfing and/or backcrossing, even if F1 hybrids were almost completely sterile. Furthermore, the experimentally generated hybrid lineages were often strongly reproductively isolated from the parental species. Estimating the frequency of homoploid hybrid speciation in nature is more difficult, and fewer than ten cases have been rigorously documented for plants40. However, these low numbers may be an artifact of the difficulty of detecting and documenting homoploid hybrid species, particularly if the hybridization events are ancient. A much larger number of hybrid species has been proposed, and molecular phylogenetic studies continually uncover unexpected cases of ancient hybridization, some of which may have led to speciation43. Also, hybridization may play an important role as the creative stimulus for speciation in small or peripheral populations. Hybridization rates appear to be highest in populations with these characteristics, and hybridization may be more plausible than population bottlenecks for generating the genetic or chromosomal reorganization proposed in founder-effect speciation40. 258
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Genetic models suggest that reproductive isolation between the hybrid neospecies and species populations can be facilitated by rapid karyotypic evolution39–41. This hypothesis has been tested by comparing genetic maps for H. anomalus and its putative parental species, H. annuus and H. petiolaris42. The mapping data revealed extensive chromosomal differentiation between the hybrid and parental genomes. Seven linkage groups in the hybrid genome differed in gene order from both parental species, and three chromosomal breakages, three fusions and one duplication are required to explain these differences. Chromosomal differentiation does appear to have facilitated reproductive isolation in H. anomalus; F1 hybrids with its putative parental species are partially sterile because of meiotic abnormalities42. The role of selection
To assess the relative contributions of deterministic and stochastic forces in the recombinational speciation process, three independent hybrid lineages were synthesized between H. annuus and H. petiolaris5. Comparison of the genomic composition of the ancient (H. anomalus) and synthetic hybrid lineages revealed that all three synthetic hybrid lineages had converged to nearly identical gene combinations, and this set of combinations was statistically concordant with that of H. anomalus (Fig. 3). Similarity in genomic composition between the synthetic and ancient hybrids suggests that deterministic forces such as selection, rather than stochastic forces, largely govern the formation of ‘recombinational’ species. Because the synthetic hybrid lineages were generated in the greenhouse, fertility selection probably played a greater role than ecological selection in shaping hybrid genomic composition. This conclusion is supported by the rapid increase in fertility observed in the three synthetic hybrid lineages; average pollen fertility increased from 4% in the F1 generation to >90% in the fifth-generation hybrids (Fig. 4). Congruence in genomic composition also implies that the genomic structure and composition of hybrid species is essentially fixed after a small number of generations of hybridization and remain relatively static thereafter. Tempo of recombinational speciation
Simulation studies suggest that recombinational speciation is punctuated: long periods of hybrid-zone stasis are followed by abrupt transitions in which parental individuals are displaced rapidly by the hybrid neospecies41. The hypothesis of rapid speciation was tested by analyzing the sizes of parental species chromosome blocks in the H. anomalus genome (M. Ungerer et al., unpublished). During the evolution of a new hybrid species, parental linkage block sizes are expected to become progressively smaller over time because of recombination. However, continued reductions in block size will be countered by structural fixation of the hybrid genome; subsequent recombination among blocks derived from the same parental species will no longer decrease block size. To estimate the number of generations of recombination required to achieve the present distribution of parental chromosomal block sizes in the H. anomalus genome, observed block sizes were compared with those of a computer-simulated hybrid population (M. Ungerer et al., unpublished). This comparison suggests that H. anomalus arose extremely rapidly, probably in fewer than 60 generations. This result is corroborated by the independent lines of evidence already discussed, which demonstrate
trends in plant science reviews significant concordance between the genomes of H. anomalus and early-generation synthetic hybrids (Fig. 3), and a rapid recovery of fertility in these synthetic hybrid lineages (Fig. 4). Conclusions
