Binucleation to breed new plant species adaptable to their environments

Abstract

Classical plant breeding approaches may fall short to breed new plant species of high environmental and ecological interests. Biotechnological and genetic manipulations, on the other hand, may hold more effective capabilities to circumvent the limitations of sexual incompatibility and conventional breeding programs. Given that plant cells encompass multiple copies of organellar genomes (mitochondrial and plastidial genomes), an important question could be raised about whether an artificial attempt to duplicate the nuclear genome might also be conceivable through a binucleation approach (generating plant cells with 2 nuclei from 2 different plant species) for potential production of new polyploidies that would characterize new plant species. Since the complexities of plant genomes are the result of multiple genome duplications, an artificial binucleation approach would thus be of some interest to eventually varying plant genomes and producing new polyploidy from related or distal plant species. Here, I discuss the potentiality of such an approach to engineer binucleated plant cells as a germ of new plant species to fulfill some environmental applications such as increasing the biodiversity and breeding new species adaptable to harsh environmental stresses and increasing green surfaces to reduce atmospheric pollutions in arid lands with poor vegetation.

Keywords:

• binucleation
• binucleated cells
• biotic and abiotic stresses
• biofuel production
• environmental stresses
• genetic engineering
• genome fusion
• genome duplication
• molecular breeding
• new plant species
• nuclear transfer
• plant breeding techniques
• plant stress tolerance
• land sea hybrid species
• plant breeding
• seawater irrigation

The growing human population, the depletion of natural resources and the potential penury of foods in upcoming years are highly stimulating factors to experiment new researches hypothesis toward breeding new plant species adaptable to their fluctuating environmental conditions and to fulfill important ecological applications. Breeding hybrid species from related or distant plants species growing in opposite ecological niches (for e.g. in cold and hot, or sea and land habitats) would be of a particular interest to create new intermediate species for temperate regions or arid lands with new potential valuable traits. Creating such species would allow solving major and challenging environmental issues, particularly to provide new source for animal and human feeding, to produce green biofuels and to protect soils against erosion while reducing atmospheric pollution in regions with poor vegetation. To attain such objectives, classical breeding approaches would be challenged by sexual incompatibility and sterility barriers between distant species and genera. To circumvent these hurdles, we would need to investigate innovative methods or new ideas, predominantly those based on genetic and biotechnological tools. Previously, I have suggested some potential methods toward breeding new plant species irrigable with seawater using for example somatic hybridization, artificially-stimulated genome fusion, and nuclear transfer between land and sea plant species.¹ Here, I'd discuss another potential approach that might be of a particular interest to breed new intermediate plant species from related or distant species.

The potential method would be based on a binucleation assay by combining 2 nuclei from 2 different plant species in one plant cell to produce binucleated plant cells with new potential polyploidy and diverse genetic information. Experiments could thus be designed to transfer the nucleus from one plant species growing in a given ecological niches to a plant cell from another plant species growing in another environmental condition. Plants from opposite environmental conditions, for example northern cold hemisphere and hot tropics or plant species from land and sea habitats, could be attempted to breed new intermediate species with new fitness characteristics. The nucleus from one species (donor) should thus be isolated and reintroduced or injected into the cell of another plant species (receiver or host plant), without any exogenous or heterogeneous vectors, so the resulting genotypes cannot be tagged as transgenic, and then follow up the fate of the potentially resulting bi-nucleated cells after stimulating them to divide and grow. Beyond theoretical and experimental complexities of such genetic manipulations, combining 2 nuclear genomes in one plant cell would hypothetically duplicate and vary the genetic information within the new cell, and would characterize completely new plant species with atypical genetic material.

A solid rationale behind such an approach already exists in higher plants, where plant cells contain multiples copies of mitochondria and chloroplasts, each with their own genomes. Based on the similarity of these organelles to free living prokaryotes, it was suggested that the chloroplasts and mitochondria originated from prokaryotic cells through an endosymbiosis process during long evolutionary events (for review see ref.²) This endosymbiotic theory and the presence of multiple copies of mitochondria and plastids in plant cells strongly suggest the ability of plant cells to integrate new entire organelles with their own genomes that would confer new characteristics and fulfill some important functions (i.e. chloroplasts are involved in photosynthesis and mitochondria in respiration). In other word, if plant cells already contain multiple copies of organelles' genomes, we could also expect/test whether 2 nuclear genomes might also be conceivable in a plant cell to confer new characteristics encoded by the new ‘imported’ nucleus.

Another argument in favor of potential binucleation approach to produce new species comes from some organisms whose cells contain 2 nuclei. For example, in Giardia lambia (a medically important flagellated protozoan parasite³ it was found that 2 nuclei have complete copies of the genome, and both nuclei are transcriptionally active and replicate nearly at the same time^4,5 In human, it was found that the nondisjunction of chromosomes results in a tetraploid genotype in bi-nucleated cell lines⁶). To explore the potential consequences of binucleation on cell division, the authors compared the fates of mono-nucleated cells with bi-nucleated cells and found that 40% of the binucleated cells divide.⁶ If a similar or even less percentage of binucleated cells can be obtained from plants, there would be a great chance that new plant species could be obtained from binucleated cells since plant cells are totipotent and can develop into entire plants (for review, see⁷) Although bi-nucleation would be associated with cancer in human,⁶ binucleation in plant cells, on the other hand, would be an advantageous approach if it induces cell proliferation since this would represent a new form of reproduction of new prospected species.

