Photosynthetica 2023, 61(4):417-424 | DOI: 10.32615/ps.2023.028

Single-cell microfluidics in combination with chlorophyll a fluorescence measurements to assess the lifetime of the Chlamydomonas PSBO protein

E. SZÉLES1, 2, S. KUNTAM1, A. VIDAL-MEIRELES1, V. NAGY1, K. NAGY3, Á. ÁBRAHÁM3, 4, L. KOVÁCS1, S.Z. TÓTH1
1 Institute of Plant Biology, Biological Research Centre, Szeged, H-6726 Szeged, Hungary
2 Doctoral School of Biology, University of Szeged, H-6722 Szeged, Hungary
3 Institute of Biophysics, Biological Research Centre, Szeged, H-6726 Szeged, Hungary
4 Doctoral School of Multidisciplinary Medical Sciences, University of Szeged, H-6720 Szeged, Hungary

PSBO is an essential subunit of the oxygen-evolving complex and we recently demonstrated that its lifetime depends on environmental conditions in Chlamydomonas reinhardtii. To assess PSBO lifetime with a high time resolution, we employed (1) a microfluidic platform enabling the trapping of single cells and the parallel measurement of photosynthetic activity, and (2) a nitrate-inducible PSBO amiRNA line. Our microfluidic platform allowed the rapid replacement of the nutrient solution necessary for induction. It also enabled the precise monitoring of the decline in the Fv/Fm value, reflecting PSBO loss. We found that in the dark, at medium and high light intensity, the Fv/Fm value decreased with halftimes of about 25, 12.5, and 5 h, respectively. We also observed that photosynthetic activity was better sustained upon carbon limitation. In the absence of acetate, the halftimes of Fv/Fm diminishment doubled to quadrupled compared with the control, acetate-supplied cultures.

Additional key words: carbon availability; chlorophyll fluorescence; oxygen-evolving complex; photoinhibition; protein lifetime.

Received: June 14, 2023; Revised: July 6, 2023; Accepted: July 11, 2023; Prepublished online: July 31, 2023; Published: December 19, 2023  Show citation

ACS AIP APA ASA Harvard Chicago Chicago Notes IEEE ISO690 MLA NLM Turabian Vancouver
SZÉLES, E., KUNTAM, S., VIDAL-MEIRELES, A., NAGY, V., NAGY, K., ÁBRAHÁM, Á., KOVÁCS, L., & TÓTH, S.Z. (2023). Single-cell microfluidics in combination with chlorophyll a fluorescence measurements to assess the lifetime of the Chlamydomonas PSBO protein. Photosynthetica, 61(SPECIAL ISSUE 2023-2), 417-424. doi: 10.32615/ps.2023.028
Share...
Download citation
  • BibTeX (.bib)
  • Bookends (.ris)
  • EasyBib (.ris)
  • EndNote (.enw)
  • EndNote 8 (.xml)
  • ISI WoS (.isi)
  • Medlars (.medlars)
  • Mendeley (.ris)
  • MODS (.xml)
  • Papers (.ris)
  • RefWorks (.txt)
  • RefManager (.ris)
  • RIS (.ris)
  • MS Word (.xml)
  • Zotero (.ris)
    • Open full article

