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Microalgae play an essential role in global net primary productivity and global biogeochemical cycling. Despite their phototrophic lifestyle, over half of algal species depend for growth on acquiring an external supply of the corrinoid vitamin B12 (cobalamin), a micronutrient produced only by a subset of prokaryotic organisms. Previous studies have identified protein components involved in vitamin B12 uptake in bacterial species and humans. However, little is known about its uptake in algae. Here, we demonstrate the essential role of a protein, cobalamin acquisition protein 1 (CBA1), in B12 uptake in Phaeodactylum tricornutum using CRISPR-Cas9 to generate targeted knockouts and in Chlamydomonas reinhardtii by insertional mutagenesis. In both cases, CBA1 knockout lines could not take up exogenous vitamin B12. Complementation of the C. reinhardtii mutants with the wild-type CBA1 gene restored B12 uptake, and regulation of CBA1 expression via a riboswitch element enabled control of the phenotype. When visualized by confocal microscopy, a YFP-fusion with C. reinhardtii CBA1 showed association with membranes. Bioinformatics analysis found that CBA1-like sequences are present in all major eukaryotic phyla. In algal taxa, the majority that encoded CBA1 also had genes for B12-dependent enzymes, suggesting CBA1 plays a conserved role. Our results thus provide insight into the molecular basis of algal B12 acquisition, a process that likely underpins many interactions in aquatic microbial communities., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2023. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Cobalamin (vitamin B 12 ) is a cofactor for essential metabolic reactions in multiple eukaryotic taxa, including major primary producers such as algae, and yet only prokaryotes can produce it. Many bacteria can colonize the algal phycosphere, forming stable communities that gain preferential access to photosynthate and in return provide compounds such as B 12 . Extended coexistence can then drive gene loss, leading to greater algal-bacterial interdependence. In this study, we investigate how a recently evolved B 12 -dependent strain of Chlamydomonas reinhardtii, metE7, forms a mutualism with certain bacteria, including the rhizobium Mesorhizobium loti and even a strain of the gut bacterium E. coli engineered to produce cobalamin. Although metE7 was supported by B 12 producers, its growth in co-culture was slower than the B 12 -independent wild-type, suggesting that high bacterial B 12 provision may be necessary to favour B 12 auxotrophs and their evolution. Moreover, we found that an E. coli strain that releases more B 12 makes a better mutualistic partner, and although this trait may be more costly in isolation, greater B 12 release provided an advantage in co-cultures. We hypothesize that, given the right conditions, bacteria that release more B 12 may be selected for, particularly if they form close interactions with B 12 -dependent algae., (© 2022 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)