Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Dynamic diel proteome and daytime nitrogenase activity supports buoyancy in the cyanobacterium Trichodesmium

Nature Microbiology volume 7, pages 300–311 (2022)Cite this article

Subjects

Abstract

Cyanobacteria of the genus Trichodesmium provide about 80 Tg of fixed nitrogen to the surface ocean per year and contribute to marine biogeochemistry, including the sequestration of carbon dioxide. Trichodesmium fixes nitrogen in the daylight, despite the incompatibility of the nitrogenase enzyme with oxygen produced during photosynthesis. While the mechanisms protecting nitrogenase remain unclear, all proposed strategies require considerable resource investment. Here we identify a crucial benefit of daytime nitrogen fixation in Trichodesmium spp. that may counteract these costs. We analysed diel proteomes of cultured and field populations of Trichodesmium in comparison with the marine diazotroph Crocosphaera watsonii WH8501, which fixes nitrogen at night. Trichodesmium’s proteome is extraordinarily dynamic and demonstrates simultaneous photosynthesis and nitrogen fixation, resulting in balanced particulate organic carbon and particulate organic nitrogen production. Unlike Crocosphaera, which produces large quantities of glycogen as an energy store for nitrogenase, proteomic evidence is consistent with the idea that Trichodesmium reduces the need to produce glycogen by supplying energy directly to nitrogenase via soluble ferredoxin charged by the photosynthesis protein PsaC. This minimizes ballast associated with glycogen, reducing cell density and decreasing sinking velocity, thus supporting Trichodesmium’s niche as a buoyant, high-light-adapted colony forming cyanobacterium. To occupy its niche of simultaneous nitrogen fixation and photosynthesis, Trichodesmium appears to be a conspicuous consumer of iron, and has therefore developed unique iron-acquisition strategies, including the use of iron-rich dust. Particle capture by buoyant Trichodesmium colonies may increase the residence time and degradation of mineral iron in the euphotic zone. These findings describe how cellular biochemistry defines and reinforces the ecological and biogeochemical function of these keystone marine diazotrophs.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Dynamics of the Trichodesmium proteome in comparison to Crocosphaera.
Fig. 2: Nitrogenase concentration and activity over the diel cycle.
Fig. 3: Temporal dynamics of biomass composition, nitrogen fixation rates, and associated proteins in Trichodesmium.
Fig. 4: A glycogen synthesis protein emerges as a hub in the Trichodesmium proteome.
Fig. 5: Buoyancy properties of Trichodesmium and other phytoplankton.
Fig. 6: Summary of physiological properties and behaviors that are reinforced by daytime nitrogen fixation.

Similar content being viewed by others

Data availability

The mass spectrometry proteomics data has been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifiers PXD016332 and https://doi.org/10.6019/PXD016332 (laboratory experiments) and identifier PXD027796 and https://doi.org/10.6019/PXD027796 (field data). The processed proteomic data are also available at the Biological and Chemical Oceanography Data Management Office (BCO-DMO) (https://www.bco-dmo.org/dataset/783873). Source data are provided for main text Figs. 1–5 and Extended Data Figs. 1–10. Source data are provided with this paper.

Code availability

Fully reproducible code for sinking velocity calculations, statistics and plotting is available at https://github.com/naheld/Held2020_TrichoDiel.

References

  1. Zehr, J. P. & Capone, D. G. Changing perspectives in marine nitrogen fixation. Science 9514, 729 (2020).

    Google Scholar 

  2. Karl, D. et al. Dinitrogen fixation in the world’s oceans. Biogeochemistry 57–58, 47–98 (2002).

    Article  Google Scholar 

  3. Dugdale, R. & Wilkerson, F. in Primary Productivity and Biogeochemical Cycles in the Sea (eds Falkowski, P. G. et al.) 107–122 (Springer, 1992).

  4. Carpenter, E. J. & Capone, D. G. in Nitrogen in the Marine Environment 2nd edn (eds Capone, D. G., Bronk, D. A., Mulholland, M. R. & Carpenter, E. J.) Ch. 4 (Elsevier, 2008).

  5. Gruber, N. & Sarmiento, J. L. Global patterns of marine nitrogen fixation and denitrification. Global Biogeochem. Cycles 11, 23–266 (1997).

    Article  Google Scholar 

  6. Buchanan, P. J., Chase, Z., Matear, R. J., Phipps, S. J. & Bindoff, N. L. Marine nitrogen fixers mediate a low latitude pathway for atmospheric CO2 drawdown. Nat. Commun. https://doi.org/10.1038/s41467-019-12549-z (2019).