    References

    1. Allahverdiyeva Y., Mamedov F., Holmström M. et al.: Comparison of the electron transport properties of the psbo1 and psbo2 mutants of Arabidopsis thaliana. - BBA-Bioenergetics 1787: 1230-1237, 2009. Go to original source...
    2. Barber R.W., Emerson D.R.: Optimal design of microfluidic networks using biologically inspired principles. - Microfluid. Nanofluid. 4: 179-191, 2008. Go to original source...
    3. Bricker T.M., Roose J.L., Fagerlund R.D. et al.: The extrinsic proteins of Photosystem II. - BBA-Bioenergetics 1817: 121-142, 2012. Go to original source...
    4. Chen G.-X., Blubaugh D.J., Homann P.H. et al.: Superoxide contributes to the rapid inactivation of specific secondary donors of the photosystem II reaction-center during photodamage of manganese-depleted photosystem II membranes. - Biochemistry 34: 2317-2332, 1995. Go to original source...
    5. Hashimoto A., Yamamoto Y., Theg S.M.: Unassembled subunits of the photosynthetic oxygen-evolving complex present in the thylakoid lumen are long-lived and assembly-competent. - FEBS Lett. 391: 29-34, 1996. Go to original source...
    6. Heifetz P.B., Förster B., Osmond C.B. et al.: Effects of acetate on facultative autotrophy in Chlamydomonas reinhardtii assessed by photosynthetic measurements and stable isotope analyses. - Plant Physiol. 122: 1439-1446, 2000. Go to original source...
    7. Ho F.M., Styring S.: Access channels and methanol binding site to the CaMn4-cluster in photosystem II based on solvent accessibility simulations, with implications for substrate water access. - BBA-Bioenergetics 1777: 140-153, 2008. Go to original source...
    8. Ifuku K., Noguchi T.: Structural coupling of extrinsic proteins with the oxygen-evolving center in photosystem II. - Front. Plant Sci. 7: 84, 2016. Go to original source...
    9. Jegerschöld C., Styring S.: Spectroscopic characterization of intermediate steps involved in donor-side-induced photoinhibition of photosystem II. - Biochemistry 35: 7794-7801, 1996. Go to original source...
    10. Kim L., Toh Y., Voldman J., Yu H.: A practical guide to microfluidic perfusion culture of adherent mammalian cells. -Lab Chip 7: 681-694, 2007. Go to original source...
    11. Kosourov S., Jokel M., Aro E.-M., Allahverdiyeva Y.: A new approach for sustained and efficient H2 photoproduction by Chlamydomonas reinhardtii. - Energy Environ. Sci. 11: 1431-1436, 2018. Go to original source...
    12. Liu H., Frankel L.K., Bricker T.M.: Functional complementation of the Arabidopsis thaliana psbo1 mutant phenotype with an N-terminally His6-tagged PsbO-1 protein in photosystem II. - BBA-Bioenergetics 1787: 1029-1038, 2009. Go to original source...
    13. López-Otín C., Galluzzi L., Freije J.M.P. et al.: Metabolic control of longevity. - Cell 166: 802-821, 2016. Go to original source...
    14. Lundin B., Hansson M., Schoefs B. et al.: The Arabidopsis PsbO2 protein regulates dephosphorylation and turnover of the photosystem II reaction centre D1 protein. - Plant J. 49: 528-539, 2007. Go to original source...
    15. Mayfield S., Bennoum P., Rochaix J.-D.: Expression of the nuclear-encoded OEE1 proteins is required for oxygen evolution and stability of photosystem II particles in Chlamydomonas reinhardtii. - EMBO J. 6: 313-318, 1987. Go to original source...
    16. Nagy V., Podmaniczki A., Vidal-Meireles A. et al.: Thin cell layer cultures of Chlamydomonas reinhardtii L159I-N230Y, pgrl1 and pgr5 mutants perform enhanced hydrogen production at sunlight intensity. - Bioresource Technol. 333: 125217, 2021. Go to original source...
    17. Nagy V., Vidal-Meireles A., Podmaniczki A. et al.: The mechanism of photosystem-II inactivation during sulphur deprivation-induced H2 production in Chlamydomonas reinhardtii. - Plant J. 94: 548-561, 2018. Go to original source...
