Skip to main content
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1984 Jun 1;98(6):2011–2018. doi: 10.1083/jcb.98.6.2011

Chloroplast ribosomal proteins of Chlamydomonas synthesized in the cytoplasm are made as precursors

PMCID: PMC2113070  PMID: 6202701

Abstract

Polyadenylated RNA from Chlamydomonas was translated in a cell-free rabbit reticulocyte system that employed [35S]methionine. Antibodies made to four chloroplast ribosomal proteins synthesized in the cytoplasm and imported into the organelle were used for indirect immunoprecipitation of the labeled translation products, which were subsequently visualized on fluorographs of SDS gels. The cytoplasmically synthesized chloroplast ribosomal proteins were first seen as precursors with apparent molecular weights of 1,000 to 6,000 greater than their respective mature forms. Processing of the ribosomal protein precursors to mature proteins was affected by adding a postribosomal supernatant that had been extracted from cells of Chlamydomonas. In contrast to the chloroplast ribosomal proteins synthesized in the cytoplasm, two such proteins made within the chloroplast were found to be synthesized in mature form in cell-free wheat germ translation systems programmed with nonpolyadenylated RNA.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Apel K., Kloppstech K. The plastid membranes of barley (Hordeum vulgare). Light-induced appearance of mRNA coding for the apoprotein of the light-harvesting chlorophyll a/b protein. Eur J Biochem. 1978 Apr 17;85(2):581–588. doi: 10.1111/j.1432-1033.1978.tb12273.x. [DOI] [PubMed] [Google Scholar]
  2. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  3. Cashmore A. R., Broadhurst M. K., Gray R. E. Cell-free synthesis of leaf protein: Identification of an apparent precursor of the small subunit of ribulose-1,5-bisphosphate carboxylase. Proc Natl Acad Sci U S A. 1978 Feb;75(2):655–659. doi: 10.1073/pnas.75.2.655. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chua N. H., Schmidt G. W. In vitro synthesis, transport, and assembly of ribulose 1,5-bisphosphate carboxylase subunits. Basic Life Sci. 1978;11:325–347. doi: 10.1007/978-1-4684-8106-8_20. [DOI] [PubMed] [Google Scholar]
  5. Chua N. H., Schmidt G. W. Post-translational transport into intact chloroplasts of a precursor to the small subunit of ribulose-1,5-bisphosphate carboxylase. Proc Natl Acad Sci U S A. 1978 Dec;75(12):6110–6114. doi: 10.1073/pnas.75.12.6110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chua N. H., Schmidt G. W. Transport of proteins into mitochondria and chloroplasts. J Cell Biol. 1979 Jun;81(3):461–483. doi: 10.1083/jcb.81.3.461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Conde M. F., Boynton J. E., Gillham N. W., Harris E. H., Tingle C. L., Wang W. L. Chloroplast genes in Chlamydomonas affecting organelle ribosomes. Genetic and biochemical analysis of analysis of antibiotic-resistant mutants at several gene loci. Mol Gen Genet. 1975 Oct 3;140(3):183–220. doi: 10.1007/BF00334266. [DOI] [PubMed] [Google Scholar]
  8. Coruzzi G., Broglie R., Cashmore A., Chua N. H. Nucleotide sequences of two pea cDNA clones encoding the small subunit of ribulose 1,5-bisphosphate carboxylase and the major chlorophyll a/b-binding thylakoid polypeptide. J Biol Chem. 1983 Feb 10;258(3):1399–1402. [PubMed] [Google Scholar]
  9. Davidson J. N., Bogorad L. Suppression of erythromycin resistance in ery-M1 mutants of Chlamydomonas reinhardi. Mol Gen Genet. 1977 Nov 29;157(1):39–46. doi: 10.1007/BF00268685. [DOI] [PubMed] [Google Scholar]
  10. Davidson J. N., Hanson M. R., Bogorad L. An altered chloroplast ribosomal protein in ery-M1 mutants of Chlamydomonas reinhardi. Mol Gen Genet. 1974;132(2):119–129. doi: 10.1007/BF00272177. [DOI] [PubMed] [Google Scholar]
  11. Dobberstein B., Blobel G., Chua N. H. In vitro synthesis and processing of a putative precursor for the small subunit of ribulose-1,5-bisphosphate carboxylase of Chlamydomonas reinhardtii. Proc Natl Acad Sci U S A. 1977 Mar;74(3):1082–1085. doi: 10.1073/pnas.74.3.1082. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gray R. E., Cashmore A. R. RNA synthesis in plant leaf tissue: the characterization of messenger RNA species lacking and containing polyadenylic acid. J Mol Biol. 1976 Dec 15;108(3):595–608. doi: 10.1016/s0022-2836(76)80139-8. [DOI] [PubMed] [Google Scholar]