  7. Monteiro, F. M., Follows, M. J. & Dutkiewicz, S. Distribution of diverse nitrogen fixers in the global ocean. Global Biogeochem. Cycles 24, 1–16 (2010).

    Article  Google Scholar 

  8. Church, M. J., Björkman, K. M., Karl, D. M., Saito, M. A. & Zehr, J. P. Regional distributions of nitrogen-fixing bacteria in the Pacific Ocean. Limnol. Oceanogr. 53, 63–77 (2008).

    Article  CAS  Google Scholar 

  9. Monteiro, F. M., Dutkiewicz, S. & Follows, M. J. Biogeographical controls on the marine nitrogen fixers. Global Biogeochem. Cycles 25, 1–8 (2011).

    Article  Google Scholar 

  10. Dutkiewicz, S., Ward, B. A., Monteiro, F. & Follows, M. J. Interconnection of nitrogen fixers and iron in the Pacific Ocean: theory and numerical simulations. Global Biogeochem. Cycles 26, 1–16 (2012).

    Article  Google Scholar 

  11. Walworth, N. G. et al. Nutrient-colimited Trichodesmium as a nitrogen source or sink in a future ocean. Appl. Environ. Microbiol. 84, 1–14 (2018).

    Article  CAS  Google Scholar 

  12. McGillicuddy, D. J. Jr. Do Trichodesmium spp. populations in the North Atlantic export most of the nitrogen they fix? Global Biogeochem. Cycles 28, 103–114 (2014).

    Article  CAS  Google Scholar 

  13. Carpenter, E. J. & Romans, K. Major role of the cyanobacterium Trichodesmium in nutrient cycling in the North Atlantic Ocean. Science 254, 1989–1992 (1991).

    Article  Google Scholar 

  14. Bergman, B., Sandh, G., Lin, S., Larsson, J. & Carpenter, E. J. Trichodesmium – a widespread marine cyanobacterium with unusual nitrogen fixation properties. FEMS Microbiol. Rev. 37, 286–302 (2013).

    Article  CAS  PubMed  Google Scholar 

  15. Capone, D. G. Trichodesmium, a globally significant marine cyanobacterium. Science 276, 1221–1229 (1997).

    Article  CAS  Google Scholar 

  16. Gallon, J. R. The oxygen sensitivity of nitrogenase: a problem for biochemists and micro-organisms. Trends Biochem. Sci. 6, 19–23 (1981).

    Article  CAS  Google Scholar 

  17. Saito, M. A. et al. Iron conservation by reduction of metalloenzyme inventories in the marine diazotroph Crocosphaera watsonii. Proc. Natl Acad. Sci. USA 108, 2184–2189 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Dron, A. et al. Light-dark (12:12) cycle of carbon and nitrogen metabolism in Crocosphaera watsonii WH8501: relation to the cell cycle. Environ. Microbiol. 14, 967–981 (2012).

    Article  CAS  PubMed  Google Scholar 

  19. Mohr, W., Intermaggio, M. P. & LaRoche, J. Diel rhythm of nitrogen and carbon metabolism in the unicellular, diazotrophic cyanobacterium Crocosphaera watsonii WH8501. Environ. Microbiol. 12, 412–421 (2010).

    Article  CAS  PubMed  Google Scholar 

  20. Flores, E. & Herrero, A. Compartmentalized function through cell differentiation in filamentous cyanobacteria. Nat. Rev. Microbiol. 8, 39–50 (2010).

    Article  CAS  PubMed  Google Scholar 

  21. Burnat, M., Herrero, A. & Flores, E. Compartmentalized cyanophycin metabolism in the diazotrophic filaments of a heterocyst-forming cyanobacterium. Proc. Natl Acad. Sci. USA 111, 3823–3828 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Sherman, D. M., Tucker, D. & Sherman, L. A. Heterocyst development and localization of cyanophycin in N2-fixing cultures of Anabaena sp. PCC 7120 (Cyanobacteria). J. Phycol. 941, 932–941 (2000).

    Article  Google Scholar 

  23. Lamont, H. C., Silvester, W. B. & Torrey, J. G. Nile red fluorescence demonstrates lipid in the envelope of vesicles from N2-fixing cultures of Frankia. Can. J. Microbiol. 34, 656–660 (1988).

    Article  CAS  Google Scholar 

  24. Saino, T. Diel variation in nitrogen fixation by a marine blue-green alga, Trichodesmium thiebautii. Deep Sea Res. 25, 1259–1263 (1978).