Understanding of how hybrid lineages are stabilized has advanced considerably over the past five years. Many of these advances are due to the application of recently developed genetic mapping technology. Indeed, reticulate modes of speciation are arguably far better understood than their divergent counterparts, such as geographical speciation. The rapid progress that has characterized the study of reticulate evolution is due in part to the application of genetic mapping tools, but has also benefited from allopolyploid and recombinational speciation being rapid modes of speciation that are easily studied experimentally. It seems likely that the combination of experimental genetic and historical approaches that have proven so powerful for the study of hybrid lineages will have a similar impact on studies of divergent modes of evolution. Acknowledgements
Our work on reticulate evolution has been funded by the National Science Foundation of the USA (DEB9615335 and DEB9701347). We thank Seung Chul Kim and Rhonda Rieseberg for helpful comments on an earlier version of the manuscript. References 01 Darwin, C. (1859) On the Origin of Species by Means of Natural Selection, or the Preservation of Favored Races in the Struggle for Life, John Murray 02 Roberts, H.F. (1929) Plant Hybridization before Mendel, Princeton University Press 03 Lotsy, J.P. (1916) Evolution by Means of Hybridization, M. Nijhoff 04 Winge, Ö. (1917) The chromosomes: their number and general importance, C. R. Trav. Lab. Carlsberg 13, 131–275 05 Rieseberg, L.H. et al. (1996) Role of gene interactions in hybrid speciation: evidence from ancient and experimental hybrids, Science 272, 741–745 06 Song, K. et al. (1995) Rapid genome change in synthetic polyploids of Brassica and its implications for polyploid evolution, Proc. Natl. Acad. Sci. U. S. A. 92, 7719–7723 07 Nogler, G.A. (1984) Gametophytic apomixis, in Embryology of Angiosperms (Johri, B.M., ed.), pp. 475–518, Springer 08 Asker, S.E. and Jerling, L. (1992) Apomixis in Plants, CRC Press 09 Grant, V. (1981) Plant Speciation, Columbia University Press 10 Stebbins, G.L. (1950) Variation and Evolution in Plants, Columbia University Press 11 Savidan, Y. (1980) Chromosomal and embryological analyses in sexual ⫻ apomictic hybrids of Panicum maximum Jacq., Theor. Appl. Genet. 57, 153–156 12 Mogie, M. (1992) The Evolution of Asexual Reproduction in Plants, Chapman & Hall 13 Carman, J.G. (1997) Asynchronous expression of duplicate genes in angiosperms may cause apomixis, bispory, tetraspory, and polyembryony, Biol. J. Linn. Soc. 61, 51–94 14 De Wet, J.M.J. and Harlan, J.R. (1970) Apomixis, polyploidy, and speciation in Dichanthium, Evolution 24, 270–277 15 Jefferson, R.A. and Bicknell, R. (1996) The potential impacts of apomixis: a molecular genetics approach, in The Impact of Plant Molecular Genetics (Sobral, B.W.S., ed.), pp. 87–101, Birkhäuser 16 Vielle-Calzada, J-P., Crane, C.F. and Stelly, D.M. (1996) Apomixis: the asexual revolution, Science 274, 1322–1323 17 Leblanc, O. et al. (1995) Detection of the apomictic mode of reproduction in maize–Tripsacum hybrids using maize RFLP markers, Theor. Appl. Genet. 90, 1198–1203 18 Kindiger, B., Bai, D. and Sokolov, V. (1996) Assignment of gene(s) conferring apomixis in Tripsacum to a chromosome arm: cytological and molecular evidence, Genome 39, 1133–1141
19 Pessino, S.C. et al. (1997) Identification of a maize linkage group related to apomixis in Brachiaria, Theor. Appl. Genet. 94, 439–444 20 Ozias-Akins, P. et al. (1993) Transmission of the apomictic mode of reproduction in Pennisetum: co-inheritance of the trait and molecular markers, Theor. Appl. Genet. 85, 632–638 21 Lubbers, E.L. et al. (1994) Molecular markers shared by diverse apomictic Pennisetum species, Theor. Appl. Genet. 89, 636–642 22 Pupilli, F. et al. (1997) Segregation analysis of RFLP markers reveals a tetrasomic inheritance in apomictic Paspalum simplex, Genome 40, 822–828 23 Soltis, D.E. and Soltis, P.S. (1993) Molecular data and the dynamic nature of polyploidy, Crit. Rev. Plant Sci. 12, 243–275 24 Sybenga, J. (1996) Chromosome pairing affinity and quadrivalent formation in polyploids: do segmental allopolyploids exist? Genome 39, 1176–1184 25 Leipold, M. and Schmidtke, J. (1982) Gene expression in phylogenetically polyploid organisms, in Genome Evolution (Dover, G. and Flavell, R., eds), pp. 219–236, Academic Press 26 Roderiguez, D.J. (1996) A model for the establishment of polyploidy in plants, Am. Nat. 147, 33–46 27 Roderiguez, D.J. (1996) A model for the establishment of polyploidy in plants: viable but infertile hybrids, iteroparity, and demographic stochasticity, J. Theor. Biol. 180, 189–196 28 Helentjaris, T., Weber D. and Wright, S. (1988) Identification of the genomic locations of duplicate nucleotide sequences in maize by analysis of restriction fragment length