    18. Offenbacher A.R., Polander B.C., Barry B.A.: An intrinsically disordered photosystem II subunit, PsbO, provides a structural template and a sensor of the hydrogen-bonding network in photosynthetic water oxidation. - J. Biol. Chem. 288: 29056-29068, 2013. Go to original source...
    19. Pigolev A.V., Klimov V.V.: The green alga Chlamydomonas reinhardtii as a tool for in vivo study of site-directed mutations in PsbO protein of photosystem II. - Biochemistry-Moscow 80: 662-673, 2015. Go to original source...
    20. Podmaniczki A., Nagy V., Vidal-Meireles A. et al.: Ascorbate inactivates the oxygen-evolving complex in prolonged darkness. - Physiol. Plantarum 171: 232-245, 2021. Go to original source...
    21. Popelkova H., Boswell N., Yocum C.: Probing the topography of the photosystem II oxygen evolving complex: PsbO is required for efficient calcium protection of the manganese cluster against dark-inhibition by an artificial reductant. - Photosynth. Res. 110: 111-121, 2011. Go to original source...
    22. Porra R.J., Thompson W.A., Kriedemann P.E.: Determination of accurate extinction coefficients and simultaneous equations for essaying chlorophylls a and b with four different solvents: verification of the concentration of chlorophyll standards by atomic absorption spectroscopy. - BBA-Bioenergetics 975: 384-394, 1989. Go to original source...
    23. Roach T., Sedoud A., Krieger-Liszkay A.: Acetate in mixotrophic growth medium affects photosystem II in Chlamydomonas reinhardtii and protects against photoinhibition. - BBA-Bioenergetics 1827: 1183-1190, 2013. Go to original source...
    24. Roose J.L., Frankel L.K., Mummadisetti M.P., Bricker T.M.: The extrinsic proteins of photosystem II: update. - Planta 243: 889-908, 2016. Go to original source...
    25. Sampaio-Marques B., Burhans W.C., Ludovico P.: Yeast at the forefront of research on ageing and age-related diseases. - In: Sá-Correia I. (ed): Yeasts in Biotechnology and Human Health. Progress in Molecular and Subcellular Biology. Vol. 58. Pp. 217-242. Springer, Cham 2019. Go to original source...
    26. Schmollinger S., Strenkert D., Schroda M.: An inducible artificial microRNA system for Chlamydomonas reinhardtii confirms a key role for heat shock factor 1 in regulating thermo­tolerance. - Curr. Genetics 56: 383-389, 2010. Go to original source...
    27. Song K., Li G., Zu X. et al.: The fabrication and application mechanism of microfluidic systems for high throughput biomedical screening: a review. - Micromachines 11: 297, 2020. Go to original source...
    28. Széles E., Nagy K., Ábrahám Á. et al.: Microfluidic platforms designed for morphological and photosynthetic investigations of Chlamydomonas reinhardtii on a single-cell level. - Cells 11: 285, 2022. Go to original source...
    29. Tóth S.Z., Nagy V., Puthur J.T. et al.: The physiological role of ascorbate as photosystem II electron donor: protection against photoinactivation in heat-stressed leaves. - Plant Physiol. 156: 382-392, 2011. Go to original source...
    30. Vajravel S., Sirin S., Kosourov S., Allahverdiyeva Y.: Towards sustainable ethylene production with cyanobacterial artificial biofilms. - Green Chem. 22: 6404-6414, 2020. Go to original source...
    31. Vidal-Meireles A., Kuntam S., Széles E. et al.: The lifetime of the oxygen-evolving complex subunit PSBO depends on light intensity and carbon availability in Chlamydomonas. - Plant Cell Environ. 46: 422-439, 2023. Go to original source...
    32. Vinyard D.J., Brudvig G.W.: Progress toward a molecular mechanism of water oxidation in photosystem II. - Annu. Rev. Phys. Chem. 68: 101-116, 2017. Go to original source...
    33. Whitesides G.M., Ostuni E., Takayama S. et al.: Soft lithography in biology and chemistry. - Annu. Rev. Biomed. Eng. 3: 335-373, 2001. Go to original source...
    34. Zamzam G., Lee C.W.J., Milne F. et al.: Live long and prosper: Acetate and its effects on longevity in batch culturing of Chlamydomonas reinhardtii. - Algal Res. 64: 102676, 2022. Go to original source...

    Return to the content