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Isolation of state transition mutants of Chlamydomonas reinhardtii by fluorescence video imaging

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Abstract

Oxygenic photosynthetic organisms adapt to varying light conditions by changing the distribution of light energy between Photosystem II (PS II) and photosystem I (PS I) during so-called state transitions. To identify the genes involved in this process, we have exploited a simple chlorophyll fluorescence video-imaging technique to screen a library of nuclear mutants of Chlamydomonas reinhardtii for colonies grown on agar plates that are disturbed in their ability to regulate light energy distribution between PS I and PS II. Subsequent modulated fluorescence measurements at room temperature and 77 K fluorescence emission spectra confirmed that 5 mutants (0.025% of total number screened) were defective in state transitions. [32P]orthophosphate phosphorylation experiments in vivo revealed that in one of these mutants, designated stm1, the level of LHC II polypeptide phosphorylation was drastically reduced compared with wild type. Despite WT levels of PS I and PS II, stm1 grew photoautotrophically at reduced rates, compared with WT especially under low light conditions, which is consistent with an important physiological role for state transitions. Our results highlight the feasibility of video imaging in tandem with mutagenesis as a means of identifying the genes involved in controlling state transitions in eukaryotic photosynthetic organisms.

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References

  • Allen JF (1992) Protein phosphorylation in regulation of photosynthesis. Biochim Biophys Acta 1098: 275–335

    PubMed  Google Scholar 

  • Allen JF (1995) Thylakoid protein phosphorylation, state 1-state 2 transitions, and Photosystem stoichiometry adjustment: Redox control at multiple levels of gene expression. Physiol Plant 93: 196–205

    Article  Google Scholar 

  • Allen JF and Holmes NG (1986) A general model for regulation of photosynthetic unit function by protein phosphorylation. FEBS Lett 202: 175–181

    Article  Google Scholar 

  • Allen, JF, Bennett J, Steinback KE and Arntzen CJ (1981) Chloroplast protein phosphorylation couples plastoquinone redox state to distribution of excitation energy between photosystems. Nature 291: 21–25

    Google Scholar 

  • Allen JF, Mullineaux CW, Sanders CE and Melis A (1989) State transitions, Photosystem stoichiometry adjustment and non-photochemical quenching in cyanobacterial cells acclimated to light absorbed by Photosystem I or Photosystem II. Photosynth Res 22: 157–166

    Google Scholar 

  • Allen KD and Staehelin LA (1994) Polypeptide composition, assembly and phosphorylation patterns of the Photosystem II system of Chlamydomonas reinhardtii. Planta 194: 42–54

    Google Scholar 

  • Andronis C, Kruse O, Deak Z, Vass I, Diner BA and Nixon PJ (1998) Mutation of residue threonine-2 of the D2 polypeptide and its effect on Photosystem II function in Chlamydomonas reinhardtii. Plant Physiol 117(2): 515–524

    PubMed  Google Scholar 

  • Aro E-M, Kettunen R and Tyystjärvi E (1992) FEBS Lett 297: 29–33

    PubMed  Google Scholar 

  • Bassi R and Wollman F-A (1991) The chlorophyll-a/b proteins of Photosystem II in Chlamydomonas reinhardtii. Planta 183: 423–433

    Article  Google Scholar 

  • Bennett J (1977) Phosphorylation of chloroplasts membrane polypeptides. Nature 269: 344–346

    Google Scholar 

  • Bennett J (1991) Protein-phosphorylation in green plant chloroplasts. Ann Rev Plant Mol Biol 42: 281–311

    Google Scholar 

  • Bennett J, Shaw EK and Michel H (1988) Cytochrome b6f complex is required for phosphorylation of light-harvesting chlorophyll a/b complex II in chloroplast photosynthetic membranes. Eur J Biochem 171: 95–100

    PubMed  Google Scholar 

  • Boichenko VA (1998) Photosynthetic generation of O2 and H2 by Photosystem I-deficient Chlamydomonas mutants. Biochemistry-Moscow 63(2): 164–170

    PubMed  Google Scholar 

  • Boichenko VA and Hoffmann P (1994) Photosynthetic hydrogenproduction in prokaryotes and eukaryotes – occurrence, mechanism and functions. Photosynthetica 30(4): 527–552

    Google Scholar 

  • Carlberg I and Andersson B (1996) Phosphatase activities in spinach thylakoid membranes – effectors, regulation and location. Photosynth Res 47: 145–156

    Google Scholar 

  • Coughlan SJ and Hind G (1986) Purification and characterisation of a membrane-bound protein kinase from spinach thylakoids. J Biol Chem 261: 11378–11385

    PubMed  Google Scholar 

  • Debuchy R, Purton S and Rochaix JD (1989) The arginine succinate lyase gene of Chlamydomonas reinhardtii: An important tool for nuclear transformation and for correlating the genetic and molecular maps of the ARG7 locus. EMBO J 8: 2803–2809

    PubMed  Google Scholar 

  • Diner BA and Wollman F-A (1980) Isolation of highly active Photosystem II particles from a mutant of Chlamydomonas reinhardtii. Eur J Biochem 110: 521–526