    Article  Google Scholar 

  25. Saino, T. & Hattori, A. Aerobic nitrogen fixation by the marine non-heterocystous cyanobacterium Trichodesmium (Oscillatoria) spp.: its protective mechanism against oxygen. Mar. Biol. 70, 251–254 (1982).

    Article  Google Scholar 

  26. Berman-Frank, I. et al. Segregation of nitrogen fixation and oxygenic photosynthesis in the marine cyanobacterium Trichodesmium. Science 294, 1534–1537 (2001).

    Article  CAS  PubMed  Google Scholar 

  27. Ohki, K. & Taniuchi, Y. Detection of nitrogenase in individual cells of a natural population of Trichodesmium using immunocytochemical methods for fluorescent cells. J. Oceanogr. 65, 427–432 (2009).

    Article  CAS  Google Scholar 

  28. Eichner, M. et al. N2 fixation in free-floating filaments of Trichodesmium is higher than in transiently suboxic colony microenvironments. New Phytol. 222, 852–863 (2019).

    Article  CAS  PubMed  Google Scholar 

  29. Ohki, K. Intercellular localization of nitrogenase in a non-heterocystous cyanobacterium (cyanophyte), Trichodesmium sp. NIBB1067. J. Oceanogr. 64, 211–216 (2008).

    Article  CAS  Google Scholar 

  30. Ohki, K., Zehr, F. & Fujita, Y. Regulation of nitrogenase activity in relation to the light-dark regime in the filamentous non-heterocystous cyanobacterium Trichodesmium sp. NIBB 1067. J. Gen. Microbiol. 138, 2679–2685 (1992).

    Article  CAS  Google Scholar 

  31. Finzi-Hart, J. A. et al. Fixation and fate of C and N in the cyanobacterium Trichodesmium using nanometer-scale secondary ion mass spectrometry. Proc. Natl Acad. Sci. USA 106, 9931 (2009).

    Article  CAS  Google Scholar 

  32. Sandh, G., El-Shehawy, R., Díez, B. & Bergman, B. Temporal separation of cell division and diazotrophy in the marine diazotrophic cyanobacterium Trichodesmium erythraeum IMS101. FEMS Microbiol. Lett. 295, 281–288 (2009).

    Article  CAS  PubMed  Google Scholar 

  33. Küpper, H. et al. Traffic lights in Trichodesmium. Regulation of photosynthesis for nitrogen fixation studied by chlorophyll fluorescence kinetic microscopy. Plant Physiol. 135, 2120–2133 (2019).

    Article  Google Scholar 

  34. Ohki, K. & Fujita, Y. Aerobic nitrogenase activity measured as acetylene reduction in the marine non-heterocystous cyanobacterium Trichodesmium spp. grown under artificial conditions. Mar. Biol. 98, 111–114 (1988).

    Article  CAS  Google Scholar 

  35. Waterbury, J. B. & Willey, J. M. Isolation and growth of marine planktonic Cyanobacteria. Methods Enzymol. 167, 100–105 (1988).

    Article  CAS  Google Scholar 

  36. Chen, Y. B., Zehr, J. P. & Mellon, M. Growth and nitrogen fixation of the diazotrophic filamentous nonheterocystous cyanobacterium Trichodesmium sp. IMS 101 in defined media: evidence for a circadian rhythm. J. Phycol. 32, 916–923 (1996).

    Article  Google Scholar 

  37. Berman-Frank, I., Bidle, K. D., Haramaty, L. & Falkowski, P. G. The demise of the marine cyanobacterium, Trichodesmium spp., via an autocatalyzed cell death pathway. Limnol. Oceanogr. 49, 997–1005 (2004).

    Article  Google Scholar 

  38. Bell, P. R. F. et al. Laboratory culture studies of Trichodesmium isolated from the Great Barrier Reef lagoon, Australia. Hydrobiologia 532, 9–21 (2005).

    Article  Google Scholar 

  39. Tzubari, Y., Magnezi, L., Be’Er, A. & Berman-Frank, I. Iron and phosphorus deprivation induce sociality in the marine bloom-forming cyanobacterium Trichodesmium. ISME J. 12, 1682–1693 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Held, N. A., McIlvin, M. R., Moran, D. M., Laub, M. T. & Saito, M. A. Unique patterns and biogeochemical relevance of two-component sensing in marine bacteria. mSystems 4, 1–16 (2019).

    Article  Google Scholar 

  41. Aryal, U. K. & Sherman, L. A. in Cyanobacteria Omics Manipulation (ed. Los, D. A.) Ch. 6 (Caister Academic Press, 2017).

