ARTICLE Received 4 Aug 2015 | Accepted 23 Dec 2015 | Published 4 Feb 2016 DOI: 10.1038/ncomms10543 OPEN A role for diatom-like silicon transporters in calcifying coccolithophores Graz˙yna M. Durak1,*,w, Alison R. Taylor2,*, Charlotte E. Walker1, Ian Probert3, Colomban de Vargas3, Stephane Audic3, Declan Schroeder1, Colin Brownlee1,4 & Glen L. Wheeler1 Biomineralization by marine phytoplankton, such as the silicifying diatoms and calcifying coccolithophores, plays an important role in carbon and nutrient cycling in the oceans. Silicification and calcification are distinct cellular processes with no known common mechanisms. It is thought that coccolithophores are able to outcompete diatoms in Si-depleted waters, which can contribute to the formation of coccolithophore blooms. Here we show that an expanded family of diatom-like silicon transporters (SITs) are present in both silicifying and calcifying haptophyte phytoplankton, including some globally important coccolithophores. Si is required for calcification in these coccolithophores, indicating that Si uptake contributes to the very different forms of biomineralization in diatoms and coccolithophores. Significantly, SITs and the requirement for Si are absent from highly abundant bloom-forming coccolithophores, such as Emiliania huxleyi. These very different requirements for Si in coccolithophores are likely to have major influence on their competitive interactions with diatoms and other siliceous phytoplankton. 1 Marine Biological Association, The Laboratory, Citadel Hill, Plymouth, Devon PL1 2PB, UK. 2 Department of Biology and Marine Biology, University of North Carolina Wilmington, 601 South College Road, Wilmington, North Carolina, 28403-5915, USA. 3 Station Biologique de Roscoff, Place Georges Teissier, 29680 Roscoff, France. 4 School of Ocean and Earth Sciences, University of Southampton, National Oceanography Centre, Southampton SO14 3ZH, UK. * These authors contributed equally to this work. w Present address: University of Konstanz, Department of Chemistry, Physical Chemistry, Universita¨tsstr. 10, Box 714, D-78457 Konstanz, Germany. Correspondence and requests for materials should be addressed to C.B. (email: [email protected]) or to G.W. (email: [email protected]). NATURE COMMUNICATIONS | 7:10543 | DOI: 10.1038/ncomms10543 | www.nature.com/naturecommunications 1 ARTICLE NATURE COMMUNICATIONS | DOI: 10.1038/ncomms10543 he biomineralized phytoplankton are major contributors to an important contributory factor in the formation of some marine primary productivity and play a major role in coccolithophore blooms. Major E. huxleyi blooms in areas such as Tcarbon export to the deep oceans by promoting the sinking the North Atlantic, the Black Sea and off the Patagonian of organic material from the photic zone1,2. The two primary shelf have been associated with low silicate availability16–18. forms of biomineralization found in marine plankton are the These observations support the view that the ecological niche of precipitation of silica (by diatoms, chrysophytes, synurophytes, coccolithophores is partly defined by conditions that reduce dictyochophytes, choanoflagellates and radiolarians) and competition with the fast-growing resource-efficient diatoms, calcium carbonate (by coccolithophores, foraminifera, ciliates such as in areas of low silicate where other nutrients (for example, and dinoflagellates)3. These processes require very different nitrate and phosphate) remain available19. chemistries and exhibit no known shared mechanisms. To further understand the evolution of biomineralization in Both silicification and calcification appear to have evolved haptophytes, we characterized the cellular mechanisms underlying independently on multiple occasions. However, since in many silica scale formation in P. neolepis. We examine commonalities cases the underlying cellular mechanisms have not been with other silicified organisms and determine whether any elucidated, the evolutionary processes remain unclear. Improved common cellular mechanisms contribute to biomineralization in knowledge of the cellular mechanisms of biomineralization will silicified and calcified haptophytes. Surprisingly, given that it is allow us to understand the impact of past climatic events on the generally assumed that coccolithophores lack a requirement for Si, major phytoplankton lineages and better predict their response to we identify that diatom-like Si transporters are present in future environmental change. haptophytes, not only in the silicified P. neolepis but also in some The haptophyte algae are of particular interest in the evolution important calcifying coccolithophore species. We demonstrate that of biomineralization as they include closely related silicified Si plays an important role in formation