Princeton scientists solve the mystery behind an enigmatic organelle, the pyrenoid

Carbon is one of the main building blocks of life on Earth. It is abundant in the atmosphere of our planet, where it is found in the form of carbon dioxide. Carbon makes its way into the bodies of earthlings primarily through the process of photosynthesis, which incorporates carbon dioxide into sugars that serve as components for important biomolecules and fuel the global food chain. About a third of this process globally is done by unicellular algae that live in the oceans (most of the rest is done by plants).

The enzyme that goes through the first step of the reaction to assimilate carbon dioxide in sugars is a bulky protein called Rubiscus assembled from eight identical small subunits and eight identical large subunits arranged symmetrically together. All parts of this assembly, which is called the holoenzyme, work together to perform Rubisco’s enzymatic duty. Rubisco’s activity rate – and by extension, the growth rate of plants and algae – is limited by its access to carbon dioxide. Free carbon dioxide can be scarce in water, so aquatic algae like Chlamydomonas reinhardtii sometimes he struggles to keep Rubisco at his best. To counter this, these algae have developed a special structure called a pyrenoid to deliver concentrated carbon dioxide to Rubisco. The pyrenoid is so important that almost every algae on the planet has one. Several species of algae are thought to have developed the structure independently.

“The defining feature of a pyrenoid is the matrix, a giant liquid-like condensate that contains nearly all of the cell’s Rubiscus,” explains Jonikas, Assistant Professor in the Princeton Department of Molecular Biology.

Rubisco is the main component of the pyrenoid matrix, but not the only one; in 2016, Jonikas’s lab discovered another protein abundant in the pyrenoid called EPYC1. In their 2016 paper, Jonikas’ group demonstrated that EPYC1 binds to Rubisco and helps concentrate Rubisco in the pyrenoid. The researchers theorized that EPYC1 works as a molecular glue to link Rubisco holoenzymes together. Postdoc Shan He, along with colleagues from Jonikas’s lab and collaborators from Germany, Singapore and England, decided to test this theory.

“In the present work, we demonstrate that this is indeed the way it works,” says Jonikas, “by showing that EPYC1 has five binding sites for Rubisco, allowing it to ‘link’ multiple Rubisco holoenzymes together.”

EPYC1 is an extended and loosely structured protein, and its five Rubisco binding sites are evenly distributed over its entire length. The researchers also found that Rubisco has eight EPYC1 binding sites evenly distributed across its ball-like surface. Computer modeling showed that the freely structured and flexible EPYC1 protein can make multiple contacts with a single Rubisco holoenzyme or link neighboring ones together. In this way, EPYC1 pushes Rubisco to group in the pyrenoid matrix.

While this offers a satisfying explanation for how the matrix is ​​assembled, it poses something of an enigma. Other proteins need to have access to Rubisco to repair it when it breaks. If the EPYC1-Rubisco network is rigid, it could prevent these proteins from accessing Rubisco. However, He and colleagues found that EPYC1’s interactions with Rubisco are quite weak, so although the two proteins can form many contacts with each other, these contacts are swapping rapidly.

“This allows EPYC1 and Rubisco to flow alongside each other while remaining in a densely packed condensate, allowing other pyrenoid proteins to access Rubisco as well,” Jonikas notes. “Our work solves the long-standing mystery of how Rubisco is held together in the pyrenoid matrix.”

Terrestrial plants do not have pyrenoids, and scientists think that engineering a pyrenoid-like structure in cultivated plants could increase their growth rates. Understanding how the pyrenoid is assembled in algae represents a significant step towards such efforts.

“He and his colleagues provide a very nice molecular study of the protein-protein interactions between the small Rubisco subunit and EPYC1,” says Dr. James Moroney, a professor of biology in the Louisiana State University department of biological sciences, whose lab studies photosynthesis in plants and algae.

“This work is encouraging for researchers looking to introduce pyrenoid-like structures into plants to improve photosynthesis,” he adds.

In a world plagued by hunger and disease, we can use any stimulus we can get.

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Funding: The work described here was supported by grants to MCJ from the National Science Foundation (# IOS-1359682 and MCB-1935444), National Institutes of Health (# DP2-GM-119137) and Simons Foundation and Howard Hughes Medical Institute (No. 55108535); a BDE of Deutsche Forschungsgemeinschaft (EN 1194 / 1-1 as part of FOR2092); to OM-C. by the Ministry of Education (MOE Singapore) Tier 2 (n. MOE2018-T2-2-059); to AJM and NA from the British Council for Research in Biotechnology and Life Sciences (n. BB / S015531 / 1) and Leverhulme Trust (n. RPG-2017-402); to FMH of NIH (R01GM071574); to SAP from the Deutsche Forschungsgemeinschaft scholarship (n. PO2195 / 1-1); and to the VKC by a training grant from the National Institute of General Medical Sciences of the Institutes of Health (n. T32GM007276).

Quote: Shan He, Hui-Ting Chou, Doreen Matthies, Tobias Wunder, Moritz T. Meyer, Nicky Atkinson, Antonio Martinez-Sanchez, Phillip D. Jeffrey, Sarah A. Port, Weronika Patena, Guanhua He, Vivian K. Chen, Frederick M. Hughson, Alistair J. McCormick, Oliver Mueller-Cajar, Benjamin D. Engel, Zhiheng, Yu and Martin C. Jonikas. The structural basis of the Rubisco phase separation in the pyrenoid. Natural plants. 2020. doi: 10.1038 / s41477-020-00811-y