polymorphisms, Genetics 118, 353–363 29 Reinisch, A.J. et al. (1994) A detailed RFLP map of cotton, Gossypium hirsutum ⫻ Gossypium barbadense: chromosome organization and evolution in a disomic polyploid genome, Genetics 138, 829–847 30 Shoemaker, R.C. et al. (1996) Genome duplication in soybean (Glycine subgenus soja), Genetics 144, 329–338 31 Sharpe, A.G. et al. (1995) Frequent nonreciprocal translocations in the amphidiploid genome of oilseed rape (Brassica napus), Genome 38, 1112–1121 32 Parkin, I.A.P. and Lydiate, D.S. (1997) Conserved patterns of chromosome pairing and recombination in Brassica napus crosses, Genome 40, 496–504 33 Ahn, S. and Tanksley, S.D. (1993) Comparative linkage maps of the rice and maize genomes, Proc. Natl. Acad. Sci. U. S. A. 90, 7980–7984 34 Lagercrantz, U. and Lydiate, D.J. (1996) Comparative genome mapping in Brassica, Genetics 144, 1903–1910 35 Gaut, B.S. and Doebley, J.F. (1997) DNA sequence evidence for the segmental allotetraploid origin of maize, Proc. Natl. Acad. Sci. U. S. A. 94, 6809–6814 36 Chittenden, L.M. et al. (1994) A detailed RFLP map of Sorghum bicolor ⫻ S. propinquum, suitable for high-density mapping, suggests ancestral duplication of Sorghum chromosomes or chromosomal segments, Theor. Appl. Genet. 87, 925–933 37 Spring, J. (1997) Vertebrate evolution by interspecific hybridization – are we polyploid? FEBS Lett. 400, 2–8 38 Wolfe, K.H. and Shields, D.C. (1997) Molecular evidence for an ancient duplication of the entire yeast genome, Nature 387, 708–713 39 Grant, V. (1958) The regulation of recombination in plants, Cold Spring Harbor Symp. Quant. Biol. 23, 337–363 40 Rieseberg, L.H. (1997) Hybrid origins of plant species, Annu. Rev. Ecol. Syst. 28, 359–389 41 McCarthy, E.M., Asmussen, M.A. and Anderson, W.W. (1995) A theoretical assessment of recombinational speciation, Heredity 74, 502–509 42 Rieseberg, L.H., Van Fossen, C. and Desrochers, A. (1995) Hybrid speciation accompanied by genomic reorganization in wild sunflowers, Nature 375, 313–316 43 Wendel, J.F., Stewart, J.M. and Rettig, J.H. (1991) Molecular evidence for homoploid reticulate evolution among Australian species of Gossypium, Evolution 45, 694–711
Loren Rieseberg* and Rick Noyes are at the Dept of Biology, Indiana University, Bloomington, IN 47405, USA. *Author for correspondence (tel +1 812 855 7614; fax +1 812 855 6705; e-mail [email protected]).
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Genetic map-based studies of reticulate evolution in plants Loren H. Rieseberg and Richard D. Noyes Botanists have long been interested in the role of hybridization in plant evolution. There are several pathways by which hybrid lineages can be stabilized (apomixis, allopolyploidy and recombinational speciation), and technological advances in genetic mapping have enhanced our understanding of these pathways. Important recent discoveries include a simple genetic basis of apomixis, the dynamic nature of nascent allopolyploid genomes and the critical role of fertility selection in recombinational speciation.
arwin1 viewed speciation as a strictly divergent process in which a new species diverged over time from an ancestral form. Although he recognized the existence of hybrids, they were not viewed as evidence for reticulate evolution (Box 1), because many hybrids were almost completely sterile (reticlate evolution refers to the grafting together of formerly separate branches on an evolutionary tree). Moreover, it had been shown that latergeneration hybrids typically revert back to the parental forms2. Near the time of Darwin’s publication, however, botanists were finding evidence that in some instances hybrids were constant and fertile and did not segregate back towards the parental species2. Later evidence indicated that the constancy of these hybrids resulted from either apomixis or allopolyploidy3,4. In the early part of this century, it was shown that hybrid reproduction could also be stabilized at the diploid level through ‘recombinational speciation’. Since then, considerable evidence has accumulated concerning the mechanistic basis of reticulate evolution in plants, and now the application of genetic mapping is having a major impact5,6. Mapping data contribute in two ways. First, the kinds of genomic changes that accompany or facilitate hybrid speciation can be inferred by tracking the linear order, number and distribution of ancestral species markers in hybrid and allopolyploid genomes. Second, mapped markers provide a powerful means of dissecting the genetic basis of complex traits that facilitate hybrid reproduction (e.g. apomixis) or that have been duplicated during allopolyploidy. Here we review several mechanisms by which fertile, stable lineages can arise via hybridization, emphasizing the contributions of genetic linkage mapping data to our understanding of these mechanisms.
