Abstract
To study the significance of Photosystem (PS) II phosphorylation for the turnover of the D1 protein, phosphorylation was compared with the synthesis and content of the D1 protein in intact chloroplasts. As shown by radioactive labelling with [32Pi] phosphorylation of PS II polypeptides was saturated at light intensities of 125 mol m-2 s-1. Under steady state conditions, in intact chloroplasts D1 protein, once it was phosphorylated, was neither dephosphorylated nor degraded in the light. D1 protein-synthesis was measured as incorporation of [14C] leucine. As shown by non-denaturing gel-electrophoresis followed by SDS-PAGE newly synthesised D1 protein was assembled to intact PS II-centres and no free D1 protein could be detected. D1 protein-synthesis was saturated at light intensities of 500 mol m-2 s-1. The content of D1 protein stayed stable even after illumination with 5000 μmol m-2 s-1 showing that D1 protein-degradation was saturated at the same light intensities. The difference in the light saturation points of phosphorylation and of D1 protein-turnover indicates a complex regulation of D1 protein-turnover by phosphorylation. Separation of the phosphorylated and dephosphorylated D1 protein by LiDS-gelelectrophoresis combined with radioactive pulse-labelling with [14C] leucine and [32Pi] revealed that D1 protein, synthesised under steady state conditions in the light, did not become phosphorylated but instead was rapidly degraded whereas the phosphorylated form of the D1 protein was not a good substrate for degradation. According to these observations phosphorylation of the D1 protein creates a pool of PS II centres which is not involved in D1 to these observations phosphorylation of the D1 protein creates a pool of PS II centres which is not involved in D1 protein-turnover. Fractionation of thylakoid membranes confirms that the phosphorylated, non-turning over pool of PS II-centres was located in the central regions of the grana, whereas PS II-centres involved in D1 protein-turnover were found exclusively in the stroma-lamellae and in the grana-margins.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Abbreviations
- chl:
-
chlorophyll
- Fv :
-
yield of variable fluorescence, difference between Fm, the maximal fluorescence yield at saturating light, when all reaction-centres are closed, and Fo, the fluorescence yield in the dark, when all reaction-centres are open
- LHC:
-
light harvesting complex
- PFD:
-
photon flux density
- PS:
-
photosystem
References
Adir N, Shochat S and Ohad I (1990) Light-dependent D1 proteinsynthesis and translocationis regulated by reaction-centre II. J Biol Chem 265: 12563–12568.
Allen JF (1992) Protein-phosphorylation in regulation of photosynthesis. Biochim BiophysAActa 1098: 275–335.
Anderson JM and Aro EM (1994) Grana stacking and protection of Photosystem II in thylakoid membranes of higher plants leaves under sustained high irradiance: Hypothesis. Photosynth Res 41: 315–326.
Arnon DI (1949) Copper enzymes in isolated chloroplasts. Polyphenol oxidase in Beta vulgaris. Plant Physiol 24: 1–5.
Aro EM, Kettunen R and Tyystjiirvi E (1992) ATP and light regulate D1 protein modification and turnover. Role of D1 in photoinhibition. FEBS Lett 297: 29–33.
Aro EM, Virgin I and Andersson B (1993) Photoinhibition of Photosystem II-inactivation, protein damage and turnover. Biochim Biophys Acta 1143: 113–134.
Aro EM, McCaffery S and Anderson JM (1994) Recovery from photoinhibition in peas acclimated to varying growth irradiances: Role of D1 protein-turnover. Plant Physiol 104: 1033–1041.
Barbato R, Friso G, Rigoni F, Dalla Vecchia F and Giacometti G (1992) Structural changes and lateral redistribution of Photosystem II during donor side photoinhibition of thylakoids. J Cell Biol 119: 325–335.
Bennett J (1991) Protein-phosphorylation in green plant chloroplasts. Annu Rev Plant Physiol Plant Mol Biol 42: 281–311.
Berthold DA, Babcock GT and Yoccum CF (1981) A highly resolved, oxygen-evolving Photosystem II preparation from spinach thylakoid membranes. FEBS Lett 134: 231–234.
Callahan FE, Ghirardi ML, Sopory SK, Metha AM, Edelman M and Mattoo AK (1990) A novel metabolic form of the 32 kDa-D1 protein in the grana-localized reaction center of Photosystem II. J Biol Chem 265: 15357–15360.
Ebbert V and Godde D (1994) Regulation of thylakoid proteinphosphorylation in intact chloroplasts by the activity of kinases and phosphatases. Biochim Biophys Acta 1187: 335–346.
Elich TD, Edelman M and Mattoo AK (1992) Identification, charaeterization, and resolution of the in vivo phosphorylated form of the D1 Photosystem II reaction center protein. J Biol Chem 167: 3523–3529.
Elich TD, Edelman M and Mattoo AK (1993) Dephosphorylation of Photosystem II core is light-regulated in vivo. EMBO J 12 4857–4862.
