Food crops worldwide are increasingly coming under pressure from extended droughts and other extreme weather events. Many food crops grow best in a cooler, wetter climate, but with global temperature increases projected to continue and even accelerate, those crops will need help to cope with the change—and also to keep pace with the nutritional needs of a burgeoning world population that has now surpassed 8 billion people. In a paper published November 15, 2024 in the journal Nature Plants, Princeton researchers Jessica Hennacy, Martin Jonikas and colleagues report on a major advance in the quest to do just that. 

Plants use a process called carbon fixation to incorporate CO₂ pulled from the environment into the starches they use to build cell walls and energy stores. Food crops such as rice and wheat are valuable because they contain large amounts of edible starches, but they aren’t the best carbon fixers on the planet. In this respect, they’re handily surpassed by humble, single-celled aquatic algae, which are responsible for around one third of global carbon fixation. This is thanks to a structure found in algae but not rice or wheat, called the pyrenoid.

“The rate of carbon fixation is limited by the low availability of CO₂ to the CO₂-fixing enzyme RuBisCO,” said Jonikas, a Professor at Princeton’s Department of Molecular Biology. “The pyrenoid overcomes this limitation by supplying concentrated CO₂ to RuBisCO.”

With the pyrenoid’s ability to concentrate CO₂ for RuBisCO, carbon fixation by algae is far more efficient and requires less water than the same process in terrestrial plants, which rely on simple diffusion to supply their Rubsico with CO₂. This presents an enormous opportunity.

“Engineering a pyrenoid into plants could increase yields of major crops by up to 50%,” said Jonikas.

There has been steady progress toward this goal, starting with efforts to understand how the pyrenoid works. Studies performed in the single-celled alga Chlamydomonas reinhardtii have demonstrated that the algal pyrenoid consists of a starch-rich outer sheath surrounding a central matrix that is densely packed with the enzyme RuBisCO. The RuBisCO matrix is traversed by pipe-like membrane tubules that serve to deliver concentrated CO₂ to the matrix. Without these tubules, it’s thought that RuBisCO in the center of the matrix would be starved of CO₂ and unable to perform its job.

In 2016, Jonikas’s team discovered an algal protein that in 2020 enabled their collaborators Nicky Atkinson and Alistair McCormick at the University of Edinburgh to generate a RuBisCO matrix similar to that observed in algae in a small terrestrial plant, the mustard family weed Arabidopsis thaliana. However, these structures lack the pyrenoid’s membrane tubules, indicating that other, as yet unidentified algal genes are involved in their formation. Until now, this has proven a difficult hurdle in the effort to construct a pyrenoid for terrestrial plants.

The groundwork for a breakthrough was laid in 2022 when Jonikas’s research group performed a large-scale genetic screen to identify genes that affect algal growth. The screen showed that algal cells with defects in two proteins, MITH1 and SAGA1, grow poorly in conditions of low CO₂, which is when the CO₂-concentrating ability of the pyrenoid tubules is most needed. Spurred this discovery, Princeton graduate student Jessica Hennacy hypothesized that MITH1 and SAGA1 could be required for tubule formation. Indeed, when Hennacy examined the mutant algae, she discovered that algae missing either protein lack pyrenoid tubules. Subsequent experiments showed that SAGA1 initiates the process of tubule formation by binding to RuBisCO, while MITH1 promotes tubule extension into the RuBisCO matrix. The piece de resistance was the demonstration that MITH1 and SAGA1 can also work together to generate tubules in Arabidopsis that already sport an engineered algal RuBisCO matrix.

“Our collaborators Nicky Atkinson and Alistair McCormick made that discovery,” said Jonikas.

It is remarkable that just four algal genes—two to generate the matrix, and two to form tubules—are needed to generate such a complex structure. These discoveries could put the goal of reconstituting a functional pyrenoid for terrestrial plants within reach.

“To achieve a minimally functional pyrenoid, the last thing we need to do is to deliver CO₂ into the pyrenoid via the traversing membranes. We think this will require expressing only two additional proteins,” said Jonikas.

The researchers have already identified proteins they think will do the job and are currently working to engineer them into plants.

Citation: Jessica H. Hennacy, Nicky Atkinson, Angelo Kayser-Browne, Sabrina L. Ergun, Eric Franklin, Lianyong Wang, Simona Eicke, Yana Kazachkova, Moshe Kafri, Friedrich Fauser, Josep Vilarrasa-Blasi, Robert E. Jinkerson, Samuel C. Zeeman, Alistair J. McCormick, Martin C. Jonikas. SAGA1 and MITH1 produce matrix-traversing membranes in the CO₂-fixing pyrenoid. 2024. Nature Plants. DOI: https://www.nature.com/articles/s41477-024-01847-0

Funding: Work described here was funded by grants from the U.S. National Institutes of Health (T32GM007388); the Howard Hughes Medical Institute; the Howard Hughes Medical Institute and Simons Foundation Faculty Scholar program (55108535); the U.S. National Science Foundation (MCB-1935444); the U.S. National Institutes of Health (1R01GM140032-01); the U.S. Department of Energy (DE-SC0020195); the Bill and Melinda Gates Foundation and the UK Foreign, Commonwealth, and Development Office (INV-054558); the UK Research and Innovation Biotechnology and Biological Sciences Research Council (BB/S015531/1) and Leverhulme Trust (RPG-2017-402); and the Wellcome Trust Multi-User Equipment Grant (WT104915MA).