D
Apomixis
Apomixis includes diverse developmental pathways by which a plant reproduces asexually via seed7,8. The most widespread form of apomixis is gametophytic apomixis, in which plants produce unreduced gametophytes via apospory or diplospory, and embryos via diploid parthenogenesis or apogamety. Most gametophytic apomicts are polyploid, of hybrid origin, and display facultative rather than obligate apomixis9. In the absence of recombination (and excepting mutation), hybrid genotypes are perpetuated indefinitely by apomixis. Early on, apomixis was considered to be a byproduct of hybridization and polyploidy and to be complex genetically10. Later work based on crosses, however, suggested monogenic inheritance and either dominant or ploidy-dependent expression7,11. A possible one-locus genetic mechanism for apomixis has been proposed12, according to which asexual seed formation results from a mutation in the gene controlling meiotic divisions in the ovule. In this model, elimination of meiotic divisions is thought to advance megagametophyte development and pleiotropically trigger parthenogenetic development. Similarly, it has been proposed that apomixis 254
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results from a complex conflict between sets of asynchronous developmental genes13. Traditionally, apomicts have been considered to be evolutionary dead ends10. However, it has been well demonstrated that, through pollen and occasional reduced ovules, apomicts can participate in crosses both with other agamosperms and with sexual plants, and might act as intermediaries in gene flow between otherwise reproductively isolated taxa14. It has even been suggested that apomictic taxa might represent an early stage in the formation of sexual polyploids characterized by atypical (polysporic or polyembryonic) developmental schemes13. Current interest in apomixis is motivated principally by the goal of introducing the trait into agricultural crops in order to fix desirable heterozygous genotypes and to facilitate propagation15,16. Thus, unlike studies of polyploid and recombinational speciation, the aim of genetic map-based studies of apomicts is to identify apomixis-linked markers to facilitate crossing programs and to use knowledge of genomic architecture to understand the inheritance and molecular biology of apomixis. The best-studied example involves hybrids synthesized to introgress apomixis into maize from apomictic Tripsacum spp. Several markers that are consistently associated with diplospory in segregating progeny of a maize– Tripsacum cross have been identified17. Comparisons with a known linkage map for maize showed that all of the relevant markers mapped to the distal end of maize chromosome 6, suggesting the presence of a single locus controlling diplosporous gametophyte development. Additional cosegregating markers corresponding to the maize 6L linkage group have been identified by subsequent work18. Hybrid aneuploid and translocation lines, in combination with marker data, showed that all of the markers, and thus the putative genes for diplospory, occur on the long arm of Tripsacum chromosome 16. Introgression of this single fragment correlates with diplospory in maize hybrids. High-density marker maps of this region are being constructed and may well reveal details of the architecture and function of the factors controlling apomixis in plants. The combination of highly detailed genome-wide genetic linkage maps and genomic synteny facilitate the identification of apomixis markers in diverse grass species. For instance, initial screenings in Brachiaria with evenly dispersed markers of known locations in the maize genome revealed a single marker segregating with apospory19. Concentrated analysis of markers belonging to the same maize linkage group were subsequently used to construct a genetic map of the region (Fig. 1). Two molecular markers that were consistently associated with apospory in hybrid progeny from several crosses between apomictic and sexual species of pearl millet (Pennisetum spp.) have also been identified20. Because the markers were not observed to segregate separately in sexual taxa, these data were interpreted as evidence that the aposporous phenotype resulted from the introgression of a
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Box 1. Glossary of terms
Brachiaria sp.
Maize chromosome 5
opc4
Allopolyploid: A polyploid in which one or more sets of chromosomes come from different species or widely different strains. Aneuploid: Individual or population that differs in the number of individual chromosomes. Apogamety: The presence of an apomictic pathway in which a cell of an unreduced megagametophyte other than the egg gives rise to the embryo. Apomixis: Asexual reproduction via seed. Apospory: In apomixis, the formation of an unreduced megagametophyte via mitotic divisions of the nucellus or other vegetative cells of the ovule. Clade: All of the organisms descended from a particular common ancestor. Diploid parthenogenesis: Apomictic pathway in which an egg cell of an unreduced gametophyte develops into the embryo. Diplospory: In apomixis, the formation of an unreduced megagametophyte via mitotic divisions of the megasporocyte. Disomic inheritance: Random segregation of alleles at two homologous loci. Gametophytic apomixis: The presence of apomictic pathways that include the development of an unreduced megagametophyte via apospory or diplospory. Homeologous chromosomes: Homologous chromosomes derived from different parental species in an allopolyploid. Reticulate evolution: The fusing of previously separated branches on an evolutionary tree. Synteny: Conserved gene order. Tetrasomic inheritance: Random segregation of alleles at four homologous loci.
