Photosynthesis is a process carried out by plants and algae that uses light energy to power the assimilation of atmospheric carbon dioxide into sugar, with oxygen produced as a by-product. Despite decades of study, we still have only a rough idea of how photosynthesis works; we haven’t even managed to identify all the proteins involved. In a paper published December 7^th, 2023 in the journal Cell, Princeton scientists Moshe Kafri, Weronika Patena, Lance Martin, Martin Wühr, Martin Jonikas, and colleagues characterize 70 new proteins involved in photosynthesis and map out many of their functions to extend our understanding of this life-sustaining process.

Photosynthesis happens in a structure called the chloroplast, which is found inside the cells of land plants, aquatic plants and algae. Proteins involved in photosynthesis belong to one of several different complexes, or functional assemblies, that enact particular steps in photosynthesis. The genes encoding instructions for making most of these proteins are located in the DNA of the cell’s nucleus.

  In an effort to identify the proteins involved, Jonikas’ research team and others conducted large-scale screens in the single-celled aquatic green alga Chlamydomonas reinhardtii, whose photosynthetic apparatus is similar to that found in plants. Such screens work by inserting a small chunk of DNA at a random location in the nuclear DNA. Insertions that land in a photosynthesis gene will inactivate it and thereby hamper the organism’s ability to grow in light, allowing scientists analyze the DNA surrounding the insertion site in defective cells to identify which gene was disrupted. However, there is a catch: if more than one gene gets disrupted it can be difficult to tell which one is responsible. 

Kafri, Patena, Jonikas, and colleagues started their work with 1,781 mutant C. reinhardtii cells that had been created as part of an earlier screen and that were known to have defects in photosynthesis. The genes linked to these defects had not yet been identified, often because the mutants had more than one disruption in their genome. 

To solve this problem, the researchers mated each mutant cell with a normal C. reinhardtii cell and then examined the progeny of the mating. If more than one gene is disrupted in the mutant parent, some progeny of such a mating will inherit only one or the other defective gene, but not both. Genes linked to photosynthesis can be identified because the progeny lacking these genes will always exhibit photosynthesis defects. While it would have been prohibitively time-consuming to do this for every mutant individually, the Princeton scientists devised a way to perform this analysis in large batches. Using this approach, the researchers identified 115 genes required for photosynthesis, 70 of which had never been characterized before. 

The team investigated why loss of these particular genes brings about defective photosynthesis. Earlier studies had shown that loss of one photosynthesis protein tends to affect the abundance of other proteins that work together with it in photosynthesis, so the researchers used mass spectrometry to investigate how the loss of each of their proteins affected the amounts of other proteins. Having narrowed down the functional roles for several previously-uncharacterized proteins on their list, they could then zero in on a few particularly interesting proteins for further study—including several new proteins that, they showed, regulate the production of one or more photosynthetic complexes. Intriguingly, one of these is a master regulator that coordinates the production of the entire photosynthetic apparatus.  

By identifying these new regulatory components and showing how other proteins slot into their functional roles, this work substantially extends our understanding of photosynthesis.

Citation: Moshe Kafri, Weronika Patena, Lance Martin, Lianyong Wang, Gillian Gomer, Sabrina L. Ergun, Arthur K. Sirkejyan, Audrey Goh, Alexandra T. Wilson, Sophia E. Gavrilenko, Michal Breker, Asael Roichman, Claire D. McWhite, Joshua D. Rabinowitz, Frederick R. Cross, Martin Wühr, and Martin C. Jonikas. 2023. Systematic identification and characterization of novel genes in the regulation and biogenesis of photosynthetic machinery. Cell. DOI: 10.1016/j.cell.2023.11.007.

Funding: This work was supported by the Princeton Catalysis Initiative, U.S. National Institutes of Health (R35GM128813), U.S. National Foundation (MCB-1914989), European Molecular Biology Organization fellowship (ALTF 1006-2017), Human Frontier Scientific Program fellowship (LT000031/2018-L), HHMI/Simons Foundation (55108535), the Lewis-Sigler Scholars Fund, and the Howard Hughes Medical Institute.

Funders: Princeton Catalysis Initiative, U.S. National Institutes of Health, U.S. National Foundation, European Molecular Biology Organization fellowship, Human Frontier Scientific Program fellowship, HHMI/Simons Foundation, Lewis-Sigler Scholars Fund, Howard Hughes Medical Institute.

Grant numbers: R35GM128813, MCB-1914989, ALTF 1006-2017, LT000031/2018-L, 55108535