- Email: [email protected]
What can we learn from natural apomicts?
trends in plant science research news
What can we learn from natural apomicts? Apomixis is an asexual breeding system in plants that allows fertile seeds to develop in the absence of fertilization. Apomicts produce progeny that are genetic copies of themselves, thus preserving the maternal genotype. Ever since its discovery1 in 1845, apomixis has been regarded as a rare and eccentric breeding system in angiosperms, but there has been much recent interest in apomixis because of its great potential for crop improvement2,3 and the possibility of changing sexual crops into apomicts4 via ‘apomixis technology’. In many sexual crops, hybrid varieties are superior, but because sexual hybrids segregate, they have to be renewed each generation by crossing the parental lines. In contrast, apomictic hybrids would breed true because meiosis and segregation are by-passed. Although apomixis does not naturally occur in today’s main crop species (and attempts to introgress apomixis into crops by interspecific hybridization with wild apomictic relatives have so far been Fig. 1. Apomixis in Taraxacum officinale. unsatisfactory), modern molecular techniques are expected to give better prospects for introducing apomixis into crops. The potential applications of ‘apomixis tech- evolution of this trait. A recent workshop* nology’ go far beyond classical apomixis was organized at the Netherlands Institute for introgression, where species barriers and dif- Ecological Research to discuss both the genferent ploidy levels usually prevented suc- etics of apomict formation and the genetical cessful transfer of apomixis from wild plants consequences of apomixis in a few model to cultivated relatives. If a sexual plant could species. The main contributions and discussions be transformed into an apomict by introduc- focused on comparable apomictic processes in ing an ‘apomixis cassette’, every superior different aposporous and diplosporous genera. multigenic genotype could be preserved and The aposporous genera, Ranunculus and Hierpropagated rapidly as a cultivar. acium, and the diplosporous genus Taraxacum, were discussed in greater detail. Genetic basis
Given the potential of apomixis for agriculture, it is unfortunate that little is known about the genetic basis of apomixis in established natural apomicts. However, it is known that the switch from sexual to apomictic seed production is largely characterized by two aspects: • Circumvention of meiotic reduction. • Parthenogenetic (autonomous) embryo development. Natural apomicts have solved the meiotic problem in different ways, either by substitution of meiosis by somatic mitosis (adventitious embryony and apospory), or by an aberrant meiosis, producing unreduced megaspores (diplospory). Insight into the genetics of natural apomixis and the type of genes involved would tell us whether transforming sexual crop species into apomicts is a real possibility. Understanding the genetic basis of apomixis is also essential for unravelling the
Single-gene model
Mike Mogie (University of Bath, UK) discussed a theoretical model for the genetic control of apomixis. In this single locus model, parthenogenesis was considered to be a pleiotropic effect of circumventing meiotic reduction. Mogie argued that some meiotic mutations in sexual organisms result in faster egg cell formation. Precocious oogenesis before anthesis prevents fertilization and could cause precocious embryony. From further analogies with known meiotic mutants, Mogie predicted that homozygosity for apomixis genes would be lethal and that the ratio of apomixis to wild-type genes that would generate a viable apomictic phenotype is most likely to occur in polyploids. The attractiveness of a *The Genetics of Apomixis, Netherlands Institute for Ecological Research, Heteren, The Netherlands, 22–23 August 1998.
1360 - 1385/99/$ – see front matter © 1999 Elsevier Science. All rights reserved.
single-gene model is that it bypasses the evolutionary problem posed by a twogene model. It would be difficult to bring together two different mutations, namely circumvention of meiosis and autonomous embryo development, because each mutation alone lowers fitness. Recessively lethal apomixis genes
Gian Nogler (University of Zürich, Switzerland) outlined the heredity of apomixis in the Ranunculus auricomus complex, based on what is probably the most comprehensive data set on the genetic inheritance of apomixis. The crossing scheme started with a diploid sexual mother and a tetraploid apomictic father and spanned five generations. Initial results in the first generation with small progeny sizes suggested that the control of apospory was recessive, but extended crosses and larger progenies unambiguously showed a monogenic control with dominance for apospory. In contrast, the genetic control of parthenogenesis was ambiguous. In some crosses between apomictic and sexual lines, parthenogenesis appeared to become uncoupled because unreduced eggsacs developed after fertilization. However, when pollination was delayed in these crosses, parthenogenetic embryo development was initiated. This indicates that early pollination can impose fertilization on unreduced eggsacs before parthenogenesis can be initiated. By contrast, diploids obtained in crosses between sexuals and apomicts were always sexual. From these results, Nogler proposed that apomixis genes are recessively lethal and cannot be transmitted through haploid gametes. This is a plausible explanation because apomixis is always associated with higher ploidy levels. The inheritance of apomixis in aposporous Hieracium subgenus Pilosella (Asteraceae) was comparable to Ranunculus. Hieracium is a facultative apomict that can have both sexual and asexual megagametogenesis in one ovule. Ross Bicknell (Crop and Food Research, Christchurch, New Zealand) argued that apomixis depended on a single dominant gene in Hieracium. Haploid gametes did not seem to transmit this apomixis gene, which Bicknell believes is due to deleterious alleles linked to the apomixis gene. Dihaploids recovered from apomicts were often apomicts, but had low fitness. Bicknell interpreted this as an indication for a large genetic load in apomictic Hieracium. This is consistent with evolutionary theories that predict an accumulation of deleterious mutations and chromosomal rearrangements in asexual lineages: it February 1999, Vol. 4, No. 2
43
trends in plant science research news could be that absence of apomixis in diploid plants is a consequence of the evolutionary history of apomixis genes, rather than direct lethality of apomixis genes in a diploid or haploid background. Although facultative apomixis might be controlled by a specific major apomixis gene(s), as in Hieracium, the degree of apomictic seed set can be highly variable. Anna Koltunow (CSIRO Plant Industry, Glen Osmond, Australia) presented an overview of this variability in Hieracium. Histological studies revealed that the number of aposporous embryo initials varied between apomictic dihaploids. Apomictic development of embryo sacs was stochastic and influenced by changes in the immediate ovule environment. Moreover, crosses between sexual and apomictic plants indicated the involvement of modifiers in addition to the apomixis locus. Genetic determination of diplospory
Although the genetics of aposporous systems are relatively well studied, the genetic determination of diplospory remains unclear. Peter van Dijk (Netherlands Institute for Oecological Research, Heteren, The Netherlands) reported on the heredity and gradual loss of apomixis in the common dandelion, Taraxacum officinale (Fig. 1). This plant is an obligate diplosporous apomict; one parthenogenetic embryo develops per ovule from an unreduced megaspore. Gradual loss of the two main components of apomixis, circumvention of meiotic reduction and autonomous embryo development, could be demonstrated in crosses between sexual diploids and apomictic tetraploids. Only one out of every nine triploid hybrids in these crosses was apomictic. This could be explained by recessive apomixis genes or by recombination between multiple dominant apomixis genes. The occurrence of recombination was inferred from the recovery of plants that make unreduced egg cells but lack parthenogenetic embryo development, indicating that diplospory and parthenogenesis are independent elements of apomixis. Dissecting the complex apomixis trait into its components could simplify its further genetic analysis. There is a general feeling among researchers that recombination is totally absent in apomicts. But obligate diplosporous apomicts (for instance Taraxacum) could produce genetically heterogeneous offspring by a process called autosegregation, as elaborated by Munikote Ramanna (Agricultural University, Wageningen, The Netherlands). In regular first division restitution mechanisms, crossing-over between a locus and centromere at first meiotic prophase would produce recombinant chromatids, and gene conversion would give rise to homozygosity. Recombinant alleles can segregate after the second 44
February 1999, Vol. 4, No. 2
meiotic division when the sister chromatids segregate. In 50% of the cases, recombinant alleles will end up in the meiotic product that will give rise to the embryo. One can speculate about the efficiency of this mechanism for creating recombinants and discarding deleterious mutations. Ramanna considers this so-called ‘pseudo-homeotypic division’ as a real, but generally overlooked aspect of meiotic diplospory. Autosegregation would lower heterozygosity and break down epistatic interactions. Codominant marker studies should be used to quantify autosegregation in diplosporous apomicts. Problems
During the meeting, it became clear that work on the genetics of natural apomixis is progressing steadily. Work is hampered by the female sexual reproductive function, which is (nearly) completely blocked and because apomicts are polyploids. Moreover, not every apomictic species is equally suited for genetic studies, due to, for example, higher ploidy levels, pseudogamy and incomplete penetrance of apomixis. An important established characteristic of both aposporic Hieracium and diplosporic Taraxacum, both Asteraceae, is that the ovule contains a protein-rich storage tissue, which nourishes the embryo, reducing the importance of endosperm function. This could explain why most of the apomicts in the Asteraceae show autonomous embryo development, a feature that is rare in other plant families. The Asteraceae could well have been pre-adapted for autonomous apomixis, because of the presence of this nutritional tissue, before the rise of apo- or diplo-spory. The presence of a somatic nutritional tissue provides a parsimonious way to achieve autonomous apomixis, because it also bypasses the general requirement of proper endosperm imprinting in plants5. The ability to feed the parthenogenetic embryo is a fundamental aspect of apomictic seed production. Most natural apomicts are pseudogamous: pollination is required for fertilization of the polar nuclei, a prerequisite for endosperm development. At least initially, it appears that future crops, transgenic for apomixis, will use a pseudogamous form of the trait. Pollen donors will therefore continue to be required for seed formation. However, transgenic apomixis does not only face technical problems. Transgenic plants might cross with wild relatives thereby passing on their modified information. Transfer of herbicide resistance from transgenic rape seed to a related weed without loss of parental vigour has been demonstrated6. One different type of problem is posed by the short-term evolutionary advantages of asexual reproduction. Suppose that the transgene ‘escapes’ from a field to its surrounding environment. Whereas transgenes now have to introgress
into a pre-existing gene pool to spread, often across partial reproductive barriers, a chimeric apomict could start a population by itself: there is no need to find a suitable partner. The evolutionary advantages will be even more pronounced when the ‘escaped’ transgenic apomict is aided by the selection pressure on other modified traits, for instance herbicide resistance. It is evident that transfer and spread of apomixis transgenes alone, or coupled to other transgenes, should become the subject of rigorous research. Conclusions
This meeting was very successful in synthesizing a conceptual model of genetic determination of apomixis. A clear indication of this success was the integration of classical and modern techniques in apomixis research, the prospects that were outlined and the questions that have been raised. Peter van Baarlen* Laboratory of Genetics, Wageningen Agricultural University, Dreyenlaan 2, 6703 HA Wageningen, The Netherlands Marije Verduijn and Peter J. van Dijk Netherlands Institute for Oecological Research, Boterhoeksestraat 22, 6666 ZG Heteren, The Netherlands
