Zebrafish can synthesize a sunscreen compound called gadusol, which was previously thought to be acquired only through the diet

Balskus, Emily P

Brotherton, Carolyn A

eLife

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Your story mattersCitation Brotherton, Carolyn A., and Emily P Balskus. 2015. “Sheddinglight on sunscreen biosynthesis in zebrafish.” eLife 4 (1): e07961.doi:10.7554/eLife.07961. http://dx.doi.org/10.7554/eLife.07961.Published Version doi:10.7554/eLife.07961Citable link http://nrs.harvard.edu/urn-3:HUL.InstRepos:16120932Terms of Use This article was downloaded from Harvard University’s DASHrepository, and is made available under the terms and conditionsapplicable to Other Posted Material, as set forth at http://nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of-use#LAAelifesciences.orgINSIGHTBIOCHEMISTRYShedding light on sunscreenbiosynthesis in zebrafishZebrafish can synthesize a sunscreen compound called gadusol, whichwas previously thought to be acquired only through the diet.CAROLYN A BROTHERTON AND EMILY P BALSKUSRelated research article Osborn A,Almabruk K, Holzwarth G, Asamizu S, LaDu J,Kean K, Karplus PA, Tanguay R, Bakalinsky A,Mahmud T. 2015. De novo synthesis ofa sunscreen compound in vertebrates.eLife 4:e05919. doi: 10.7554/eLife.05919Image Species including fish, birds andreptiles have genes that may enable themto produce sunscreen moleculesJust as many people apply sunscreen beforethey go to the beach, bacteria, fungi andalgae that spend much of their time in thesun also need to protect themselves from solarradiation. These organisms make small moleculesthat act as sunscreens by absorbing harmfulultraviolet (UV) radiation. Small-molecule sun-screens—which include compounds called themycosporine-like amino acids (MAAs) and a re-lated compound called gadusol—are also foundin corals, marine invertebrates and fish.Sunscreen compounds like gadusol and theMAAs are important for the development,lifestyle and health of marine-dwelling organisms.For example, gadusol is found in high concen-trations in the roes of fish, such as cod (Placket al., 1981), as well as in sea urchin eggs(Chioccara et al., 1986), and it has beensuggested that gadusol helps with embryonicdevelopment in these organisms. Sunscreencompounds also perform an array of roles inother organisms, including enhancing the UV-light vision of mantis shrimp (Bok et al., 2014).It was thought for many years that the abilityto synthesize small-molecule sunscreens waslimited to microbes, and that higher marineorganisms obtained these compounds exclu-sively from their diet. Now, in eLife, TaifoMahmud and co-workers at Oregon State Uni-versity—including Andrew Osborn, KhaledAlmabruk and Garrett Holzwarth as joint firstauthors—show that zebrafish can synthesizegadusol (Osborn et al., 2015). They also presentevidence that the pathway used by zebrafish tomake gadusol is distinct from the pathway usedby microorganisms to synthesize MAAs. More-over, they show that the gadusol biosyntheticgenes are also found in the genomes of birds,reptiles and other organisms.In microorganisms that biosynthesize MAAs, thebiosynthetic pathway involves three core enzymesand uses a compound called sedoheptulose-7-phosphate as the starting material (Balskus andWalsh, 2010). The first step is performed by adehydroquinate synthase-like enzyme. This enzymeis a member of a family of enzymes called the sugarphosphate cyclases, which catalyze the conversionof sugar molecules to products that contain astructure called a cyclohexane ring (Asamizu et al.,2012). Dehydroquinate synthase has a central rolein the biosynthesis of aromatic amino acids, and isfound in many branches of the tree of life—such asbacteria, archaea, fungi, plants and algae—but notin vertebrates.Given the lack of sugar phosphate cyclases invertebrates, the Oregon State team was sur-prised to find a gene that encoded a relatedenzyme in the genomes of fish, including zebra-fish (Danio rerio), a well-known model organism.This gene was similar to the EEVS (short for 2-epi-5-epi-valiolone synthase) genes involved in theCopyright Brotherton and Balskus.This article is distributed under theterms of the Creative CommonsAttribution License, which permitsunrestricted use and redistributionprovided that the original author andsource are credited.Brotherton and Balskus. eLife 2015;4:e07961. DOI: 10.7554/eLife.07961 1 of 3biosynthesis of bacterial natural products such asantibiotics and fungicides (Yu et al., 2005).The EEVS-like gene was clustered in the zebrafishgenome with a gene of unknown function namedMT-Ox (for methylation and oxidation). Surround-ing these two genes were genes encoding severaltranscription factors that regulate essential cellu-lar processes. The similarity of the EEVS-like geneto the genes that encode sugar phosphatecyclases in microbes raised the possibility thatthese zebrafish genes might be involved in thesynthesis of related small molecules.To explore this further, the team firstexpressed the zebrafish EEVS-like protein inEscherichia coli and incubated it alongsidesedoheptulose-7-phosphate (the starting mate-rial used by microorganisms to make MAAs).The product of this reaction was 2-epi-5-epi-valiolone, which contains a cyclohexane ring. Thisconfirms that the EEVS-like protein is a functionalsugar phosphate cyclase.The MT-Ox protein was then expressed inE. coli and incubated with its predicted cofactorsand the 2-epi-5-epi-valiolone produced by theEEVS-like protein. This reaction produced a mol-ecule that strongly absorbed UV light; structuralstudies revealed this was the small-moleculesunscreen gadusol. This work represents the firstdiscovery of a gadusol biosynthetic pathway.Unexpectedly, the enzymes involved are distinctfrom those used by microbes to produce theclosely related MAAs.Having demonstrated that these two zebrafishenzymes could synthesize a small-molecule sun-screen, the Oregon State team next examinedwhether gadusol is actually produced in zebra-fish. Quantitative reverse transcription-PCR anal-ysis and organic extraction of zebrafish embryosshowed that the gadusol biosynthetic genes areexpressed in the developing embryo and thatgadusol is indeed produced. These results clearlyindicate that the gadusol found in zebrafish doesnot come from the dietary intake of this molecule.Instead, this sunscreen is biosynthesized from theprimary metabolite sedoheptulose-7-phosphateby the activities of the EEVS-like and MT-Oxenzymes.The team then used bioinformatics to examinewhether the gadusol biosynthetic gene clusteroccurs in other organisms, including algae,invertebrates and chordates. Unexpectedly, thegene cluster is found in many organisms that arenot known to produce gadusol, including otherfish, birds, reptiles and amphibians. However, it isnot present in mammals or in marine animalscalled tunicates and lancelets. Furthermore, closerelatives of these genes are not found in thegenomes of non-vertebrate organisms, with twoexceptions: a stramenopile and a microalgae.The EEVS-like genes from these two algal speciesare more similar to those found in vertebratesthan those found in bacterial genomes. Withthese data in hand, the Oregon State teampropose that the gadusol pathway identified inhigher organisms could have come from a hori-zontal gene transfer event from an algal speciesto an ancestor of animals.Overall, this work illuminates a novel pathwaythat constructs an important biological sun-screen, but it also raises a number of questions.How did this biosynthetic gene cluster evolve inanimals, and why was it lost in some branches ofthe phylogenetic tree? Do the other animals thatharbor the gene cluster actually produce gadu-sol, and what role does the biological sunscreenplay in those organisms? Finally, why do marineanimals that obtain MAAs from their diet alsoproduce gadusol de novo? These lines of inquirywill likely reveal new roles for these intriguingsmall-molecule sunscreens.Carolyn A Brotherton is in the Department ofChemistry and Chemical Biology, Harvard University,Cambridge, United StatesEmily P Balskus is in the Department of Chemistry andChemical Biology, Harvard University, Cambridge,United Statesbalskus@chemistry.harvard.eduFigure 1. Many species produce small molecules thatprotect them from the sun by absorbing harmfulultraviolet radiation. The genome of the zebrafish(Danio rerio) contains a cluster of genes for a set ofproteins that can make the sunscreen compoundgadusol from sedoheptulose-7-phosphate. Microbesalso use this molecule to make sunscreen compounds,but do so via a different pathway. The cluster of genesfound in zebrafish is also present in several othervertebrates, including amphibians, reptiles and birds.FIGURE CREDIT: PHOTOGRAPH TAKEN BY KAMUJP (CC BY-SA2.0)Brotherton and Balskus. eLife 2015;4:e07961. DOI: 10.7554/eLife.07961 2 of 3Insight Biochemistry | Shedding light on sunscreen biosynthesis in zebrafishCompeting interests: The authors declare that nocompeting interests exist.Published 12 May 2015ReferencesAsamizu S, Xie P, Brumsted CJ, Flatt PM, Mahmud T.2012. Evolutionary divergence of sedoheptulose7-phosphate cyclases leads to several distinct cyclicproducts. Journal of the American Chemical Society134:12219–12229. doi: 10.1021/ja3041866.Balskus EP, Walsh CT. 2010. The genetic andmolecular basis for sunscreen biosynthesis incyanobacteria. Science 329:1653–1656. doi: 10.1126/science.1193637.Bok MJ, Porter ML, Place AR, Cronin TW. 2014.Biological sunscreens tune polychromatic ultravioletvision in mantis shrimp. Current Biology 24:1636–1642.doi: 10.1016/j.cub.2014.05.071.Chioccara F, Zeuli L, Novellino E. 1986. Occurrence ofmycosporine related compounds in sea urchin eggs.Comparative Biochemistry and Physiology Part B:Comparative Biochemistry 85:459–461. doi: 10.1016/0305-0491(86)90027-1.Plack PA, Fraser NW, Grant PT, Middleton C, MitchellAI, Thomson RH. 1981. Gadusol, an enolic derivative ofcyclohexane-1,3-dione present in the roes of cod andother marine fish. Biochemical Journal 199:741–747.Osborn A, Almabruk K, Holzwarth G, Asamizu S, LaDu J,Kean K, Karplus PA, Tanguay R, Bakalinsky A, Mahmud T.2015. De novo synthesis of a sunscreen compound invertebrates. eLife 4:e05919. doi: 10.7554/eLife.05919.Yu Y, Bai L, Minagawa K, Jian X, Li L, Li J, Chen S, CaoE, Mahmud T, Floss HG. 2005. Gene cluster responsiblefor validamycin biosynthesis in Streptomyceshygroscopicus subsp. jinggangensis 5008. Applied andEnvironmental Microbiology 71:5066–5076. doi: 10.1128/AEM.71.9.5066-5076.2005.Brotherton and Balskus. eLife 2015;4:e07961. DOI: 10.7554/eLife.07961 3 of 3Insight Biochemistry | Shedding light on sunscreen biosynthesis in zebrafish

Shedding light on sunscreen biosynthesis in zebrafish

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Shedding light on sunscreen biosynthesis in zebrafish

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