  42. Held, N. A. et al. Co-occurrence of Fe and P stress in natural populations of the marine diazotroph Trichodesmium. Biogeosciences 17, 2537–2551 (2020).

    Article  Google Scholar 

  43. Klugkist, J., Haaker, H., Wassink, H. & Veeger, C. The catalytic activity of nitrogenase in intact Azotobacter vinelandii cells. Eur. J. Biochem. 146, 509–515 (1985).

    Article  CAS  PubMed  Google Scholar 

  44. Zehr, J. P., Wyman, M., Miller, V., Capone, D. G. & Duguay, L. Modification of the Fe protein of nitrogenase in natural populations of Trichodesmium thiebautii. Appl. Environ. Microbiol. 59, 669–676 (1993).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Rodriguez, I. B. & Ho, T.-Y. Diel nitrogen fixation pattern of Trichodesmium: the interactive control of light and Ni. Sci. Rep. 4, 4445 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  46. Eichner, M., Kranz, S. A. & Rost, B. Combined effects of different CO2 levels and N sources on the diazotrophic cyanobacterium Trichodesmium. Physiol. Plant. 152, 316–330 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Hutchins, D. A. et al. Irreversibly increased nitrogen fixation in Trichodesmium experimentally adapted to elevated carbon dioxide. Nat. Commun. 6, 1–7 (2015).

    Article  Google Scholar 

  48. Levitan, O. et al. Combined effects of CO2 and light on the N2-fixing cyanobacterium Trichodesmium IMS101: a mechanistic view. Plant Physiol. 154, 346–356 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Villareal, T. A. & Carpenter, E. J. Buoyancy regulation and the potential for vertical migration in the oceanic cyanobacterium Trichodesmium. Microb. Ecol. 45, 1–10 (2003).

    Article  CAS  PubMed  Google Scholar 

  50. Rabouille, S., Staal, M., Stal, L. J. & Soetaert, K. Modeling the dynamic regulation of nitrogen fixation in the cyanobacterium Trichodesmium sp. Appl. Environ. Microbiol. 72, 3217–3227 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Breitbarth, E., Wohlers, J., Kläs, J., LaRoche, J. & Peeken, I. Nitrogen fixation and growth rates of Trichodesmium IMS-101 as a function of light intensity. Mar. Ecol. Prog. Ser. 359, 25–36 (2008).

    Article  CAS  Google Scholar 

  52. Chen, Y. B. et al. Circadian rhythm of nitrogenase gene expression in the diazotrophic filamentous nonheterocystous cyanobacterium Trichodesmium sp. strain IMS101. J. Bacteriol. 180, 3598–3605 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Rabouille, S., Staal, M., Stal, L. J. & Soetaert, K. Modeling the dynamic regulation of nitrogen fixation in the Cyanobacterium Trichodesmium sp. Appl. Environ. Microbiol. 72, 3217–3227 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Capone, D. G., O’Neill, J. M., Zehr, J. & Carpenter, E. J. Basis for diel variation in nitrogenase activity in the marine planktonic cyanobacterium Trichodesmium thiebautti. Appl. Environ. Microbiol. 56, 3532–3536 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Gründel, M., Scheunemann, R., Lockau, W. & Zilliges, Y. Impaired glycogen synthesis causes metabolic overflow reactions and affects stress responses in the cyanobacterium Synechocystis sp. PCC 6803. Microbiology 158, 3032–3043 (2012).

    Article  PubMed  Google Scholar 

  56. Jackson, S. A., Eaton-Rye, J. J., Bryant, D. A., Posewitz, M. C. & Davies, F. K. Dynamics of photosynthesis in a glycogen-deficient glgC mutant of Synechococcus sp. strain PCC 7002. Appl. Environ. Microbiol. 81, 6210–6222 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Boatman, T. G., Davey, P. A., Lawson, T. & Geider, R. J. The physiological cost of diazotrophy for Trichodesmium erythraeum IMS101. PLoS ONE 13, 1–24 (2018).

    Article  Google Scholar 

  58. Chappell, P. D., Moffett, J. W., Hynes, A. M. & Webb, E. A. Molecular evidence of iron limitation and availability in the global diazotroph Trichodesmium. ISME J. 6, 1728–1739 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Chappell, P. D. & Webb, E. A. A molecular assessment of the iron stress response in the two phylogenetic clades of Trichodesmium. Environ. Microbiol. 12, 13–27 (2010).

    Article  CAS  PubMed  Google Scholar 

  60. Walsby, A. E. The properties and buoyancy-providing role of gas vacuoles in Trichodesmium Ehrenberg. Br. Phycol. J. 13, 103–116 (1978).