*Author for correspondence (Fax 131 317 483146; e-mail [email protected]) References 1 Smith, J. (1845) Notice of a plant which produces seeds without any apparent action of pollen, Trans. Linnean Soc. P18 2 Vielle-Calzada, J.P., Crane, C.F. and Stelly, D.M. (1996) Apomixis: the asexual revolution, Science 274, 1322–1323 3 Grossniklaus, U., Koltunow, A. and van Lookeren Campagne, M. (1998) A bright future for apomixis, Trends Plant Sci. 3, 415–416 4 Koltunov, A.M., Bicknell, R.A. and Chaudhury, A.M. (1995) Apomixis: molecular strategies for the generation of genetically identical seeds without fertilization, Plant Physiol. 108, 1345–1352 5 Haig, D. and Westoby, M. (1991) Genomic imprinting in endosperm: its effect on seed development in crosses between species, and between different ploidies of the same species, and its implications for the evolution of apomixis, Philos. Trans. R. Soc. London. Ser. B 333, 1–13 6 Dove, A., Marshall, A. and Raz, R. (1998) Promiscuous pollen, Nat. Biotechnol. 16, 805
What can we learn from natural apomicts? Apomixis is an asexual breeding system in plants that allows fertile seeds to develop in the absence of fertilization. Apomicts produce progeny that are genetic copies of themselves, thus preserving the maternal genotype. Ever since its discovery1 in 1845, apomixis has been regarded as a rare and eccentric breeding system in angiosperms, but there has been much recent interest in apomixis because of its great potential for crop improvement2,3 and the possibility of changing sexual crops into apomicts4 via ‘apomixis technology’. In many sexual crops, hybrid varieties are superior, but because sexual hybrids segregate, they have to be renewed each generation by crossing the parental lines. In contrast, apomictic hybrids would breed true because meiosis and segregation are by-passed. Although apomixis does not naturally occur in today’s main crop species (and attempts to introgress apomixis into crops by interspecific hybridization with wild apomictic relatives have so far been Fig. 1. Apomixis in Taraxacum officinale. unsatisfactory), modern molecular techniques are expected to give better prospects for introducing apomixis into crops. The potential applications of ‘apomixis tech- evolution of this trait. A recent workshop* nology’ go far beyond classical apomixis was organized at the Netherlands Institute for introgression, where species barriers and dif- Ecological Research to discuss both the genferent ploidy levels usually prevented suc- etics of apomict formation and the genetical cessful transfer of apomixis from wild plants consequences of apomixis in a few model to cultivated relatives. If a sexual plant could species. The main contributions and discussions be transformed into an apomict by introduc- focused on comparable apomictic processes in ing an ‘apomixis cassette’, every superior different aposporous and diplosporous genera. multigenic genotype could be preserved and The aposporous genera, Ranunculus and Hierpropagated rapidly as a cultivar. acium, and the diplosporous genus Taraxacum, were discussed in greater detail. Genetic basis
Given the potential of apomixis for agriculture, it is unfortunate that little is known about the genetic basis of apomixis in established natural apomicts. However, it is known that the switch from sexual to apomictic seed production is largely characterized by two aspects: • Circumvention of meiotic reduction. • Parthenogenetic (autonomous) embryo development. Natural apomicts have solved the meiotic problem in different ways, either by substitution of meiosis by somatic mitosis (adventitious embryony and apospory), or by an aberrant meiosis, producing unreduced megaspores (diplospory). Insight into the genetics of natural apomixis and the type of genes involved would tell us whether transforming sexual crop species into apomicts is a real possibility. Understanding the genetic basis of apomixis is also essential for unravelling the
Single-gene model
Mike Mogie (University of Bath, UK) discussed a theoretical model for the genetic control of apomixis. In this single locus model, parthenogenesis was considered to be a pleiotropic effect of circumventing meiotic reduction. Mogie argued that some meiotic mutations in sexual organisms result in faster egg cell formation. Precocious oogenesis before anthesis prevents fertilization and could cause precocious embryony. From further analogies with known meiotic mutants, Mogie predicted that homozygosity for apomixis genes would be lethal and that the ratio of apomixis to wild-type genes that would generate a viable apomictic phenotype is most likely to occur in polyploids. The attractiveness of a *The Genetics of Apomixis, Netherlands Institute for Ecological Research, Heteren, The Netherlands, 22–23 August 1998.
1360 - 1385/99/$ – see front matter © 1999 Elsevier Science. All rights reserved.
