In a significant stride towards more sustainable forestry, a team of researchers at North Carolina State University (NC State) has successfully used CRISPR gene-editing techniques to breed poplar trees with lower levels of lignin. Lignin, a substance that poses a significant challenge to the sustainable production of wood fibers, has decreased in these trees while their wood properties have markedly improved.

The findings of this research, published recently in Science, promise to greenify and economize the production of fiber for a wide range of products, from paper to diapers, thereby making it more efficient.

CRISPR pioneer Rodolphe Barrangou and tree geneticist Jack Wang led the team. The duo, along with their team, relied on predictive modeling to set goals of not just reducing the lignin levels but also increasing the carbohydrate to lignin ratio (C/L) and the syringyl to guaiacyl (S/G) ratio, both essential lignin building blocks, in the poplar trees.

“CRISPR systems provide the flexibility to edit more than just single genes or gene families, allowing for greater improvement to wood properties,” said Barrangou, the Todd R. Klaenhammer Distinguished Professor of Food, Bioprocessing, and Nutrition Sciences at NC State. 

The innovative use of machine-learning models aided the researchers in sifting through approximately 70,000 gene-editing strategies, all targeting 21 crucial genes related to lignin production. Of these strategies, the model identified 347 as being the most promising, with more than 99% of these targeting at least three genes.

The team then focused on seven strategies that were predicted to achieve the desired chemical balance. These strategies resulted in trees with 35% less lignin than their wild counterparts, C/L ratios that were more than 200% higher, and S/G ratios that were similarly amplified. Crucially, the growth rates of these gene-edited trees were found to be comparable to those of wild trees.

Of the seven strategies, the researchers utilized CRISPR gene editing to create 174 lines of poplar trees. Upon inspection after six months in an NC State greenhouse, some varieties of these trees showed a lignin content reduction of up to 50%, and others exhibited a 228% increase in the C-L ratio.

“We show that multiplex CRISPR editing enables precise woody feedstock design for combinatorial improvement of lignin composition and wood properties, the authors wrote. “By assessing every possible combination of 69,123 multigenic editing strategies for 21 lignin biosynthesis genes, we deduced seven different genome editing strategies targeting the concurrent alteration of up to six genes and produced 174 edited poplar variants. CRISPR editing increased the wood carbohydrate-to-lignin ratio up to 228% that of wild type, leading to more-efficient fiber pulping.”

Reducing lignin content is expected to enhance pulp yield and minimize the creation of black liquor, a byproduct of pulping. According to pulp production mill models, these advancements could lead to a 40% increase in sustainable fiber production. Further, these efficiencies could reduce the greenhouse gases linked to pulp production by up to 20%.

In addition to their economic impact, the ecological implications of this research are profound. Forest trees are key players in efforts to mitigate climate change as they represent the largest biogenic carbon sink on earth. In North Carolina alone, forestry contributes over $35 billion to the local economy while supporting around 140,000 jobs.

Wang, an assistant professor and the director of the Forest Biotechnology Group at NC State, emphasized the crucial role of these gene-edited trees in a world facing climate change and an increased need for sustainable biomaterials.

The next stage of this research involves further greenhouse tests to compare the performance of gene-edited trees to wild trees. The team hopes to subsequently conduct field trials to assess how well these trees adapt to life outdoors, outside of the controlled greenhouse environment.

“An interdisciplinary approach to tree breeding that combines genetics, computational biology, CRISPR tools, and bio-economics has profoundly expanded our knowledge of tree growth, development, and forest applications,” said Daniel Sulis, a postdoctoral scholar at NC State.

Reflecting NC State's commitment to innovation in plant sciences and forestry, Barrangou, and Wang have established a startup company, TreeCo. This venture seeks to harness the power of genome editing to create healthier, more sustainable forests and to foster a more eco-friendly future.