    Article  Google Scholar 

  61. Villareal, T. A. & Carpenter, E. J. Diel buoyancy regulation in the marine diazotrophic cyanobacterium Trichodesmium thiebautii. Limnol. Oceanogr. 35, 1832–1837 (1990).

    Article  Google Scholar 

  62. Romans, K. M., Carpenter, E. J. & Bergman, B. Buoyancy regulation in the colonial diazotrophic cyanobacterium Trichodesmium tenue: ultrastructure and storage of carbohydrate, polyphosphate, and nitrogen. J. Phycol. 30, 935–942 (1994).

    Article  Google Scholar 

  63. Wang, L. et al. Molecular structure of glycogen in Escherichia coli. Biomacromolecules 20, 2821–2829 (2019).

    Article  CAS  PubMed  Google Scholar 

  64. Berman-Frank, I., Cullen, J. T., Shaked, Y., Sherrell, R. M. & Falkowski, P. G. Iron availability, cellular iron quotas, and nitrogen fixation in Trichodesmium. Limnol. Oceanogr. 46, 1249–1260 (2001).

    Article  CAS  Google Scholar 

  65. Kustka, A. B. et al. Iron requirements for dinitrogen- and ammonium-supported growth in cultures of Trichodesmium (IMS 101): comparison with nitrogen fixation rates and iron:carbon ratios of field populations. Limnol. Oceanogr. 49, 1224 (2004).

    Article  CAS  Google Scholar 

  66. Paerl, H. W., Prufert-Bebout, I. L. E., Guo, C. & Carolina, N. Iron-stimulated N2 fixation and growth in natural and cultured populations of the planktonic marine cyanobacteria Trichodesmium spp. Appl. Environ. Microbiol. 60, 1044–1047 (1994).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Rubin, M., Berman-Frank, I. & Shaked, Y. Dust- and mineral-iron utilization by the marine dinitrogen-fixer Trichodesmium. Nat. Geosci. 4, 529–534 (2011).

    Article  CAS  Google Scholar 

  68. Polyviou, D. et al. Desert dust as a source of iron to the globally important diazotroph Trichodesmium. Front. Microbiol. 8, 1–12 (2018).

    Article  Google Scholar 

  69. Basu, S. & Shaked, Y. Mineral iron utilization by natural and cultured Trichodesmium and associated bacteria. Limnol. Oceanogr. 63, 2307–2320 (2018).

    Article  CAS  Google Scholar 

  70. Held, N. A. et al. Mechanisms and heterogeneity of in situ mineral processing by the marine nitrogen fixer Trichodesmium revealed by single-colony metaproteomics. ISME Commun. 1, 35 (2021).

    Article  Google Scholar 

  71. Basu, S., Gledhill, M., de Beer, D., Prabhu Matondkar, S. G. & Shaked, Y. Colonies of marine cyanobacteria Trichodesmium interact with associated bacteria to acquire iron from dust. Commun. Biol. 2, 1–8 (2019).

    Article  CAS  Google Scholar 

  72. Tyrrell, T. et al. Large-scale latitudinal distribution of Trichodesmium spp. in the Atlantic Ocean. J. Plankton Res. 25, 405–416 (2003).

    Article  CAS  Google Scholar 

  73. Robson, R. L. & Postgate, J. R. Oxygen and hydrogen in biological nitrogen fixation. Annu. Rev. Microbiol. 34, 183–207 (1980).

  74. Zehr, J. P. Nitrogen fixation by marine cyanobacteria. Trends Microbiol. 19, 162–173 (2011).

    Article  CAS  PubMed  Google Scholar 

  75. Bergman, B. & Carpenter, E. J. Nitrogenase confined to randomly distributed trichomes in the marine cyanobacterium Trichodesmium thiebautii. J. Phycol. 27, 158–165 (1991).

    Article  CAS  Google Scholar 

  76. Inomura, K., Wilson, S. T. & Deutsch, C. Mechanistic model for the coexistence of nitrogen fixation and photosynthesis in marine Trichodesmium. mSystems 4, 1–13 (2019).

    Article  Google Scholar 

  77. Janson, S., Matveyev, A. & Bergman, B. The presence and expression of hetR in the non-heterocystous cyanobacterium Symploca PCC 8002. FEMS Microbiol. Lett. 168, 173–179 (1998).