    PubMed  Google Scholar 

  • Ehlich TD, Edelman M and Mattoo AK (1992) Identification, characterization and resolution of the in vivo phosphorylated form of the D1 Photosystem II reaction center. Protein. J Biol Chem 267: 3523–2529

    Google Scholar 

  • Emlyn-Jones D, Ashby MK and Mullineaux CW (1999) A gene required for the regulation of photosynthetic light harvesting in cyanobacteria (submitted)

  • Farchaus J, Dilley RA and Cramer WA (1985) Selective inhibition of the spinach thylakoid LHC II protein kinase. Biochim Biophys Acta 809: 17–26

    Google Scholar 

  • Gal A, Herrmann RG, Lottspeich F and Ohad I (1992) Phosphorylation of cytochrome b6 by the LHC II kinase associated with the cytochrome complex. FEBS Lett 298(1): 33–35

    PubMed  Google Scholar 

  • Garnier J, Wu B, Maroc J, Guyon D and Trémolières A (1990) Restoration of both an oligomeric form of the light-harvesting antenna CPII and a fluorescence state II-state I transition by Δ 3-trans-hexadecanoic acid-containing phosphatidylglycerol, in cells of a mutant of Chlamydomonas reinhardtii. Biochim Biophys Acta 1020: 153–162

    Google Scholar 

  • Gans P and Wollman F-A (1995) The effect of cyanide on state transitions in Chlamydomonas reinhardtii. Biochim Biophys Acta 1228: 51–57

    Google Scholar 

  • Gumpel NJ and Purton S (1994) Playing tag with Chlamydomonas. Trends Cell Biol 4: 299–301

    PubMed  Google Scholar 

  • Harris E (1989) The Chlamydomonas Sourcebook: A Comprehensive Guide to Biology and Laboratory Use. Academic Press, San Diego, CA

    Google Scholar 

  • Holzwarth AR (1986) Fluorescence lifetimes in photosynthetic systems. Photochem Photobiol 43: 707–725

    Google Scholar 

  • Kindle KL (1990) High frequency nuclear transformation of Chlamydomonas-reinhardtii. Proc Natl Acad Sci USA 87(3): 1228–1232

    PubMed  Google Scholar 

  • Krebs EG (1994) The growth of research on protein phosphorylation. TIBS 19: 439–440

    PubMed  Google Scholar 

  • Kruse O, Zheleva D and Barber J (1997) Stabilization of photosystem two dimers by phosphorylation: Implication for the regulation of the turnover of D1 protein. FEBS Lett 408(3): 276–280

    PubMed  Google Scholar 

  • Michel HP and Bennett J (1987) Identification of the phosphorylation site of an 8.3 kda protein from Photosystem II of spinach. FEBS Lett 212: 103–108

    Article  Google Scholar 

  • Michel HP, Hunt DF, Shabanowitz J and Bennett J (1988) Tandem Mass spectrometry reveals that three Photosystem II proteins of spinach chloroplasts contain N-acetyl-O-phosphothreonine at their NH2 termini. J Biol Chem 25: 1123–1130

    Google Scholar 

  • Purton S and Rochaix JD (1995) Characterisation of the Arg7 gene of Chlamydomonas reinhardtii and its application to nuclear transformation. Eur J Phycol 30: 141–148

    Google Scholar 

  • Rochaix JD (1995) Chlamydomonas reinhardtii as the photosynthetic yeast. Annu Rev Genet 29: 209–230

    PubMed  Google Scholar 

  • Schuster G, Dewit M, Staehelin L and Ohad I (1986) Transient inactivation of the thylakoid Photosystem II light-harvesting protein kinase and concomitant changes in intramembrane particle sizes during photoinhibition of Chlamydomonas reinhardtii. J Cell Biol 103: 71–80

    PubMed  Google Scholar 

  • Telfer A, Allen JF, Barber J and Bennett J (1983) Thylakoid protein phosphorylation during state 1-state 2 transitions in osmotically shocked pea chloroplasts. Biochim Biophys Acta 722: 176–181

    Google Scholar 

  • Wollman FA and Lemaire C (1988) Studies on kinase-controlled state transitions in Photosystem II and b 6f mutants from Chlamydomonas reinhardtii which lack quinone-binding proteins. Biochim Biophys Acta 933: 85–94.

    Google Scholar 

  • Zhang L, Niyogi KK, Baroli I, Nemson JA, Grossman AR and Melis A (1997) DNA insertional mutagenesis for the elucidation of a Photosystem II repair process in the green alga Chlamydomonas reinhardtii. Photosynth Res 53: 173–184

    Google Scholar 

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Kruse, O., Nixon, P.J., Schmid, G.H. et al. Isolation of state transition mutants of Chlamydomonas reinhardtii by fluorescence video imaging. Photosynthesis Research 61, 43–51 (1999). https://doi.org/10.1023/A:1006229308606

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  • DOI: https://doi.org/10.1023/A:1006229308606

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