Gal A, Schuster G, Frid D, Canaani O, Schwieger HG and Ohad I (1988) Role of cytochrome b6lf complex in the redox-controlled activity of the Acetabularia thylakoid protein kinase. J Biol Chem 263: 7785–7791.
Ghirardi MT (1993) Phosphorylation and dissambly of the Photosystem II core as an early stage of photoinhibition. Planta 90: 107–113.
Gedde D, Buehhold J, Ehhert V and Oettmeier W (1992) Photoinhibition in intact spinach plants: effect of high light lntensities on the function of the two photosystems and on the content of the D1 protein under nitrogen. Biochim Biophys Acta 1140: 69–77.
Heinze J (1992) Inaktivierung und Abbau von Proteinen in der Photoinhibierung von isolierten Thylakoidmembranen. Dissertation, Ruhr-Universität, Germany, Bochum.
Kettunen R, Tyystjärvi E and Aro EM (1991) D1 protein-degradation during photoinhibition of intact leaves. A modification of the D1 protein precedes degradation. FEBS Lett 290: 153–156.
Kim JH, Nemson JA and Melis A (1993) Photosystem II reaction center damage and repair in Dunaliclla salina (green alga). Plant Physiol 103: 181–189.
Koivuniemi A, Aro E-A and Andersson B (1995) Degradation of the D1-and D2 proteins of Photosystem II in higher plants is regulated by reversible phosphorylation. Biochemistry 34: 16022–16029.
Komenda J and Barber J (1995) Comparison of psbO and psbH deletion tion mutants of Synechocystis PCC 6803 indicates that degradation of D1 protein is regulated by the QB site and dependent on protein synthesis. Biochemistry 34: 9625–9631.
Krause GH and Weis E (1991) Chlorophyll fluorescence and photosynthesis: The basics. Annu Rev Plant Physiol Plant Mol Biol 42: 313–349.
Leto KJ, Bell E and MeIntosh L (1985) Nuclear mutation leads to an accelerated turnover of chloroplast-encoded 48 kDa polypeptides in thylakoids lacking Photosystem II. EMBO J 4: 1645–1653.
Mattoo AK and Edelman M (1987) Intramembrane translocation and posttranslational palmitoylation of the chloroplast 32-kDa herbicide binding protein. Proc Natl Acad Sci USA 84: 1497–1501.
Melis A (1991) Dynamics of photosynthetic membrane composition and function. Biochim Biophys Acta 1058: 87–106.
Nanba O and Satoh K (1987) Isolation of a Photosystem II reaction center consisting of D1 and D2 polypeptides and cytochrome b559. Proc Natl Acad Sci USA 84: 109–112.
Ohad I, Keren N, Zer H, Gong H, Mor TS, Gal A, Tal S and Domovich Y (1994) Light-induced degradation of the Photosystem II reaction-centre D1 protein. In: Baker NR and Bowyer JR (eds) Photoinhibition of Photosynthesis: From Molecular Mechanisms to the Field, pp 161–178. Bios Scientific Publishers, Oxford.
Park Y-I, Anderson JM and Chow WS (1996) Photoinactivation of functional Photosystem II and D1-protein-synthesis in vivo are independent of the modulation of the photosynthetic apparatus by growth irradiance. Planta 198: 300–309.
Peter GF and Thornber JP (1991a) Biochemical composition and organization of higher plant Photosystem II light-harvesting pigment-proteins. J Biol Chem 266: 16745–16754.
Peter GF and Thornber JP (1991b) Biochemical evidence that higher plant Photosystem II core complex is a organized as a dimer. Plant Cell Physiol 32: 1237–1250.
Rintamäki E, Kettunen R, Tyystjärvi E and Aro EM (1995) Lightdependent phosphorylation of D1 reaction-centre protein of Photosystem II: hypothesis for the functional role in vivo. Physiol Plant 93: 191–193.
Rögner M, Boekema EJ and Barber J (1996) How does Photosystem 2 split water? the structural basis of efficient energy conversion. TIBS 2: 44–49.
Schägger H, Borchart U, Aquila H, Link TA and vonJagow G (1985) Isolation and aminoacid composition of the smallest subunit of beef heart b/c I complex. FEBS Lett 190: 89–94.
Schreiber U, Schliwa U and Bilger W (1986) Continuous recording of photochemical and non-photochemical fluorescence quenching with a new type of modulation fluorometer. Photosynth Res 10: 51–62.
Stefansson H, Wollenberger L and Albertsson PA (1994) Eragmentation and separation of the thylakoid membrane. Effect of lightinduced protein-phosphorylation on domain composition. J Chromatogr 668: 191–200.
Walker D (1987) The Use of the Oxygen Electrode and Fluorescence Probes in Simple Measurements of Photosynthesis. Oxygraphic Limited, Sheffield, UK.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Ebbert, V., Godde, D. Phosphorylation of PS II polypeptides inhibits D1 protein-degradation and increases PS II stability. Photosynth Res 50, 257–269 (1996). https://doi.org/10.1007/BF00033124
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00033124