Skip to main content
NCBI home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1989 Sep 1;109(3):991–1006. doi: 10.1083/jcb.109.3.991

Posttranslational events leading to the assembly of photosystem II protein complex: a study using photosynthesis mutants from Chlamydomonas reinhardtii

PMCID: PMC2115777  PMID: 2670960

Abstract

We studied the assembly of photosystem II (PSII) in several mutants from Chlamydomonas reinhardtii which were unable to synthesize either one PSII core subunit (P6 [43 kD], D1, or D2) or one oxygen-evolving enhancer (OEE1 or OEE2) subunit. Synthesis of the PSII subunits was analyzed on electrophoretograms of cells pulse labeled with [14C]acetate. Their accumulation in thylakoid membranes was studied on immunoblots, their chlorophyll-binding ability on nondenaturating gels, their assembly by detergent fractionation, their stability by pulse- chase experiments and determination of in vitro protease sensitivity, and their localization by immunocytochemistry. In Chlamydomonas, the PSII core subunits P5 (47 kD), D1, and D2 are synthesized in a concerted manner while P6 synthesis is independent. P5 and P6 accumulate independently of each other in the stacked membranes. They bind chlorophyll soon after, or concomitantly with, their synthesis and independently of the presence of the other PSII subunits. Resistance to degradation increases step by step: beginning with assembly of P5, D1, and D2, then with binding of P6, and, finally, with binding of the OEE subunits on two independent high affinity sites (one for OEE1 and another for OEE2 to which OEE3 binds). In the absence of PSII cores, the OEE subunits accumulate independently in the thylakoid lumen and bind loosely to the membranes; OEE1 was found on stacked membranes, but OEE2 was found on either stacked or unstacked membranes depending on whether or not P6 was synthesized.