single-gene model is that it bypasses the evolutionary problem posed by a twogene model. It would be difficult to bring together two different mutations, namely circumvention of meiosis and autonomous embryo development, because each mutation alone lowers fitness. Recessively lethal apomixis genes
Gian Nogler (University of Zürich, Switzerland) outlined the heredity of apomixis in the Ranunculus auricomus complex, based on what is probably the most comprehensive data set on the genetic inheritance of apomixis. The crossing scheme started with a diploid sexual mother and a tetraploid apomictic father and spanned five generations. Initial results in the first generation with small progeny sizes suggested that the control of apospory was recessive, but extended crosses and larger progenies unambiguously showed a monogenic control with dominance for apospory. In contrast, the genetic control of parthenogenesis was ambiguous. In some crosses between apomictic and sexual lines, parthenogenesis appeared to become uncoupled because unreduced eggsacs developed after fertilization. However, when pollination was delayed in these crosses, parthenogenetic embryo development was initiated. This indicates that early pollination can impose fertilization on unreduced eggsacs before parthenogenesis can be initiated. By contrast, diploids obtained in crosses between sexuals and apomicts were always sexual. From these results, Nogler proposed that apomixis genes are recessively lethal and cannot be transmitted through haploid gametes. This is a plausible explanation because apomixis is always associated with higher ploidy levels. The inheritance of apomixis in aposporous Hieracium subgenus Pilosella (Asteraceae) was comparable to Ranunculus. Hieracium is a facultative apomict that can have both sexual and asexual megagametogenesis in one ovule. Ross Bicknell (Crop and Food Research, Christchurch, New Zealand) argued that apomixis depended on a single dominant gene in Hieracium. Haploid gametes did not seem to transmit this apomixis gene, which Bicknell believes is due to deleterious alleles linked to the apomixis gene. Dihaploids recovered from apomicts were often apomicts, but had low fitness. Bicknell interpreted this as an indication for a large genetic load in apomictic Hieracium. This is consistent with evolutionary theories that predict an accumulation of deleterious mutations and chromosomal rearrangements in asexual lineages: it February 1999, Vol. 4, No. 2
43
trends in plant science research news could be that absence of apomixis in diploid plants is a consequence of the evolutionary history of apomixis genes, rather than direct lethality of apomixis genes in a diploid or haploid background. Although facultative apomixis might be controlled by a specific major apomixis gene(s), as in Hieracium, the degree of apomictic seed set can be highly variable. Anna Koltunow (CSIRO Plant Industry, Glen Osmond, Australia) presented an overview of this variability in Hieracium. Histological studies revealed that the number of aposporous embryo initials varied between apomictic dihaploids. Apomictic development of embryo sacs was stochastic and influenced by changes in the immediate ovule environment. Moreover, crosses between sexual and apomictic plants indicated the involvement of modifiers in addition to the apomixis locus. Genetic determination of diplospory
Although the genetics of aposporous systems are relatively well studied, the genetic determination of diplospory remains unclear. Peter van Dijk (Netherlands Institute for Oecological Research, Heteren, The Netherlands) reported on the heredity and gradual loss of apomixis in the common dandelion, Taraxacum officinale (Fig. 1). This plant is an obligate diplosporous apomict; one parthenogenetic embryo develops per ovule from an unreduced megaspore. Gradual loss of the two main components of apomixis, circumvention of meiotic reduction and autonomous embryo development, could be demonstrated in crosses between sexual diploids and apomictic tetraploids. Only one out of every nine triploid hybrids in these crosses was apomictic. This could be explained by recessive apomixis genes or by recombination between multiple dominant apomixis genes. The occurrence of recombination was inferred from the recovery of plants that make unreduced egg cells but lack parthenogenetic embryo development, indicating that diplospory and parthenogenesis are independent elements of apomixis. Dissecting the complex apomixis trait into its components could simplify its further genetic analysis. There is a general feeling among researchers that recombination is totally absent in apomicts. But obligate diplosporous apomicts (for instance Taraxacum) could produce genetically heterogeneous offspring by a process called autosegregation, as elaborated by Munikote Ramanna (Agricultural University, Wageningen, The Netherlands). In regular first division restitution mechanisms, crossing-over between a locus and centromere at first meiotic prophase would produce recombinant chromatids, and gene conversion would give rise to homozygosity. Recombinant alleles can segregate after the second 44