A third and important argument for an artificial binucleation in plant is that, the genomes' complexity of flowering plants would be explained by genome duplication, rearrangements, crossing-over and cross-hybridizations. Genome-doubling events would have contributed to an important increase in the number of species in several angiosperm families, such as Brassicaceae, Fabaceae, Poaceae and Solanaceae.⁸ While the ancestral genome size of major clades of angiosperms and gymnosperms seems to be small,⁹, the increases of genome sizes in crops would have occurred naturally and slowly over long time by genetic duplication and polyploidy.¹⁰ Such duplications may significantly modify the plant's genetic make-up and its morphology within a few generations and allow some polyploids to colonize, and adapt to, new habitats and fluctuating conditions.¹¹ If genome's complexities have thus occurred naturally by iterative duplications/polyploidies during long time, we should wonder if an artificial genome transfer from one species to another through a binucleation approach, as described here, would produce new genetic diversifications in a shorter time than what happened naturally, over millions of years.

We thus could try to ‘imitate nature’ in doubling/varying genomes by producing cells with 2 nuclei. Theoretically, if the genomes of 2 nuclei would express, fully, partially or co-dominantly, they would generate new traits that should characterize new plant species with novel karyotype. In the same objective as previously described,¹ one nucleus from a sea plant species can be isolated and reintroduced into a land plant species, or inversely, in an attempt to potentially create new intermediate polyploidy land-sea hybrid plant species potentially irrigable with seawater.

However, if we try to transform a plant cell with a new nucleus (an entire genome), desirable or undesirable outcomes would be expected. First, nothing would happen (complete failure!), which would be unlikely because plant cells offer a great flexibility in term of totipotence.⁷ Moreover, the relative absence of ethical issues in manipulating plant genomes (compared to animals' genome manipulations) represents a major advantage for trying/retrying till a worthwhile outcome is achieved. Failure, however, is not uncommon in the scientific realm and success may be obtained only after multiple unsuccessful attempts. A second possibility of events upon a binucleation approach is that each nuclear genome may remain independent (likewise the organellar genomes) and would multiply (divide) independently within the same cell, as is the case in Giardia lambia.⁴ If this would happen successfully, a new species should be expected with a complex genetic program encoded by 2 co-existing nuclei whose the genetic makeup needs to be dissected a posteriori to assess how the 2 genomes behave in term of duplication, inheritance, dominance, co-dominance, cell division and karyotype, etc. A third and more complex pattern of binucleation is that potential interactions, loss or exchange of genetic materials may happen partially, irregularly, or randomly between the chromosomes of 2 neighboring nuclei, though usually only homologous chromosomes within each genome would in principle be involved in the crossing-over. If other than homologous chromosomes would cross-over, then, new genetic or genotypes would be expected and need to be explored closely. Another potential limitation is that, the new nucleus would be rejected or non-recognized by the host cell, so no effect would result.

Simultaneous co-existing/co-expressing of 2 nuclear genomes within a same plant cell would hypothetically reflect the sum of, or the interactions between, the genetic information encoded by the 2 coexisting nuclear genomes, and potentially producing unprecedented plant species. This is a hypothetic complex situation of coexisting genomes that, however, would deserve investigation in plants. One also should assume that unexpected outcomes may happen, such as the loss of chromosomes, the instability of genome or simply the inviability of bi-nucleated plant cells. In successfully-generated binucleated cells, a new geno-pheno-type should also be expectable.

Decades ago, genetic engineering and transformation methods have been started with the manipulation of merely one single gene. Later on, it was possible to transform cells with multiple genes at a time.^12,13 The sequencing of the human genome, and more generally the genomes of many living species, were unthinkable years ago. Now, the human's genome and an increasing number of other genomes are fully sequenced. From the same angle, complex and inconceivable approaches today might be realized in the future with the progress of science tool and knowledge. We might need to experiment genetic transformation with an entire nucleus (or complete genomes) at least in vitro studies on some model organisms, by transferring one nucleus from one plant species to another, and then observe the outcome, especially no restrictive ethical issues in plants would prevent such experimentations. New plant species with any shape, color or morphological characteristics that might result from such manipulations would not matter much. No foreign genetic material should be used in the transfer of nucleus between 2 cells but direct ‘nuclear injection’. Nucleus should be transferred from one cell into the cytoplasm of another cell, so no concerns about GMO risks. In animal research, on the other hand, the outcome of such genetic manipulations would be highly questionable from an ethical viewpoint as we cannot expect which outcome would be obtained from a binucleation approach.

Finally, since the nucleus is the cornerstone of any genetic and functional programs in living cells, transferring the nucleus from one species to another in an attempt to build new genetic blueprints would deserve the trial toward breeding new plant species. Instead of using pollen and classical breeding approaches between related species, a binucleation approach would be a potential and worthy method to consider between distant species and genera. Any probable technical and experimental limitations would be solved and optimized progressively, along with experimentation, testing and retesting, and all potential unanswered questions might be properly addressed once an experimental work is performed.

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.