A new kind of plant breeding February 2022

By Rebecca Guenard

In This Section

February 2022

  • Researchers can breed traits into plants without changing their DNA sequence.
  • The epigenome is a set of molecules that swarm the genome and determine the activity of genes based on environmental influences.
  • If researchers can identify and select for epigenetic traits with long-term heritability, they could endow agricultural crops with the resilience to withstand an unpredictable environment caused by climate change.
  • Two start-ups have already acquired investments and hope to capitalize on this technology.

Armies of caterpillars and worms will annihilate unprotected corn crops, but proteins from a naturally occurring bacterium kill the pests. For decades, the bacterium, known as Bacillus thuringiensis, or Bt for short, has been a reliable organic pesticide spray. However, as a spray application, Bt faces challenges. Its proteins decompose in the sun and wash away in the rain, which led researchers to invent a more reliable way to use the bacterium to protect corn.

In the mid-1990s, scientists genetically engineered the DNA of corn plants to include genes from Bt. The genetically modified plant had built-in protection from caterpillars and other insect larva. The crop was so economically and environmentally successful that most of corn grown in the United States is Bt-corn. According to the US Food and Drug Administration, in 2018, 92% of corn planted in the United States was genetically modified.

In the decades since the development of genetically modified crops, consumer and environmental groups have voiced concerns about their use. Editing techniques have evolved from inserting foreign DNA with large altered sequences into a plant’s genome to much more precise modifications that alter just a few DNA letters without the need to transplant genes from another species. Now researchers are considering whether they can develop desirable traits in crops without changing even one letter in the code.

“Classical genetics tools alone cannot help us solve complex problems, like climate change,” says Dragana Miladinović, a plant biologist at the Institute of Field and Vegetable Crops in Novi Sad, Serbia. “Researchers observed that a plant’s response to environmental stresses could be controlled using more modern tools.”

A DNA sequence functions beyond its collection of genes. Whether genes operate or remain dormant depends on the molecules surrounding them. Researchers see potential in breeding desired traits into plants by manipulating these molecular switches that turn genes off or on while leaving a DNA sequence unaltered (Fig. 1).

FIG. 1. Physical and environmental threats to crops leave a memory in the plant through changes in the epigenome. Researchers are taking control of these changes to breed more climate-tolerant crops.  Source: Mladenov, V., et al., Int. J. Mol. Sci., 22, 13, 7118, 2021.
FIG. 1. Physical and environmental threats to crops leave a memory in the plant through changes in the epigenome. Researchers are taking control of these changes to breed more climate-tolerant crops. Source: Mladenov, V., et al., Int. J. Mol. Sci., 22, 13, 7118, 2021.

Epigenetics

Heredity is not determined solely by the genes coded in DNA. The molecular environment around a gene can also be passed down to offspring as cells multiply through mitosis or meiosis. Researchers currently label this type of heredity, epigenetics.

Pull DNA from any cell within an organism, and they will all contain the exact same genetic code. The instructions to make a complete organism is contained in every cell, but each cell has a unique job within an organism. Epigenetics gives cells their individuality. When a group of cells form the stalk of a plant, for example, the genes that control flowering are silenced within the DNA of those stalk cells, since flowering is meant to occur elsewhere.

A gene is silenced or activated depending on whether transcription molecules can access that stretch of DNA to read it. The ways parts of the code are blocked or exposed varies. A gene may be silenced when a series of enzymes create opposing charges that lead stretches of DNA to coil tightly around protein spools, called histones. By contrast, loosely wound DNA means an active gene. Acetyl groups astride a gene cause the DNA to be repelled from the histone because of similar charges. Whereas proteins or methyl groups restrict access to genes by physically surrounding them so they cannot be read.

The variety of modifications available to tweak the genetic code gives an organism the versatility to adjust to inputs from the environment. A plant that survives drought could potentially pass along methylation patterns that increase drought tolerance in its offspring. Unlike typical patterns of inheritance, such epigenetic states are fluid. The activity of a gene can be influenced by the experience of a parent and passed on to later generations.

Researchers are in the process of categorizing the effects of specific instances of altered epigenetic states in plants and how they contribute to cellular function. Once they understand a plant’s epigenetics they can use it to breed in traits that have commercial benefits, like increased protein or fatty acid content. Scientists are also hoping this type of breeding will contribute to a sustainable food supply by making plants more robust to changes in climate.

“Agriculture is facing more and more problems. Crops need to produce more with less input from humans, due to regulatory and consumer constraints,” says Miladinović. She is part of a collaborative, multidisciplinary group of scientists from 22 countries gathering and sharing findings on plant gene expression to solve the problem of making future food production adaptable to climate change.

“The application of epigenetics is no different than applying genetics in breeding, you are just using a different mechanism,” she says.

Artificial epigenetics

After observing the methylation changes that occur naturally in plants, researchers were interested in the effects of inducing epigenetic changes in the laboratory. They have since determined that epigenetic alterations can be performed in a number of ways, including abiotic stress, tissue culture, RNA-based methods, and grafting. The following paragraphs describe some of their preliminary research efforts.