February 1999, Vol. 4, No. 2
meiotic division when the sister chromatids segregate. In 50% of the cases, recombinant alleles will end up in the meiotic product that will give rise to the embryo. One can speculate about the efficiency of this mechanism for creating recombinants and discarding deleterious mutations. Ramanna considers this so-called ‘pseudo-homeotypic division’ as a real, but generally overlooked aspect of meiotic diplospory. Autosegregation would lower heterozygosity and break down epistatic interactions. Codominant marker studies should be used to quantify autosegregation in diplosporous apomicts. Problems
During the meeting, it became clear that work on the genetics of natural apomixis is progressing steadily. Work is hampered by the female sexual reproductive function, which is (nearly) completely blocked and because apomicts are polyploids. Moreover, not every apomictic species is equally suited for genetic studies, due to, for example, higher ploidy levels, pseudogamy and incomplete penetrance of apomixis. An important established characteristic of both aposporic Hieracium and diplosporic Taraxacum, both Asteraceae, is that the ovule contains a protein-rich storage tissue, which nourishes the embryo, reducing the importance of endosperm function. This could explain why most of the apomicts in the Asteraceae show autonomous embryo development, a feature that is rare in other plant families. The Asteraceae could well have been pre-adapted for autonomous apomixis, because of the presence of this nutritional tissue, before the rise of apo- or diplo-spory. The presence of a somatic nutritional tissue provides a parsimonious way to achieve autonomous apomixis, because it also bypasses the general requirement of proper endosperm imprinting in plants5. The ability to feed the parthenogenetic embryo is a fundamental aspect of apomictic seed production. Most natural apomicts are pseudogamous: pollination is required for fertilization of the polar nuclei, a prerequisite for endosperm development. At least initially, it appears that future crops, transgenic for apomixis, will use a pseudogamous form of the trait. Pollen donors will therefore continue to be required for seed formation. However, transgenic apomixis does not only face technical problems. Transgenic plants might cross with wild relatives thereby passing on their modified information. Transfer of herbicide resistance from transgenic rape seed to a related weed without loss of parental vigour has been demonstrated6. One different type of problem is posed by the short-term evolutionary advantages of asexual reproduction. Suppose that the transgene ‘escapes’ from a field to its surrounding environment. Whereas transgenes now have to introgress
into a pre-existing gene pool to spread, often across partial reproductive barriers, a chimeric apomict could start a population by itself: there is no need to find a suitable partner. The evolutionary advantages will be even more pronounced when the ‘escaped’ transgenic apomict is aided by the selection pressure on other modified traits, for instance herbicide resistance. It is evident that transfer and spread of apomixis transgenes alone, or coupled to other transgenes, should become the subject of rigorous research. Conclusions
This meeting was very successful in synthesizing a conceptual model of genetic determination of apomixis. A clear indication of this success was the integration of classical and modern techniques in apomixis research, the prospects that were outlined and the questions that have been raised. Peter van Baarlen* Laboratory of Genetics, Wageningen Agricultural University, Dreyenlaan 2, 6703 HA Wageningen, The Netherlands Marije Verduijn and Peter J. van Dijk Netherlands Institute for Oecological Research, Boterhoeksestraat 22, 6666 ZG Heteren, The Netherlands
*Author for correspondence (Fax 131 317 483146; e-mail [email protected]) References 1 Smith, J. (1845) Notice of a plant which produces seeds without any apparent action of pollen, Trans. Linnean Soc. P18 2 Vielle-Calzada, J.P., Crane, C.F. and Stelly, D.M. (1996) Apomixis: the asexual revolution, Science 274, 1322–1323 3 Grossniklaus, U., Koltunow, A. and van Lookeren Campagne, M. (1998) A bright future for apomixis, Trends Plant Sci. 3, 415–416 4 Koltunov, A.M., Bicknell, R.A. and Chaudhury, A.M. (1995) Apomixis: molecular strategies for the generation of genetically identical seeds without fertilization, Plant Physiol. 108, 1345–1352 5 Haig, D. and Westoby, M. (1991) Genomic imprinting in endosperm: its effect on seed development in crosses between species, and between different ploidies of the same species, and its implications for the evolution of apomixis, Philos. Trans. R. Soc. London. Ser. B 333, 1–13 6 Dove, A., Marshall, A. and Raz, R. (1998) Promiscuous pollen, Nat. Biotechnol. 16, 805