  13. Grossman A. R., Bartlett S. G., Schmidt G. W., Mullet J. E., Chua N. H. Optimal conditions for post-translational uptake of proteins by isolated chloroplasts. In vitro synthesis and transport of plastocyanin, ferredoxin-NADP+ oxidoreductase, and fructose-1,6-bisphosphatase. J Biol Chem. 1982 Feb 10;257(3):1558–1563. [PubMed] [Google Scholar]
  14. Hjelm H., Hjelm K., Sjöquist J. Protein A from Staphylococcus aureus. Its isolation by affinity chromatography and its use as an immunosorbent for isolation of immunoglobulins. FEBS Lett. 1972 Nov 15;28(1):73–76. doi: 10.1016/0014-5793(72)80680-x. [DOI] [PubMed] [Google Scholar]
  15. Huisman J. G., Moorman A. F., Verkley F. N. In vitro synthesis of chloroplast ferredoxin as a high molecular weight precursor in a cell-free protein synthesizing system from wheat germs. Biochem Biophys Res Commun. 1978 Jun 29;82(4):1121–1131. doi: 10.1016/0006-291x(78)90303-0. [DOI] [PubMed] [Google Scholar]
  16. Marcus A., Efron D., Weeks D. P. The wheat embryo cell-free system. Methods Enzymol. 1974;30:749–754. doi: 10.1016/0076-6879(74)30073-0. [DOI] [PubMed] [Google Scholar]
  17. Martin N. C., Chiang K. S., Goodenough U. W. Turnover of chloroplast and cytoplasmic ribosomes during gametogenesis in Chlamydomonas reinhardi. Dev Biol. 1976 Jul 15;51(2):190–201. doi: 10.1016/0012-1606(76)90137-8. [DOI] [PubMed] [Google Scholar]
  18. Mets L., Bogorad L. Altered chlorplast ribosomal proteins associated with erythromycin-resistant mutants in two genetic systems of Chlamydomonas reinhardi. Proc Natl Acad Sci U S A. 1972 Dec;69(12):3779–3783. doi: 10.1073/pnas.69.12.3779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Pelham H. R., Jackson R. J. An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976 Aug 1;67(1):247–256. doi: 10.1111/j.1432-1033.1976.tb10656.x. [DOI] [PubMed] [Google Scholar]
  20. Schmidt G. W., Bartlett S. G., Grossman A. R., Cashmore A. R., Chua N. H. Biosynthetic pathways of two polypeptide subunits of the light-harvesting chlorophyll a/b protein complex. J Cell Biol. 1981 Nov;91(2 Pt 1):468–478. doi: 10.1083/jcb.91.2.468. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Schmidt G. W., Devillers-Thiery A., Desruisseaux H., Blobel G., Chua N. H. NH2-terminal amino acid sequences of precursor and mature forms of the ribulose-1,5-bisphosphate carboxylase small subunit from Chlamydomonas reinhardtii. J Cell Biol. 1979 Dec;83(3):615–622. doi: 10.1083/jcb.83.3.615. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Schmidt R. J., Richardson C. B., Gillham N. W., Boynton J. E. Sites of synthesis of chloroplast ribosomal proteins in Chlamydomonas. J Cell Biol. 1983 May;96(5):1451–1463. doi: 10.1083/jcb.96.5.1451. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Shepherd H. S., Boynton J. E., Gillham N. W. Mutations in nine chloroplast loci of Chlamydomonas affecting different photosynthetic functions. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1353–1357. doi: 10.1073/pnas.76.3.1353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Siersma P. W., Chiang K. S. Conservation and degradation of cytoplasmic and chloroplast ribosomes in Chlamydomonas reinhardtii. J Mol Biol. 1971 May 28;58(1):167–185. doi: 10.1016/0022-2836(71)90239-7. [DOI] [PubMed] [Google Scholar]
  25. Sueoka N. MITOTIC REPLICATION OF DEOXYRIBONUCLEIC ACID IN CHLAMYDOMONAS REINHARDI. Proc Natl Acad Sci U S A. 1960 Jan;46(1):83–91. doi: 10.1073/pnas.46.1.83. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Van Loon A. P., Kreike J., De Ronde A., Van der Horst G. T., Gasser S. M., Grivell L. A. Biosynthesis of the ubiquinol-cytochrome c reductase complex in yeast. Characterization of precursor forms of the 44-kDa, 40-kDa and 17-kDa subunits and identification of individual messenger RNAs for these and other imported subunits of the complex. Eur J Biochem. 1983 Oct 3;135(3):457–463. doi: 10.1111/j.1432-1033.1983.tb07673.x. [DOI] [PubMed] [Google Scholar]
  27. Wollgiehn R., Parthier B. RNA and protein synthesis in plastid differentiation. Results Probl Cell Differ. 1980;10:97–145. doi: 10.1007/978-3-540-38255-3_4. [DOI] [PubMed] [Google Scholar]

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

RESOURCES