    Article  CAS  PubMed  Google Scholar 

  78. Zhang, J. Y., Chen, W. L. & Zhang, C. C. hetR and patS, two genes necessary for heterocyst pattern formation, are widespread in filamentous nonheterocyst-forming cyanobacteria. Microbiology 155, 1418–1426 (2009).

    Article  CAS  PubMed  Google Scholar 

  79. Moore, J. K., Doney, S. C., Glover, D. M. & Fung, I. Y. Iron cycling and nutrient-limitation patterns in surface waters of the world ocean. Deep Sea Res. 2 Top. Stud. Oceanogr. 49, 463–507 (2001).

    Article  Google Scholar 

  80. Chisholm, S. W. in Primary Productivity and Biogeochemical Cycles in the Sea (eds Falkowski, P. G. et al.) 213–237 (Springer, 1992).https://doi.org/10.1007/978-1-4899-0762-2_12

  81. Young, K. D. The selective value of bacterial shape. Microbiol. Mol. Biol. Rev. 70, 660–703 (2006).

    Article  PubMed  PubMed Central  Google Scholar 

  82. Lu, X. & Zhu, H. Tube-gel digestion: a novel proteomic approach for high-throughput analysis of membrane proteins. Mol. Cell Proteom. 4, 1948–1958 (2005).

    Article  CAS  Google Scholar 

  83. Saito, M. A. et al. Multiple nutrient stresses at intersecting Pacific Ocean biomes detected by protein biomarkers. Science 345, 1173–1177 (2014).

    Article  CAS  PubMed  Google Scholar 

  84. McIlvin, M. R. & Saito, M. A. Online nanoflow two-dimension comprehensive active modulation reversed phase-reversed phase liquid chromatography high-resolution mass spectrometry for metaproteomics of environmental and microbiome samples. J. Proteome Res. 20, 4589–4597 (2021).

    Article  CAS  PubMed  Google Scholar 

  85. Lee, M. D. et al. Transcriptional activities of the microbial consortium living with the marine nitrogen-fixing cyanobacterium Trichodesmium reveal potential roles in community-level nitrogen cycling. Appl. Environ. Microbiol. 84, AEM.02026-17 (2017).

    Google Scholar 

  86. Zhang, Y., Wen, Z., Washburn, M. P. & Florens, L. Refinements to label-free proteome quantitation: how to deal with peptides shared by multiple proteins. Anal. Chem. 82, 2272–2281 (2010).

    Article  CAS  PubMed  Google Scholar 

  87. Gallien, S., Bourmaud, A., Kim, S. Y. & Domon, B. Technical considerations for large-scale parallel reaction monitoring analysis. J. Proteom. 100, 147–159 (2014).

    Article  CAS  Google Scholar 

  88. Pino, L. K. et al. The skyline ecosystem: informatics for quantitative mass spectrometry proteomics. Mass Spectrom. Rev. 176, 139–148 (2019).

    Google Scholar 

  89. Held, N. A. et al. Mechanisms and heterogeneity of in situ mineral processing by the marine nitrogen fixer Trichodesmium revealed by single-colony metaproteomics. ISME Commun. https://doi.org/10.1038/s43705-021-00034-y (2021).

  90. White, A. E., Spitz, Y. H. & Letelier, R. M. Modeling carbohydrate ballasting by Trichodesmium spp. Mar. Ecol. Prog. Ser. 323, 35–45 (2006).

    Article  Google Scholar 

  91. Morrison, F. A. An Introduction to Fluid Mechanics (Cambridge Univ. Press, 2013).

  92. Hunter, J. D. Matplotlib: a 2D graphics environment. Comput. Sci. Eng. 9, 90–95 (2007).

    Article  Google Scholar 

  93. Virtanen, P. et al. SciPy 1.0: fundamental algorithms for scientific computing in Python. Nat. Methods 17, 261–272 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  94. Hagberg, A. A., Schult, D. A. & Swart, P. J. Exploring network structure, dynamics, and function using NetworkX. In 7th Python Scientific Conference (SciPy 2008) 11–15 (2008).

Download references

Acknowledgements

This work was supported by NSF Graduate Research Fellowship grant 1122274 (N.A.H.), Gordon and Betty Moore Foundation grant GBMF-3782 (M.A.S.), National Science Foundation grants OCE-1657766, OCE-1850719 and OCE-1924554 (M.A.S.), National Institutes of Health grant GM135709-02 (M.A.S.), and the Woods Hole Oceanographic Institution Ocean Ventures Fund (N.A.H.). N.A.H. was additionally supported by Principles of Microbial Ecosystems collaboration of the Simons Foundation (grant ID 542379). We thank the scientific staff and crew of the AT39-05/Tricolim research expedition, particularly chief scientist D. Hutchins, and the JC150/Ziploc expedition, particularly chief scientist C. Mahaffey. Special thanks to B. White.