of calcite coccoliths in and calcified representatives. The coccolithophores (Calcihapto- these coccolithophores, but that the requirement for Si is phycidae)4 produce an extracellular covering of ornate significantly absent from the most abundant species in present calcium carbonate plates (coccoliths) and are major day oceans, E. huxleyi. The findings have important implications contributors to biogenic calcification in the ocean3. The most for the evolution of the biomineralized phytoplankton and their abundant coccolithophore species in modern oceans are distribution in both past and modern oceans. Emiliania huxleyi and Gephyrocapsa oceanica, which belong to the Noelarhabdaceae. These species have a small cell size and are able to form extensive blooms. Larger coccolithophores species Results such as Coccolithus braarudii and Calcidiscus leptoporus are less Mechanisms of biomineralization in a silicifying haptophyte. numerous, but as they are heavily calcified they are important The known mechanisms of biosilicification in eukaryotes involve contributors to global calcification5. Much of our understanding a number of common elements; a mechanism for Si uptake, an of coccolithophore biology comes from the study of E. huxleyi, acidic silica deposition vesicle and an organic matrix for but emerging evidence suggests that there is considerable catalysing and organizing silica precipitation20. However, there is physiological diversity among coccolithophores6. little evidence for shared mechanisms at the molecular level, Though the biomineralized haptophytes are predominately suggesting that silicification has evolved independently in many calcified, a representative was recently described, Prymnesium lineages. We therefore examined the mechanisms of silicification neolepis (formerly Hyalolithus neolepis), which is covered with in P. neolepis, using both molecular and physiological approaches. silica scales and resembles a ‘silicified coccolithophore’7–9. The At low Si concentrations, Si uptake in diatoms is performed by a silica scales are produced intracellularly and then deposited family of Na þ -coupled high-affinity Si transporters (SITs), outside the plasma membrane, in a manner analogous to although diatoms may also acquire Si by diffusive entry at coccolith secretion8,10. The Prymnesiales are estimated to have higher Si concentrations21,22. Silicified sponges and land plants diverged from the coccolithophores around 280 Myr ago11 and do not contain SITs, but use alternative mechanisms for Si P. neolepis is the only known extensively silicified haptophyte. transport23,24. A search for putative Si transporters in the Understanding whether common cellular mechanisms contribute transcriptome of P. neolepis strain PZ241 (Supplementary to silica scale production in P. neolepis and coccolith formation in Fig. 1) identified a single gene bearing similarity to the SITs the coccolithophores may help us to understand how these (PnSIT1). PnSIT1 exhibits 24.9–29.3% identity and 39.8–47.0% different forms of biomineralization have evolved in the similarity to diatom SITs at the amino-acid level (sequences used haptophytes and also in other phytoplankton lineages. for comparison were Cylindrotheca. fusiformis AAC49653.1, Silicification by marine phytoplankton has both contributed to Thalassiosira. pseudonana ABB81826.1 and Phaeodactylum and been influenced by the marked changes in the biogeo- tricornutum ACJ65494.1). SITs have only previously been chemistry of Si in the surface ocean. The diatoms, representing identified in siliceous stramenopiles (diatoms and chrysophytes) the dominant silicifying phytoplankton in current oceans, and choanoflagellates25–27. Many features of PnSIT1 are appeared only relatively recently in the fossil record (120 Myr conserved with these SITs, including the 10 predicted ago) and their expansion in the Cenozoic resulted in the extensive transmembrane regions and the pair of motifs (EGxQ and depletion of silicate from the surface ocean, leading to the decline GRQ) between TM2-3 and TM7-8 (ref. 27; Supplementary of heavily silicified sponges and decreased silicification in Fig. 2). We also identified a homologue of the Si efflux protein, radiolarians12–15. Si has therefore become a limiting nutrient Lsi2 in P. neolepis (Supplementary Table 1). Lsi2 is related to the for modern silicifying phytoplankton and is an important factor bacterial arsenate transporter ArsB and mediates Si efflux in plant in competitive interactions with non-silicifying taxa. As the cells28. Lsi2 is also present in diatoms and its transcriptional regeneration of available Si from silica dissolution is slow, regulation is highly similar to SIT2 in Thalassiosira pseudonana, diatom blooms can deplete Si in the surface ocean sufficiently to although