Abiotic stress in plants generally includes drought, heat, cold, and salinity. The exact mechanism for how abiotic stress affects the epigenome is not yet completely clear. Experiments show that even the application of phytohormones associated with these kinds of stresses modify the plant epigenome and lead to lasting variation. Collectively, studies show that plants do not exhibit the same response to a particular stress. Each plant responds with different methylation effects. More research is needed to identify the genome-wide epigenetic changes that result from exposure to abiotic stress or the phytohormones it causes in order to determine its most beneficial manipulation (Table 1).

TABLE. 1. A few examples of epigenetic mechanisms involved in crop response to different abiotic stress.  Source: Varotto, S., et al., J Exp Bot, 71, 17, 5223–5236, 2020.
TABLE. 1. A few examples of epigenetic mechanisms involved in crop response to different abiotic stress. Source: Varotto, S., et al., J Exp Bot, 71, 17, 5223–5236, 2020.

Plant regeneration and propagation were once believed to produce a plant identical to the donor; however, researchers discovered that during tissue culture cells could erase some of their epigenetic markers. They observed that the hormones used in the growing medium can lead to a regenerated plant with unique epigenetic profiles. A complete assessment of these changes still needs to be explored.

Bacterial and fungal pathogens may also affect a plant’s epigenome. Pathogens transfer RNA to their host to silence the host’s defense genes. It is possible that pathogen RNA could also disrupt the host’s DNA methylation, but so far no one has reported this type of experimentally induced epigenetic alteration in plants.

Grafting the shoot of one plant onto another rooted plant originated as a means of harnessing the robustness of that plant and passing it on to the shoot. In grafting studies on a variety of plant species, researchers have determined that the technique also alters DNA methylation of the shoot. The graft-induced epigenetic changes remain intact even after pollination and the formation of progeny. Some researchers see grafting as the optimal way to manipulate the epigenome of a crop.

Of course, scientists have methods to directly alter the places within the genome where the epigenetic instructions are written. In contrast to the methods mentioned above, which affect the global genome, gene-specific insertions and deletions that control DNA methylation or histone interaction would be a more precise way to conduct epigenetic breeding. Although potentially effective, such human-guided gene-editing methods are not accepted in many countries.

Finally, researchers have experimented with externally applying RNA molecules to instruct specific genes to make epigenetic changes. They found that spraying foliage or soaking roots with RNA that promotes methylation triggered an increase in DNA methylation. More research is needed to improve RNA delivery, but the researchers are hopeful this type of fine-tuned epigenetic breeding would fall outside of current gene-editing regulations.

Lasting change

The range of examples given above show the complexity of options available when considering how best to use epigenetics in agriculture. For instance, some chemicals applied to seedlings are taken up by cells and incorporated into their DNA as they replicate. However, the epigenome is typically restored to its original state when chemical treatment is removed, rendering this method impractical for crop applications. To harness the epigenome, researchers must identify changes that transmit across generations.

In experiments performed on the model plant species Arabidopsis, they found that the plant carried an epigenetic memory from its ancestors. Over three generations, in accordance with a moderate ambient temperature increase, the plant produced less of an RNA involved in silencing a gene. After three generations, the effect’s strength declined. The researchers observed more enduring epigenetic memory associated with pathogen or UV light-induced stress. Such abiotic stressors may even induce heritable changes leading to more environmentally robust phenotypes.

Taking epigenetic knowledge from the laboratory to the field is the next step in realizing the potential of this breeding strategy. The epigenomes of critical staple crops like rice, corn, wheat, and barely have been picked over in recent years to identify epigenetic memory markers on stress-responsive genes that could be useful. AOCS relevant crops that have been investigated include rapeseed and, to a lesser extent, soybeans.

Since rapeseed is a recently domesticated hybrid crop, it has yet to develop extensive genetic diversity. Less diversity in rapeseed DNA provides an opportunity for greater influence from epigenetic changes. Studies on DNA methylation of rapeseed genes indicate a sensitivity to heat and salinity. Overall, it seems that rapeseed genotypes with less methylation also have higher stress tolerance. Hence, crop varieties with this epigenetic profile could be agriculturally more resilient.

Researchers identified the epigenetic component that determines how efficiently a rapeseed plant uses energy. The way the plant uses energy predicts its vigor and yield. Artificial selection of the plants with this desired epigenetic trait became heritable upon self-fertilization. Furthermore, hybrids from parental lines selected for high energy use efficiencies had a 5% yield increase.

“According to data available so far, recurrent selection seems to be the best tool for introducing epigenetic traits in crops,” says Miladinović. She cautions that, aside from rapeseed, most of this research has been carried out on model plants and not yet applied to crops. “As the genome becomes more complex, it is more difficult to control some things,” she says. This challenge has not deterred a few entrepreneurs.

Commercializing epigenetic breeding

According to Forbes, the overall epigenetics market will grow to $35 billion by 2028 (https://tinyurl.com/44d32hfr). Most of that value will likely come from diagnostic and treatment products for human health, but crop enhancement companies are also gaining traction.

This year, a start-up company called Sound Agriculture, established in Emeryville, California in 2013, announced it had secured $45 million in funding that it will use to support two new platforms for creating climate-resilient crops (https://tinyurl.com/8z5abt8j). The first focuses on adjusting a soil’s microbiome to increase the nutrient uptake efficiency of a crop. The second is aimed at developing specific heritable traits through altered plant epigenetics.

On-demand breeding, as it is called, accelerates plant trait development 10 times faster than gene editing, according to the company’s website (https://www.sound.ag/). The company hopes to identify and optimize traits that make plants more resilient with respect to climate change, diseases, and reduced chemical use. They are also considering traits that optimize nutrition, appearance, and flavor. Sound claims that breeders can progress from concept to a plantlet with a new trait in 15 days, compared to the 150 days needed for gene editing.

A similar company was founded by plant biologist Sally Mackenzie, currently at Pennsylvania State University in University Park, Pennsylvania, USA. Mackenzie discovered that a plant protein, known as MSH1, encodes for processes that determine DNA binding and recombination suppression (Fig.2). She argues that evidence indicates MSH1 is environmentally adaptive, so breeders can direct how a plant adjusts to its environment by targeting methylation on the gene that encodes MSH1 (http://dx.doi.org/10.1098/rstb.2019.0182).

Epicrop Technologies was established in 2013 in Lincoln, Nebraska, where Mackenzie was previously a professor at the University of Nebraska-Lincoln. After receiving funding from TechAccel, the company launched platforms to use epigenetics to improve two crops: strawberries and canola. They hope to improve the strawberry’s disease resistance and environmental range, while their research efforts for canola focus on yield.

FIG. 2. a. Seedlings from grafted Arabidopsis plants comparing a control graft (left) with a graft from a plant containing a mutation on the msh1 gene. Research shows mutations on this gene trigger epigenetic reprogramming in the plant. b-e. Examples of greater vigor and seed yield for msh1 mutated tomato plants over multiple generations in different field locations. Source: Kundariya, H., et al., Nat. Comm., 11, 5343, 2020.
FIG. 2. a. Seedlings from grafted Arabidopsis plants comparing a control graft (left) with a graft from a plant containing a mutation on the msh1 gene. Research shows mutations on this gene trigger epigenetic reprogramming in the plant. b-e. Examples of greater vigor and seed yield for msh1 mutated tomato plants over multiple generations in different field locations. Source: Kundariya, H., et al., Nat. Comm., 11, 5343, 2020.

The future of epigenomic breeding has many exciting prospects, according to Miladinović. The involvement of a variety of scientists using a range of tools means a quicker pace of discovery. A combination of epigenetics, genomics, and other omics tools with high-throughput analytics tools and artificial intelligence allows more data to be gathered and evaluated faster.

Miladinović says that since the concept for epigenetic breeding is still unproven in some crops, there is no way to know if desired traits will be heritable over many generations of a plant. “We still need to go case by case, crop by crop, and trait by trait,” she says. Even with classical breeding there are some traits that are easier to maintain than others. She says researchers will have to be careful to first identify traits with potential for heritable change.

One limitation of epigenetics is made obvious by our earlier example of Bt-corn. Scientists cannot add or remove any information from a plant’s genetic code. This means that incorporating the gene from a natural pesticide into a crop plant is off-limits in the epigenetic breeding playbook. If a gene responsible for some self-protective measure to ward off pests is not already present in the genome, there is no epigenetic course of action to address such a threat.

Another issue is that a plant epigenetically bred to resist heat may be left vulnerable if exposed to flooding. Miladinović says epigenetics is not going to produce crop varieties that can be planted anywhere. More likely, the discipline will assist with the new trend of agroecology, where crops are specialized for specific regions. “Farmers will not grow the same variety for different countries around the world, like they do now with seeds from a big corporation,” she says. Instead, farmers will select crops with one or two traits best suited for their agroecological area.

“We have discovered something beyond the classical genetics alphabet,” says Miladinović. “We have the potential to create a plant genotype that is really adaptive to a certain climate or a certain soil. This new breeding tool opens new ways to explore genetic diversity so we can assure stability of crops production as it faces environmental change.”

About the Author

Rebecca Guenard is the associate editor of Inform at AOCS. She can be contacted at rebecca.guenard@aocs.org

Information

Deciphering the epigenetic alphabet involved in transgenerational stress memory in crops, Mladenov, V., et al., Int. J. Mol. Sci., 22, 13, 7118, 2021.

Epigenetic approaches to crop breeding: current status and perspectives, Dalakouras, A. and Vlachostergios, D., J. Exp. Bot., 72, 15, 5356–5371, 2021.

MSH1-induced heritable enhanced growth vigor through grafting is associated with the RdDM pathway in plants, Kundariya, H., et al., Nat. Comm., 11, 5343, 2020.

Epigenetics: possible applications in climate-smart crop breeding, Varotto, S., et al., J. Exp. Bot., 71, 17, 5223–5236, 2020.

She has her mother’s laugh: The power, perversions, and potential of heredity by Carl Zimmer, Penguin Random House, LLC, 2018.

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