Harnessing the Potential of Genome Editing for Tomorrow’s Agriculture

The fourth day of the Plant Biology Worldwide submit was introduced by Asia Hightower, a PhD student at Michigan State University who is also the Early Career Representative on the ASPB Equity, Diversity, and Inclusion (EDI) committee. Asia relayed how becoming involved in ASPB provided many opportunities to build her network and gave her access to an inclusive, supportive community. As a member of the EDI Committee, Asia has been involved in making ASPB (and the wider plant science community) more equitable, diverse, and inclusive for everyone. These initiatives include ensuring speaker selections are more reflective of the APSB membership as well as collaborating with other societies to further the shared diversity goals. Check out this Plantae article for more information on the ASPB and NAASC “Changing Cultures and Climates” Initiative.

Overview by Chair: Catherine Feuillet

The session organizer Catherine Feuillet introduced Plenary Symposium IV, setting the scene by explaining that the successes of today’s agriculture have had a heavy cost on the environment. The agriculture of tomorrow will look very different in different countries and production systems- but all have common themes: use fewer natural resources, but be more efficient to produce enough food, feed, and fiber. To do so, we need to harness the power of new technologies. We have witnessed many technological advances, namely in genomics, data analytics, and artificial intelligence. By connecting these areas, we will be able to transform plant breeding and modify genomes in very precise ways.

The speakers then provided a broad overview of the potential for genome editing to revolutionize future agriculture.

Pamela Ronald- Harnessing the Potential of Genome Editing for Targeted Insertion in Plants

Conventional gene-editing techniques such as agrobacterium mediated transformation and particle bombardment are suboptimal, as the insertion site of the transgene cassette is random and thus genes important for growth and development can be accidentally disrupted. Furthermore, conventional techniques require a selectable marker linked to the gene of interest which can cause regulatory concerns. First published in 2005 golden rice was genetically engineered using conventional approaches to synthesize beta-carotene, a precursor to vitamin A, the deficiency of which causes 500,000 children to go blind annually. Faced with regulatory hurdles, golden rice has yet to be distributed to growers.

To develop golden rice with fewer regulatory concerns, Pamela Ronald’s group inserted the golden rice cassette into targeted sites in the rice genome using CRISPR-Cas9. CRISPR-Cas9 is an attractive alternative to conventional gene editing approaches as it allows for targeted insertion of transgenes at DNA double-stranded breaks. The Ronald lab identified five genomic “safe harbors” (chromosomal regions that can accommodate a transgene without causing adverse effects on the host) that are optimal for gene insertion in rice using mutant screens, the data of which is available at KitBase. The carotenoid cassette was generated consisting of the psy (phytoene synthase) and crtI (phytoene desaturase) genes accompanied by endosperm specific promoter sequences and was successfully inserted into the rice genome at two designated targets. Marker-free rice with high β-carotene levels was then obtained with no detectable consequences in morphology or yield (see the paper here).

Furthermore, Pamela suggested that gene stacking approaches can be applied using targeted gene insertion methods to insert transgenes at the same loci (colocalized to reduce the possibility of segregation in subsequent generations). For example, the stacking of NLR genes in the same genomic region could enhance the robustness and durability of disease resistance.

Thus, with the advancement in gene insertion techniques, the knowledge of genomic safe harbors, and the understanding of genes to enhance stress tolerance and disease resistance it is certainly an exciting time to work in plant biotechnology.

Omar Abudayyeh and Jonathan Gootenberg- Novel CRISPR Systems for Genome Engineering and Human Health

The Cas9 enzyme in the currently most widely utilized CRISPR system is only a snippet of the actual genetic diversity that exists. Omar Abudayyeh and Jonathan Gootenberg from MIT sought to find new CRISPR systems that could be developed as tools.

Omar described how a computational pipeline was developed, and subsequently, several new CRISPR systems were found, but a particularly fascinating example is Cas13a. This enzyme is an RNA-guided RNase, meaning that it will target RNA instead of DNA and cut the RNA several times, effectively degrading the RNA. An interesting observation was that Cas13a has “collateral activity”. This means that once the Cas13a binds its target it will cleave any RNA in solution and not just complementary to the guide RNA. Using fluorophore-quenchers the potential of this phenomenon was harnessed for nucleic acid diagnostics, the Specific High-sensitivity Enzymatic Reporter Unlocking (SHERLOCK) tool was developed. Omar outlined two applications of SHERLOCK. SHERLOCK has been used for GMO detection in soybean, with a test developed in less than one week, attesting to the applicability of these assays to plant science questions. Furthermore, SHERLOCK is currently being applied for COVID-19 detection.