single chromosome fragment and possibly a single ‘apomixis locus’ to which both markers were linked. Surveys of these markers across 19 sexual and apomictic Pennisetum spp.21 revealed that one of them, UGT197, is consistently associated with apomictic taxa in all but one section of the genus surveyed. This study is significant because its demonstration of the fidelity of markers linked to apomictic development across species suggests that phylogenetic methods may potentially be used to test hypotheses that apomixis genes move among and across clades via introgressive hybridization. Because research has been so keenly focused on the chromosome fragment specific for apomixis, general knowledge of the structure and recombinational behavior of the genomes of apomictic species is in its infancy. However, in a recent study of the tetraploid apomict Paspalum simplex, analysis of heterozygous RFLP profiles in first generation (F1) progeny resulting from crosses with sexual plants revealed patterns consistent with tetrasomic inheritance, thus providing evidence for an autotetraploid origin of the apomictic lineage22. Studies like this are just a first step in developing genome maps of apomicts. Construction of such detailed genetic maps would be highly informative about the nature of recombination (rates and distribution) in apomictic lineages. Furthermore, comparative studies of detailed linkage maps among apomicts and putative sexual progenitors could well help to identify the origin and architecture of apomictic genomes and be used to study genome evolution under relaxed recombination. Although many of the studies discussed provide evidence that gametophytic apomixis is associated with a single chromosome fragment, the number of genes at the locus or the specific function of the genes has not yet been determined, and the interplay between heterozygosity, polyploidy and expression of the apomictic phenotypes is not well understood. However, with ever more detailed genetic maps in hand, the cloning of apomixis-associated
csu134 csu149 Apomixisrelated locus
umc147
umc147
umc72 csu137 csu134
umc72 umc90
csu149
umc90
csu137
Fig. 1. Comparison of the chromosomal region related to apomixis in Brachiaria and the homeologous fragment of maize chromosome 5. Molecular markers mapped in both taxa are shown. Broken lines indicate homologous markers. Modified from Ref. 19.
chromosomal fragments for use in transformation studies appears to be imminent. This avenue of research, perhaps in combination with comparative transcription studies, may reveal the mechanistic basis of apomixis and permit the manipulation of the trait for agronomic purposes. Allopolyploidy
Allopolyploidy is the most frequent solution to the problems of hybrid sterility and segregation10. In its simplest form, genome duplication in hybrids leads to the formation of meiotically normal and gametically fertile allopolyploids. Genome duplication also leads to instantaneous reproductive isolation between the new allopolyploid species and its parents. The situation is more complex when the polyploid hybrids are derived from more closely related genomes. In the absence of substantial chromosomal or genic differentiation (i.e. autopolyploidy), homeologous chromosome pairing and tetrasomic inheritance can be observed23. This may or may not involve multivalent formation at meiosis or a reduction in fertility. It also is possible that, within a polyploid hybrid, some sets of chromosomes will be well-differentiated and exhibit disomic inheritance, whereas other sets will show little differentiation and display tetrasomic inheritance10. This ‘segmental allopolyploid’ behavior may be common in early-generation synthetic polyploids, but tends to be absent in established polyploids24. In some instances, segmental allopolyploids appear quickly to assume the behavior of autopolyploids because of occasional homeologous pairing24. This leads to a feedback effect: recombination between homeologs increases their similarity, July 1998, Vol. 3, No. 7
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G1
G2
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The utility of genetic mapping for studying allopolyploid speciation was first demonC strated in maize28. Maize is considered to D be an ancient polyploid, and RFLP markers H D1 were used to determine the number and loC cation of duplicated sequences in its genD ome28. Although approximately 30% of the E maize loci examined were duplicated, they H2 were distributed in a complex manner not consistent with simple allopolyploidy. It was B F therefore suggested that either the dupliA B G cated sequences were produced by segmental duplication in the absence of polyploidy G C E or that numerous translocations and inverD2 sions had occurred in maize following polyploidization28. Follow-up studies of RFLP B variation in maize suggested that a much F higher proportion of loci (>70%) are dupliH E A cated33, providing further support for the A polyploidy argument. Moreover, analyses of coalescence times for duplicated sequences F in the maize genome revealed two distinctive groups of sequences, which correspond roughly to 20.5 and 11.4 million years ago35. E G This suggests that the diploid progenitors of maize diverged approximately 20.5 million years ago, and the allotetraploid event ocFig. 