Author information

Author notes

  1. Noelle A. Held

    Present address: Department of Environmental Systems Science, ETH, Zurich, Switzerland

  2. Kevin M. Sutherland

    Present address: Department of Earth and Planetary Sciences, Harvard University, Cambridge, MA, USA

Authors and Affiliations

  1. Marine Chemistry and Geochemistry Department, Woods Hole Oceanographic Institution, Woods Hole, MA, USA

    Noelle A. Held, Riss M. Kellogg, Matthew R. McIlvin, Dawn M. Moran, Kevin M. Sutherland & Mak A. Saito

  2. Department of Earth, Atmospheric and Planetary Science, Massachusetts Institute of Technology, Cambridge, MA, USA

    Noelle A. Held, Riss M. Kellogg & Kevin M. Sutherland

  3. Department of Biology, Woods Hole Oceanographic Institution, Woods Hole, MA, USA

    John B. Waterbury & Frederica W. Valois

  4. Marine and Environmental Biology, Department of Biological Sciences, University of Southern California, Los Angeles, CA, USA

    Eric A. Webb

  5. Department of Applied Ocean Physics and Engineering, Woods Hole Oceanographic Institution, Woods Hole, MA, USA

    Michael Jakuba

Authors
  1. Noelle A. Held
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. John B. Waterbury
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Eric A. Webb
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Riss M. Kellogg
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Matthew R. McIlvin
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Michael Jakuba
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Frederica W. Valois
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Dawn M. Moran
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Kevin M. Sutherland
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Mak A. Saito
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

N.A.H., J.B.W. and M.A.S. conceived and designed the experiment. N.A.H., R.M.K, D.M.M. and F.W.V. performed the experiments and analyses. M.R.M. and N.A.H. performed mass spectrometry analyses. M.J., N.A.H. and M.A.S. developed sinking velocity calculations. K.M.S. performed synchrotron element map analyses. All authors contributed to the writing and editing of the manuscript.

Corresponding author

Correspondence to Mak A. Saito.

Ethics declarations

Competing interests

The authors declare no competing interests.

Peer review information

Nature Microbiology thanks Ilka Axmann, David Karl, Adam Kustka and Marc Strous for their contribution to the peer review of this work.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Extended Data Fig. 1 Total spectral counts and average protein content across the diel cycle.

a) Average total spectral counts (peptide to spectrum matches) with error bar representing +/- one standard deviation, at each time point. Each data point is also shown individually as black scatter points. Yellow and indigo bars indicate the light and dark periods, respectively. Total spectral counts were relatively uniform and do not vary systematically throughout the diel cycle, implying consistency in the proteome analyses. b) Total protein content in the culture shown with error bar representing +/- one standard deviation, for biological duplicates after protein precipitation and purification, measured by a colorimetric assay. Higher protein abundances at night may suggest nighttime cell growth. Again, each data point is also shown individually as black scatter points. Yellow and indigo bars indicate the light and dark periods, respectively.

Source data

Extended Data Fig. 2 Dynamics of the entire proteome of Trichodesmium erythraeum sp. IMS101 over the diel cycle.

The dynamic range of the normalized spectral count data can be observed, as well as fluctuations in protein abundance occurring throughout the experiment.

Source data

Extended Data Fig. 3 Proteome dynamics of separate replicate laboratory experiment over the diel cycle.

a) Clustered heatmap of a replicate diel experiment conducted one year prior to the main experiment, under the same experimental conditions. Protein abundances were summed for each KO module and normalized across each row. b) Dynamics of the proteome clusters over the diel cycle, with each KO module represented as a line and colored based on the clustering in panel (A). Rapid oscillations of the proteome and clustering of the nitrogenase/nitrogen metabolism proteins with the photosystems are similar in the main experiment. Yellow and dark purple bars indicate the light and dark periods, respectively.

Source data

Extended Data Fig. 4 Proteome dynamics of field Trichodesmium population over the diel cycle.

a) Clustered heatmap of the proteome of a field Trichodesmium population sampled in situ over a diel cycle. Protein abundances were summed for each KO module and normalized across each row. b) Dynamics of the proteome clusters over the diel cycle, with each KO module represented as a line and colored based on the clustering in panel (A). Though the sampling was lower resolution than in the laboratory experiments, the rapid oscillations of the proteome are reproduced. Yellow and dark purple bars indicate the light and dark periods, respectively.

