Anika Ajgaonkar's notes on CRISPR-Cas edited trees.
Among scientists and students well-acquainted in the world of biotechnology, CRISPR has become a familiar concept for those exploring how to tinker with the genetic code of life itself: DNA, short for deoxyribonucleic acid. Though its potential to alter human genes has been the main topic of discussion in the media, such as eliminating genetic diseases in our bodies, its industrial and agricultural applications are somewhat lesser known. A new such usage has arisen thanks to researchers at North Carolina State University, who recently published an article in the journal Science detailing their work in editing poplar trees with CRISPR and improving their wood quality for use in fiber production.
CRISPR, an abbreviation for Clustered Regularly Interspaced Short Palindromic Repeats, is essentially a bacterial defense system that forms the backbone of CRISPR-Cas9 genome editing technology. It edits genes by using CRISPR RNAs to guide the system to the corresponding DNA sequences, then precisely cut the genes with the enzyme Cas9, which shuts the targeted gene off. Modified versions of Cas9 can also activate gene expression, which makes it a powerful tool for scientists to deliberately manipulate gene expression to correct errors in DNA.Rodolphe Barrangou, NC State University team lead in CRISPR pursuits, and Jack Wang, a well-known tree geneticist at NC State, led a team of researchers with three important goals for this research: lowering lignin levels, increasing the carbohydrate to lignin ratio (C/L) ratio, and increasing the ratio of syringyl to guaicyl (S/G, two important building blocks for lignin) in poplar trees.
Lignin is a major component of trees and plants alongside cellulose. It provides rigidity and support to trees, but also makes them resistant to water and physical degradation, which interferes with the production of fiber for industrial purposes.Combined, these factors create the perfect conditions for fiber and pulp production. Barrangou, the Todd R. Klaenhammer Distinguished Professor of Food, Bioprocessing, and Nutrition Sciences at NC State and co-corresponding author of the paper, says that CRISPR is especially useful because of its ability to edit beyond just single genes or gene families, enabling more improvement of the properties of wood.
Using predictive modeling, the software predicted and sorted through nearly 70,000 strategies for gene editing that targeted 21 genes associated with producing lignin. Some strategies used a combination of editing several genes at a time. In total, it yielded 327 strategies to do so, with 99% of the strategies targeting three genes or more. After that, the researchers chose seven strategies created by the model that would create the best possible conditions for fiber production: 35% less lignin than wild trees, C/L ratios, and S/G ratios over 200% higher than wild trees, and similar growth rates.
CRISPR gene editing was then used to produce 174 lines of poplar trees; after half a year in a greenhouse, certain trees were demonstrating up to 50% reduced lignin content, and others showed more than a 228% increase in the C-L ratio. The researchers noted that the most lignin reduction occurred in trees with four to six edits, while trees that had three edits showed more limited lignin reduction. Single-gene edits did not yield noticeable changes, suggesting that CRISPR’s use in fiber production to make multigene changes offers many advantages.
Other types of modeling used in the study, such as pulp production mill models, suggested that reducing lignin content in trees could increase pulp yield and also decrease the amount of “black liquor,” a major pulping by-product. If eliminated, it could help mills produce fibers up to 40% more sustainable. And if the reduced lignin production and increased C/L and S/G ratios are achieved in trees at an industrial level, the associated greenhouse gas production of fiber production could be reduced by up to 20%. Wang, assistant professor and director of the Forest Biotechnology Group at NC State and co-corresponding author of the paper, remarked that genome editing is a great opportunity to improve the sustainability of our forests at a time when they are challenged by climate change.
The team will continue to observe how the gene-edited trees fare in the greenhouse compared to wild trees and perform field trials to see if the tree can withstand real-world, outdoor stressors.
The researchers highlighted the importance of an interdisciplinary approach to tree breeding, combining computational biology, CRISPR tools, and bio-economics to carry out their research and expand their knowledge of the growth and development of trees, as well as forest applications. Daniel Sulis, a postdoctoral scholar at NC State and the first author of the paper, remarked, “This powerful approach has transformed our ability to unravel the complexity of tree genetics and deduce integrated solutions that could improve ecologically and economically important wood traits while reducing the carbon footprint of fiber production.”
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