2. Genetic map of Brassica nigra showing the distribution of eight triplicated sets of chromosomal segments (A–H). The triplicated segments, which carry homologous sets of curred nine to ten million years later. loci, are indicated by letters (A–H) and color. In two instances, a segment has been broken Genetic mapping of other crop plants has up and now occurs in more than one place in the genome (i.e. D1/D2 and H1/H2). revealed several additional examples of anHomeologous triplicated segments differ in length, suggesting that recombination rates cient polyploidy30,34,36. For example, an RFLP may not be conserved following karyotypic evolution. Modified from Ref. 34. map of ‘diploid’ Brassica nigra (n = 8, where n is the haploid chromosome number) reveals that its genome is extensively duplicated which in turn increases the frequency of pairing, and so forth. (Fig. 2) and appears to consist of three complete, but rearranged, Alternatively, rapid divergence between genomes may lead to a copies of a common ancestral genome34. Comparison of the B. cessation of homeologous recombination and stable allopolyploid nigra map with that of two other ‘diploid’ Brassica spp., B. rapa behavior25. (n = 10) and B. oleracea (n = 9), indicates that the three are reTheoretical studies have identified several factors that favor the markably similar in genome content, although some differences in establishment of polyploids in nature26,27. These include differen- gene order are apparent. The simplest explanation for this pattern tial niche preference, a selfing mating system, high fecundity and is that Brassica descended from an ancestral hexaploid approxian iteroparous life history (repeated opportunities for reproduc- mately ten million years ago. However, the comparative mapping tion). Niche separation and selfing enhance the probability of suc- data also suggest considerable karyotypic evolution since the cessful matings during the early stages of establishment. Random polyploidization event: a minimum of 24 chromosomal rearrangemating would result in a preponderance of mating with the diploid ments are required to explain observed differences in gene order34. parental species and subsequent production of sterile triploid seed. In addition to providing evidence of duplicated genes and genStochastic events due to a small number of polyploid colonizers omes, mapping data can be used to examine the evolutionary fate of decrease the chance of establishment, but this barrier is minimized duplicated sequences. An example of this application comes from by high fecundity and iteroparity. In fact, Roderiguez27 concludes studies of ancient polyploid soybean, in which quantitative trait loci that ‘the establishment of polyploids in higher plants is not an un- for seed protein and oil occurred in concordant positions across likely event’, a conclusion that is supported by the high frequency homeologous chromosomes30. Thus, genes affecting these traits apof polyploidy in plants and recent evidence for the multiple origins pear to have retained similar functions following polyploidization. of many polyploid species, even including those originating in Evidence for ancient polyploidy is not limited to plants. For this century23. example, accumulating data hint at an ancient polyploid origin for Mapping data indicate that considerable genetic change accom- most vertebrates. Many single invertebrate genes correspond to as panies allopolyploid speciation. Homeologous genomes in allo- many as four equally related vertebrate genes on different chromopolyploid species appear to be highly rearranged relative to each somes37. Jawless hagfish and lampreys appear to be allotetraploids, other28–30 and to their diploid progenitors31 (although see Ref. 32). whereas jawed vertebrates from fish to humans are likely to be alloMany of these changes appear to occur rapidly following poly- octoploids37. Likewise, analyses of duplicate genes suggest that ploidization6, suggesting that they are an important component of yeast represents a degenerate tetraploid that arose roughly 100 milpolyploid speciation. Mapping data have also served to confirm lion years ago38. Although most of the duplicated genes have apsuspected examples of ancient polyploidy33 and to indicate some un- parently since been deleted, and gene order is highly rearranged, expected and even more ancient polyploid events34. the evidence for allotetraploidy is quite strong. For 50 out of 55 H1
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trends in plant science reviews duplicate regions, gene orientation with respect to the centromere is the same in both copies38. Additionally, if this pattern arose by successive duplication rather than allotetraploidy, at least some triplicated regions should be expected: none has been reported.