Source data

Extended Data Fig. 5 In vivo specific activity of the nitrogenase NifH protein over the diel cycle for Crocosphaera watsonii.

In vivo specific activity of the nitrogenase NifH protein (nmol ethyelene produced per min per mg NifH) over the diel cycle for Crocosphaera watsonii34. Unlike in Trichodesmium which exhibits significant variability in nitrogenase activity throughout the diel cycle, in Crocosphaera nitrogenase is either not present or highly present and very active.

Source data

Extended Data Fig. 6 POC content versus total protein spectral counts.

POC content versus total protein spectral counts in the main laboratory experiment. These are weakly correlated suggesting that POC content is driven mainly by carbohydrate content, not protein abundance.

Source data

Extended Data Fig. 7 Glycogen content of Trichodesmium populations sampled in situ by depth.

The populations were sampled on August 7, 2017 at 31°W 22°N in the early morning. Error bars are standard deviations of the mean value of the biological triplicates, and corresponding data points are plotted in grey circles. For each depth, n = 3 samples collected from replicate phytoplankton net sampling events, n = 2 samples for depth = 160 m.

Source data

Extended Data Fig. 8 Glycogen content of Trichodesmium colonies separated by morphology.

Glycogen content of Trichodesmium colonies sampled in situ at the surface and separated by morphology. The populations were sampled from the surface on March 10, 2018 at 65 22.420 °W 17 0.284 °N and separated by morphology at the time of picking.

Source data

Extended Data Fig. 9 Synchrotron-based element maps used to determine mass of particulate iron associated with a puff-type colony.

Synchrotron-based element maps used to determine mass of particulate iron associated with a puff-type colony, data originally collected as in Held et al., 202020. The left image is the X-ray fluorescence-based concentration, the middle image represents pixels with sufficiently high Fe to be considered a particle, and the right image is the product of the left and middle images. The total particulate Fe was determined as the area integrated Fe of the right image. The scale bar represents 180 microns. As detailed in Held et al., 2021, five Trichodesmium colonies of differing morphologies and degrees of particle association were examined in this way. These images are representative of a Trichodesmium colony with average-to-high particle loading.

Extended Data Fig. 10 Calibration curves for 15N labeled standard peptides used for absolute quantitation of the nitrogenase proteins.

Precursor ion intensities were linearly correlated with analyzed peptide concentrations between 0-10 fmol μL−1.

Source data

Supplementary information

Supplementary Information

Supplementary Discussion, Table 1, Box 1 and Figs. 1–5.

Reporting Summary

Supplementary Table

Supplementary Tables 2–9.

Peer Review File

Source data

Source Data Fig. 1

Diel proteome data of Trichodesmium (main laboratory experiment) and Crocosphaera with annotated KO modules.

Source Data Fig. 2

Quantitative data for nitrogenase proteins and specific activity data for NifH.

Source Data Fig. 3

Selected protein and physiological data from main laboratory experiment.

Source Data Fig. 4

Spearmann correlation statistics for significant positive and negative correlations between protein pairs, and selected protein data used to generate the network.

Source Data Fig. 5

Modelling input and output data; Trichodesmium and Crocosphaera glycogen content over the diel cycle.

Source Data Extended Data Fig. 1

Total spectral counts and total protein content for samples in main laboratory experiment.

Source Data Extended Data Fig. 2

Global proteome data for main laboratory experiment.

Source Data Extended Data Fig. 3

Global proteome data organized into KO modules for separate replicate replicate experiment.

Source Data Extended Data Fig. 4

Global proteome data organized into KO modules for field samples.

Source Data Extended Data Fig. 5

NifH specific activity for Crocosphaera watsonii.

Source Data Extended Data Fig. 6

Total spectrum counts and POC content for samples in main laboratory experiment.

Source Data Extended Data Fig. 7

Glycogen content data for field samples at different depths.

Source Data Extended Data Fig. 8

Glycogen content for field samples separated by morphology.

Source Data Extended Data Fig. 10

Calibration data for 15N-labelled peptide standards.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Held, N.A., Waterbury, J.B., Webb, E.A. et al. Dynamic diel proteome and daytime nitrogenase activity supports buoyancy in the cyanobacterium Trichodesmium. Nat Microbiol 7, 300–311 (2022). https://doi.org/10.1038/s41564-021-01028-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41564-021-01028-1

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Translational Research

Sign up for the Nature Briefing: Translational Research newsletter — top stories in biotechnology, drug discovery and pharma.

Get what matters in translational research, free to your inbox weekly. Sign up for Nature Briefing: Translational Research