Full Text

The Full Text of this article is available as a PDF (4.4 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  2. Chua N. H., Bennoun P. Thylakoid membrane polypeptides of Chlamydomonas reinhardtii: wild-type and mutant strains deficient in photosystem II reaction center. Proc Natl Acad Sci U S A. 1975 Jun;72(6):2175–2179. doi: 10.1073/pnas.72.6.2175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Delepelaire P., Chua N. H. Lithium dodecyl sulfate/polyacrylamide gel electrophoresis of thylakoid membranes at 4 degrees C: Characterizations of two additional chlorophyll a-protein complexes. Proc Natl Acad Sci U S A. 1979 Jan;76(1):111–115. doi: 10.1073/pnas.76.1.111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Delepelaire P. Partial characterization of the biosynthesis and integration of the Photosystem II reaction centers in the thylakoid membrane of Chlamydomonas reinhardtii. EMBO J. 1984 Apr;3(4):701–706. doi: 10.1002/j.1460-2075.1984.tb01872.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Diner B. A., Ries D. F., Cohen B. N., Metz J. G. COOH-terminal processing of polypeptide D1 of the photosystem II reaction center of Scenedesmus obliquus is necessary for the assembly of the oxygen-evolving complex. J Biol Chem. 1988 Jun 25;263(18):8972–8980. [PubMed] [Google Scholar]
  6. Diner B. A., Wollman F. A. Isolation of highly active photosystem II particles from a mutant of Chlamydomonas reinhardtii. Eur J Biochem. 1980 Sep;110(2):521–526. doi: 10.1111/j.1432-1033.1980.tb04894.x. [DOI] [PubMed] [Google Scholar]
  7. Drummond I. A., Chisholm R. L. The effect of caffeine, adenosine, and buffer ionic composition on the induction of cell-surface cAMP binding during starvation of Dictyostelium discoideum. Dev Genet. 1988;9(4-5):293–301. doi: 10.1002/dvg.1020090411. [DOI] [PubMed] [Google Scholar]
  8. Erickson J. M., Rahire M., Malnoë P., Girard-Bascou J., Pierre Y., Bennoun P., Rochaix J. D. Lack of the D2 protein in a Chlamydomonas reinhardtii psbD mutant affects photosystem II stability and D1 expression. EMBO J. 1986 Aug;5(8):1745–1754. doi: 10.1002/j.1460-2075.1986.tb04422.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Erickson J. M., Rahire M., Rochaix J. D. Chlamydomonas reinhardii gene for the 32 000 mol. wt. protein of photosystem II contains four large introns and is located entirely within the chloroplast inverted repeat. EMBO J. 1984 Dec 1;3(12):2753–2762. doi: 10.1002/j.1460-2075.1984.tb02206.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Greer K. L., Plumley F. G., Schmidt G. W. The Water Oxidation Complex of Chlamydomonas: Accumulation and Maturation of the Largest Subunit in Photosystem II Mutants. Plant Physiol. 1986 Sep;82(1):114–120. doi: 10.1104/pp.82.1.114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Holschuh K., Bottomley W., Whitfeld P. R. Structure of the spinach chloroplast genes for the D2 and 44 kd reaction-centre proteins of photosystem II and for tRNASer (UGA). Nucleic Acids Res. 1984 Dec 11;12(23):8819–8834. doi: 10.1093/nar/12.23.8819. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hovanessian A. G., Galabru J., Rivière Y., Montagnier L. Efficiency of poly(A).poly(U) as an adjuvant. Immunol Today. 1988 Jun;9(6):161–162. doi: 10.1016/0167-5699(88)91288-1. [DOI] [PubMed] [Google Scholar]
  13. Jacobs E., Clad A. Electroelution of fixed and stained membrane proteins from preparative sodium dodecyl sulfate-polyacrylamide gels into a membrane trap. Anal Biochem. 1986 May 1;154(2):583–589. doi: 10.1016/0003-2697(86)90033-3. [DOI] [PubMed] [Google Scholar]
  14. Klein R. R., Mullet J. E. Regulation of chloroplast-encoded chlorophyll-binding protein translation during higher plant chloroplast biogenesis. J Biol Chem. 1986 Aug 25;261(24):11138–11145. [PubMed] [Google Scholar]
  15. Kuchka M. R., Mayfield S. P., Rochaix J. D. Nuclear mutations specifically affect the synthesis and/or degradation of the chloroplast-encoded D2 polypeptide of photosystem II in Chlamydomonas reinhardtii. EMBO J. 1988 Feb;7(2):319–324. doi: 10.1002/j.1460-2075.1988.tb02815.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kyle D. J., Ohad I., Arntzen C. J. Membrane protein damage and repair: Selective loss of a quinone-protein function in chloroplast membranes. Proc Natl Acad Sci U S A. 1984 Jul;81(13):4070–4074. doi: 10.1073/pnas.81.13.4070. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Leto K. J., Bell E., McIntosh L. Nuclear mutation leads to an accelerated turnover of chloroplast-encoded 48 kd and 34.5 kd polypeptides in thylakoids lacking photosystem II. EMBO J. 1985 Jul;4(7):1645–1653. doi: 10.1002/j.1460-2075.1985.tb03832.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Ljungberg U., Akerlund H. E., Andersson B. Isolation and characterization of the 10-kDa and 22-kDa polypeptides of higher plant photosystem 2. Eur J Biochem. 1986 Aug 1;158(3):477–482. doi: 10.1111/j.1432-1033.1986.tb09779.x. [DOI] [PubMed] [Google Scholar]