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The distribution of duplicated sequences has Q R 199 B6 also been studied in several neopolyploid 313 taxa. In tetraploid cotton, for example, map443 ping of duplicate loci allowed identification of 11 of the 13 expected homeologous pairs 227 rp1 between the A and D subgenomes29. Two of nf73 these pairs are collinear; four differ by at least one inversion; two differ by at least two inversions; and at least one pair shows a Synthesized H. anomalus hybrids translocation. To determine whether the ex239 tensive chromosomal reorganization between Synthesized H. anomalus hybrids the A and D genomes occurred before or after 103 polyploidization, the genomes of the A and D diploid progenitors have also been mapped (C.L. Brubaker et al., unpublished). Comparison of the diploid and polyploid genH. petiolaris markers or S linkage blocks 295 omes revealed that the A and D genomes of the diploid species were most similar in A15 gene order, and the A and D subgenomes of H. annuus markers or the tetraploid were most divergent. This linkage blocks suggests that rates of structural evolution increased following polyploidization. Experimental study has also revealed evi40 cM dence for rapid genomic evolution following 471 6 polyploidization . The most detailed studies come from synthetic Brassica allotetraploids generated by interspecific hybridizations between ‘diploid’ Brassica spp.: B. rapa (A genome); B. nigra (B genome); and B. oleracea Synthesized H. anomalus hybrids (C genome). The crosses were: A ⫻ B; B ⫻ A; C ⫻ A; and A ⫻ C (Ref. 6). The AB and Fig. 3. Genomic composition of ancient and experimental hybrid lineages for three selected BA synthetic tetraploids have the same nulinkage groups. Letters at the left of each linkage group designate linkage blocks in Helianthus clear genome complement as the natural neoanomalus and indicate homology to linkages previously mapped in the parental species, tetraploid, B. juncea (n = 18), and the AC H. annuus and H. petiolaris5 (the broken line indicates the break-point between the R and and CA synthetic tetraploids have the same S blocks of that linkage group). The distribution of parental chromosomal blocks in the synthesized hybrids (left linkage group) and H. anomalus (right linkage group) is indicated by nuclear genome complement as B. napus colored bars. Regions harboring recombination points are indicated by a color scale, with (n = 19). RFLP comparisons of F2 and F5 the intensity of shading indicating the likelihood that a particular region was derived from plants revealed evidence for rapid genome one parent or the other. Modified from Ref. 5. change. On average, 9.6% of restriction fragments were dissimilar between F2 and F5 plants from the AB line, 8.2% from the BA line, 4.1% from the AC line, and 3.7% from the CA line. Notably, provide a source of genetic variation for adaptive evolution and dithe paternal subgenome in the AB and BA tetraploids showed versification within polyploid lineages. Additionally, if the kinds greater differentiation from its diploid parent than did the mater- of genomic changes observed here are arbitrary, it seems likely that nal subgenome, but no significant differences in subgenome evo- polyploid lineages independently derived from the same parental lution were observed in the AC and CA tetraploids. This suggests material can rapidly diverge and potentially achieve reproductive that cytoplasmic–nuclear interactions influenced genomic change isolation. This could be explored by testing the compatibility of in the AB and BA tetraploids, but not in the AC and CA tetra- polyploid populations that molecular evidence has shown to be ploids. Presumably, the lack of significant cytoplasmic effects in independently derived from the same parental species. the AC and CA tetraploids reflects higher levels of cytonuclear compatibility, because the A and C subgenomes are more closely Recombinational speciation related phylogenetically than are the A and B subgenomes. The most widely accepted model for diploid or homoploid hybrid As stressed by Song et al.6, these data have significant impli- speciation is the recombinational model described by Grant39. cations for our understanding of polyploid evolution. They demon- According to this model, the sorting of chromosomal rearrangestrate that much of the genomic change found in natural polyploids ments in later-generation hybrids could, by chance, lead to the probably accompanies polyploid formation. These changes possibly formation of new populations that are homozygous for a novel July 1998, Vol. 3, No. 7
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trends in plant science reviews In comparison to allopolyploidy, researchers have expended much less effort towards understanding the mechanistic basis of recombinational speciation. The only relevant studies concern the origin of a wild hybrid sunflower species, Helianthus anomalus5,42.
100
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0 0
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Fig. 4. Mean pollen fertility in three synthetic hybrid lineages between Helianthus annuus and H. petiolaris. Lineage pedigrees are as follows (generation number in parentheses): lineage I, P(0)F1(1)-BC1(2)-BC2(3)-F2(4)-F3(5); lineage II, P(0)-F1(1)-F2(2)BC1(3)-BC2(4)-F3(5); and lineage III, P(0)-F1(1)-F2(2)-F3(3)BC1(4)-BC2(5). Abbreviations: P, parental; F, sibmating; BC, backcrossing. All backcrosses were in the direction of H. annuus. For each line, 100 pollen grains from each of 20 plants per generation were tested by viability staining. Standard error bars are omitted for clarity.
combination of rearrangements. The new hybrid population would be fertile, stable and have the same ploidy as its parents, yet would be at least partially isolated from both parental species by a chromosomal sterility barrier. Although this model focused on chromosomal rearrangements, it is clear that the sorting of genic sterility factors could generate similar results. Thus, current models incorporate both genic and chromosomal sterility factors40. Other factors that appear to play a critical role in recombinational speciation include: strong natural selection for the most fertile or viable hybrid segregants5,41; rapid chromosomal evolution42; and the availability of habitats suitable for the establishment of hybrid neospecies40. The feasibility of the recombinational model has been explored experimentally via crossing studies40. These studies validated the recombinational model by demonstrating that fertile and viable hybrid lineages can be obtained after only a small number of generations (<10) of selfing and/or backcrossing, even if F1 hybrids were almost completely sterile. Furthermore, the experimentally generated hybrid lineages were often strongly reproductively isolated from the parental species. Estimating the frequency of homoploid hybrid speciation in nature is more difficult, and fewer than ten cases have been rigorously documented for plants40. However, these low numbers may be an artifact of the difficulty of detecting and documenting homoploid hybrid species, particularly if the hybridization events are ancient. A much larger number of hybrid species has been proposed, and molecular phylogenetic studies continually uncover unexpected cases of ancient hybridization, some of which may have led to speciation43. Also, hybridization may play an important role as the creative stimulus for speciation in small or peripheral populations. Hybridization rates appear to be highest in populations with these characteristics, and hybridization may be more plausible than population bottlenecks for generating the genetic or chromosomal reorganization proposed in founder-effect speciation40. 258
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Genetic models suggest that reproductive isolation between the hybrid neospecies and species populations can be facilitated by rapid karyotypic evolution39–41. This hypothesis has been tested by comparing genetic maps for H. anomalus and its putative parental species, H. annuus and H. petiolaris42. The mapping data revealed extensive chromosomal differentiation between the hybrid and parental genomes. Seven linkage groups in the hybrid genome differed in gene order from both parental species, and three chromosomal breakages, three fusions and one duplication are required to explain these differences. Chromosomal differentiation does appear to have facilitated reproductive isolation in H. anomalus; F1 hybrids with its putative parental species are partially sterile because of meiotic abnormalities42. The role of selection
To assess the relative contributions of deterministic and stochastic forces in the recombinational speciation process, three independent hybrid lineages were synthesized between H. annuus and H. petiolaris5. Comparison of the genomic composition of the ancient (H. anomalus) and synthetic hybrid lineages revealed that all three synthetic hybrid lineages had converged to nearly identical gene combinations, and this set of combinations was statistically concordant with that of H. anomalus (Fig. 3). Similarity in genomic composition between the synthetic and ancient hybrids suggests that deterministic forces such as selection, rather than stochastic forces, largely govern the formation of ‘recombinational’ species. Because the synthetic hybrid lineages were generated in the greenhouse, fertility selection probably played a greater role than ecological selection in shaping hybrid genomic composition. This conclusion is supported by the rapid increase in fertility observed in the three synthetic hybrid lineages; average pollen fertility increased from 4% in the F1 generation to >90% in the fifth-generation hybrids (Fig. 4). Congruence in genomic composition also implies that the genomic structure and composition of hybrid species is essentially fixed after a small number of generations of hybridization and remain relatively static thereafter. Tempo of recombinational speciation
Simulation studies suggest that recombinational speciation is punctuated: long periods of hybrid-zone stasis are followed by abrupt transitions in which parental individuals are displaced rapidly by the hybrid neospecies41. The hypothesis of rapid speciation was tested by analyzing the sizes of parental species chromosome blocks in the H. anomalus genome (M. Ungerer et al., unpublished). During the evolution of a new hybrid species, parental linkage block sizes are expected to become progressively smaller over time because of recombination. However, continued reductions in block size will be countered by structural fixation of the hybrid genome; subsequent recombination among blocks derived from the same parental species will no longer decrease block size. To estimate the number of generations of recombination required to achieve the present distribution of parental chromosomal block sizes in the H. anomalus genome, observed block sizes were compared with those of a computer-simulated hybrid population (M. Ungerer et al., unpublished). This comparison suggests that H. anomalus arose extremely rapidly, probably in fewer than 60 generations. This result is corroborated by the independent lines of evidence already discussed, which demonstrate
trends in plant science reviews significant concordance between the genomes of H. anomalus and early-generation synthetic hybrids (Fig. 3), and a rapid recovery of fertility in these synthetic hybrid lineages (Fig. 4). Conclusions
Understanding of how hybrid lineages are stabilized has advanced considerably over the past five years. Many of these advances are due to the application of recently developed genetic mapping technology. Indeed, reticulate modes of speciation are arguably far better understood than their divergent counterparts, such as geographical speciation. The rapid progress that has characterized the study of reticulate evolution is due in part to the application of genetic mapping tools, but has also benefited from allopolyploid and recombinational speciation being rapid modes of speciation that are easily studied experimentally. It seems likely that the combination of experimental genetic and historical approaches that have proven so powerful for the study of hybrid lineages will have a similar impact on studies of divergent modes of evolution. Acknowledgements
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Loren Rieseberg* and Rick Noyes are at the Dept of Biology, Indiana University, Bloomington, IN 47405, USA. *Author for correspondence (tel +1 812 855 7614; fax +1 812 855 6705; e-mail [email protected]).
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