Jonathan then described the applications of Cas13a as an RNA-targeting tool. RNA-knockdown has been successfully conducted using Cas13 in mammalian cells and rice protoplasts and Cas13 also has higher specificity than RNA interference. Base editing of RNA using Cas13 has also been successful in technologies called REPAIR (RNA Editing for Programmable A to I Replacement) and RESCUE (RNA Editing for Specific C to U Exchange). But as Jonathan outlines this is just the tip of the iceberg in RNA editing tools, and now as the DNA editing toolbox fills it is time to explore that of RNA editing.

Dirk Inzé- Plant Science for Climate Emergency: The Pivotal Role of Genome Editing

Climate change needs to be addressed on all fronts, and while we can adapt plants to withstand and be resilient in future climatic conditions, we should also invest in the mitigatory role plants can play for example in bioenergy and protein crops. Based at VIB in Ghent University Dirk Inzé’s research focus is biomass productivity and while many genetic pathways have been identified as having a role in biomass accumulation/yield, the underlying molecular mechanisms remain poorly understood. Few genes effectively translate from model species to boost productivity in crop species. An example of an exception is the PEAPOD module which is a master regulator of biomass productivity and a growth repressor in Arabidopsis that when downregulated increases plant growth and seed size in soybean and Medicago. However, many growth regulatory genes characterized in Arabidopsis do not affect growth when mutated or over-expressed in crops such as maize. This means that there is a very high attrition rate when it comes to improving biomass productivity in crop species, and Dirk offered some suggestions as to how to improve the efficiency of this research area.

1. “Walk the extra mile and go to the field”. Field trials are essential to accurately evaluate traits, and while some traits such as final plant height can be well predicted by greenhouse experiments other traits like seed yield parameters are impossible to predict.

2. “Learn how genes behave in the field”. From comparative transcriptomic analyses, the Inzé lab found that some genes were significantly differentially expressed between growth chambers and the field. For example, shade avoidance genes were upregulated in field-grown plants, as individuals are cultivated at a higher density than in growth chambers.

3. “Network engineering”. The Inzé lab developed BREEDIT (a combination of breeding and gene editing). BREEDIT involves selecting ~70 yield-related genes to be candidates for gene editing in maize. An “editor plant” constitutively expresses Cas9 and then different combinations of guide RNAs specific to the ~70 candidate genes are introduced. By conducting crosses between edited plants, the collaborative effect of mutations can be investigated and the genotypes of favorable phenotypes can be analyzed.

A major hurdle to integrating gene-edited crops into production systems, particularly in Europe, is the regulatory status of GMOs. Dirk expressed his frustration at the outdated European legislation and outlines the costs if Europe cannot use gene editing e.g. slow adaptation of crops for climate resilience. Over 130 plant science centers have come together in an organization called EU-SAGE (European Sustainable Agriculture through Genome Editing) to urge the EU to reconsider their stance on genome editing. Dirk left us with a very important message: As plant scientists, we have a moral duty to stand up for science, and even when the answers are complex and difficult to explain we need to communicate that this is the right path to take.

Catherine Feuillet – Transforming Plant Breeding at Inari: Challenges and Opportunities

Catherine Feuillet of Inari wrapped up Plenary IV. Catherine emphasized that “seeds are at the core of the global food system”, and while there have been major innovations in food, farming, and other crop inputs, it all comes back to the seed. The seed determines a lot of factors- fertilizer inputs, yield potential, and land footprint. Inari is asking the question: How do we capture (and improve) the full potential of seeds? Catherine presented Inari’s aim to “reimagine biotechnology” and to take advantage of the new technologies in order to “unlock the power of seeds”. This involves employing nature’s intricacies via “breeding by editing” to deliver profound reductions in inputs and increases in yield.

Inari has developed a new model in its Seed Foundry. The Seed Foundry involves predictive design (integrating data from all levels to drive breeding and gene prioritization), a multiplex genetic toolbox (gene editing using a proprietary nuclease system, with efforts to multiplex the system to edit several genes concurrently), step-change products, and a democratized market. Catherine presented Inari’s several successful projects for example a promoter fine-tuning technology was developed with Zach Lippman’s lab at CSHL and a successful example of multiplex gene editing that improved plant architecture in soybean and maize using various combinations of genes.

Catherine highlighted Inari’s alternative business model of fostering a culture of “opportunity through partnership” which emphasizes collaborations and connecting with independent seed companies who possess the knowledge, experience, and relationships with farmers. Finally, Catherine highlighted the diversity of experience and talents of the Inari team, with an emphasis on multidisciplinary work, and highlighted the current vacancies at Inari both in the USA and Europe.

“Now is the time to apply all of these resources and understand biology better” – Catherine Feuillet

Each talk was followed by excellent live Q&A sessions and through the discussion window attendees could submit questions as well as interact and network with each other. Plenary IV was inspiring and motivational and demonstrated that it is a very exciting time to be a plant scientist, and while the future challenges are great, the ongoing technological innovations have huge potential to help overcome these hurdles.

Resources

(In his seminar Dirk recommends “Our Final Warning: Six Degrees of Climate Emergency” by Mark Lynas as a must-read.)