  20. Mattoo A. K., Edelman M. Intramembrane translocation and posttranslational palmitoylation of the chloroplast 32-kDa herbicide-binding protein. Proc Natl Acad Sci U S A. 1987 Mar;84(6):1497–1501. doi: 10.1073/pnas.84.6.1497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Mayfield S. P., Bennoun P., Rochaix J. D. Expression of the nuclear encoded OEE1 protein is required for oxygen evolution and stability of photosystem II particles in Chlamydomonas reinhardtii. EMBO J. 1987 Feb;6(2):313–318. doi: 10.1002/j.1460-2075.1987.tb04756.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mayfield S. P., Rahire M., Frank G., Zuber H., Rochaix J. D. Expression of the nuclear gene encoding oxygen-evolving enhancer protein 2 is required for high levels of photosynthetic oxygen evolution in Chlamydomonas reinhardtii. Proc Natl Acad Sci U S A. 1987 Feb;84(3):749–753. doi: 10.1073/pnas.84.3.749. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Michel H., Epp O., Deisenhofer J. Pigment-protein interactions in the photosynthetic reaction centre from Rhodopseudomonas viridis. EMBO J. 1986 Oct;5(10):2445–2451. doi: 10.1002/j.1460-2075.1986.tb04520.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nanba O., Satoh K. Isolation of a photosystem II reaction center consisting of D-1 and D-2 polypeptides and cytochrome b-559. Proc Natl Acad Sci U S A. 1987 Jan;84(1):109–112. doi: 10.1073/pnas.84.1.109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Piccioni R. G., Bennoun P., Chua N. H. A nuclear mutant of Chlamydomonas reinhardtii defective in photosynthetic photophosphorylation. Characterization of the algal coupling factor ATPase. Eur J Biochem. 1981 Jun;117(1):93–102. doi: 10.1111/j.1432-1033.1981.tb06307.x. [DOI] [PubMed] [Google Scholar]
  26. Rochaix J. D., Erickson J. Function and assembly of photosystem II: genetic and molecular analysis. Trends Biochem Sci. 1988 Feb;13(2):56–59. doi: 10.1016/0968-0004(88)90029-1. [DOI] [PubMed] [Google Scholar]
  27. Schmidt G. W., Mishkind M. L. Rapid degradation of unassembled ribulose 1,5-bisphosphate carboxylase small subunits in chloroplasts. Proc Natl Acad Sci U S A. 1983 May;80(9):2632–2636. doi: 10.1073/pnas.80.9.2632. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Staehelin L. A., Arntzen C. J. Regulation of chloroplast membrane function: protein phosphorylation changes the spatial organization of membrane components. J Cell Biol. 1983 Nov;97(5 Pt 1):1327–1337. doi: 10.1083/jcb.97.5.1327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sutton A., Sieburth L. E., Bennett J. Light-dependent accumulation and localization of photosystem II proteins in maize. Eur J Biochem. 1987 May 4;164(3):571–578. doi: 10.1111/j.1432-1033.1987.tb11165.x. [DOI] [PubMed] [Google Scholar]
  30. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Wollman F. A., Delepelaire P. Correlation between changes in light energy distribution and changes in thylakoid membrane polypeptide phosphorylation in Chlamydomonas reinhardtii. J Cell Biol. 1984 Jan;98(1):1–7. doi: 10.1083/jcb.98.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Wollman F. A., Diner B. A. Cation control of fluorescence emission, light scatter, and membrane stacking in pigment mutants of Chlamydomonas reinhardi. Arch Biochem Biophys. 1980 May;201(2):646–659. doi: 10.1016/0003-9861(80)90555-x. [DOI] [PubMed] [Google Scholar]
  33. Wollman F. A., Olive J., Bennoun P., Recouvreur M. Organization of the photosystem II centers and their associated antennae in the thylakoid membranes: a comparative ultrastructural, biochemical, and biophysical study of Chlamydomonas wild type and mutants lacking in photosystem II reaction centers. J Cell Biol. 1980 Dec;87(3 Pt 1):728–735. doi: 10.1083/jcb.87.3.728. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zurawski G., Bohnert H. J., Whitfeld P. R., Bottomley W. Nucleotide sequence of the gene for the M(r) 32,000 thylakoid membrane protein from Spinacia oleracea and Nicotiana debneyi predicts a totally conserved primary translation product of M(r) 38,950. Proc Natl Acad Sci U S A. 1982 Dec;79(24):7699–7703. doi: 10.1073/pnas.79.24.7699. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES