Skip to main content
PMC home page
Genetics logoLink to Genetics
. 2003 Mar;163(3):895–904. doi: 10.1093/genetics/163.3.895

Multiple translational control sequences in the 5' leader of the chloroplast psbC mRNA interact with nuclear gene products in Chlamydomonas reinhardtii.

William Zerges 1, Andrea H Auchincloss 1, Jean-David Rochaix 1
PMCID: PMC1462503  PMID: 12663530

Abstract

Translation of the chloroplast psbC mRNA in the unicellular eukaryotic alga Chlamydomonas reinhardtii is controlled by interactions between its 547-base 5' untranslated region and the products of the nuclear loci TBC1, TBC2, and possibly TBC3. In this study, a series of site-directed mutations in this region was generated and the ability of these constructs to drive expression of a reporter gene was assayed in chloroplast transformants that are wild type or mutant at these nuclear loci. Two regions located in the middle of the 5' leader and near the initiation codon are important for translation. Other deletions still allow for partial expression of the reporter gene in the wild-type background. Regions with target sites for TBC1 and TBC2 were identified by estimating the residual translation activity in the respective mutant backgrounds. TBC1 targets include mostly the central part of the leader and the translation initiation region whereas the only detected TBC2 targets are in the 3' part. The 5'-most 93 nt of the leader are required for wild-type levels of transcription and/or mRNA stabilization. The results indicate that TBC1 and TBC2 function independently and further support the possibility that TBC1 acts together with TBC3.

Full Text

The Full Text of this article is available as a PDF (339.4 KB).

Selected References

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

  1. Auchincloss Andrea H., Zerges William, Perron Karl, Girard-Bascou Jacqueline, Rochaix Jean-David. Characterization of Tbc2, a nucleus-encoded factor specifically required for translation of the chloroplast psbC mRNA in Chlamydomonas reinhardtii. J Cell Biol. 2002 Jun 3;157(6):953–962. doi: 10.1083/jcb.200201060. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Barkan A., Goldschmidt-Clermont M. Participation of nuclear genes in chloroplast gene expression. Biochimie. 2000 Jun-Jul;82(6-7):559–572. doi: 10.1016/s0300-9084(00)00602-7. [DOI] [PubMed] [Google Scholar]
  3. Bruick RK, Mayfield SP. Light-activated translation of chloroplast mRNAs. Trends Plant Sci. 1999 May;4(5):190–195. doi: 10.1016/s1360-1385(99)01402-8. [DOI] [PubMed] [Google Scholar]
  4. Choquet Y., Wostrikoff K., Rimbault B., Zito F., Girard-Bascou J., Drapier D., Wollman F. A. Assembly-controlled regulation of chloroplast gene translation. Biochem Soc Trans. 2001 Aug;29(Pt 4):421–426. doi: 10.1042/bst0290421. [DOI] [PubMed] [Google Scholar]
  5. Danon A. Translational regulation in the chloroplast. Plant Physiol. 1997 Dec;115(4):1293–1298. doi: 10.1104/pp.115.4.1293. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dron M., Rahire M., Rochaix J. D. Sequence of the chloroplast DNA region of Chlamydomonas reinhardii containing the gene of the large subunit of ribulose bisphosphate carboxylase and parts of its flanking genes. J Mol Biol. 1982 Dec 25;162(4):775–793. doi: 10.1016/0022-2836(82)90547-2. [DOI] [PubMed] [Google Scholar]
  7. Eibl C, Zou Z, Beck a, Kim M, Mullet J, Koop HU. In vivo analysis of plastid psbA, rbcL and rpl32 UTR elements by chloroplast transformation: tobacco plastid gene expression is controlled by modulation of transcript levels and translation efficiency. Plant J. 1999 Aug;19(3):333–345. doi: 10.1046/j.1365-313x.1999.00543.x. [DOI] [PubMed] [Google Scholar]
  8. Esposito D., Hicks A. J., Stern D. B. A role for initiation codon context in chloroplast translation. Plant Cell. 2001 Oct;13(10):2373–2384. doi: 10.1105/tpc.010236. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fargo D. C., Boynton J. E., Gillham N. W. Chloroplast ribosomal protein S7 of Chlamydomonas binds to chloroplast mRNA leader sequences and may be involved in translation initiation. Plant Cell. 2001 Jan;13(1):207–218. doi: 10.1105/tpc.13.1.207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fargo D. C., Zhang M., Gillham N. W., Boynton J. E. Shine-Dalgarno-like sequences are not required for translation of chloroplast mRNAs in Chlamydomonas reinhardtii chloroplasts or in Escherichia coli. Mol Gen Genet. 1998 Feb;257(3):271–282. doi: 10.1007/s004380050648. [DOI] [PubMed] [Google Scholar]
  11. Goldschmidt-Clermont M. Coordination of nuclear and chloroplast gene expression in plant cells. Int Rev Cytol. 1998;177:115–180. doi: 10.1016/s0074-7696(08)62232-9. [DOI] [PubMed] [Google Scholar]
  12. Goldschmidt-Clermont M., Girard-Bascou J., Choquet Y., Rochaix J. D. Trans-splicing mutants of Chlamydomonas reinhardtii. Mol Gen Genet. 1990 Sep;223(3):417–425. doi: 10.1007/BF00264448. [DOI] [PubMed] [Google Scholar]
  13. Goldschmidt-Clermont M., Rahire M. Sequence, evolution and differential expression of the two genes encoding variant small subunits of ribulose bisphosphate carboxylase/oxygenase in Chlamydomonas reinhardtii. J Mol Biol. 1986 Oct 5;191(3):421–432. doi: 10.1016/0022-2836(86)90137-3. [DOI] [PubMed] [Google Scholar]
  14. Goldschmidt-Clermont M. Transgenic expression of aminoglycoside adenine transferase in the chloroplast: a selectable marker of site-directed transformation of chlamydomonas. Nucleic Acids Res. 1991 Aug 11;19(15):4083–4089. doi: 10.1093/nar/19.15.4083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gorman D. S., Levine R. P. Cytochrome f and plastocyanin: their sequence in the photosynthetic electron transport chain of Chlamydomonas reinhardi. Proc Natl Acad Sci U S A. 1965 Dec;54(6):1665–1669. doi: 10.1073/pnas.54.6.1665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Higgs D. C., Shapiro R. S., Kindle K. L., Stern D. B. Small cis-acting sequences that specify secondary structures in a chloroplast mRNA are essential for RNA stability and translation. Mol Cell Biol. 1999 Dec;19(12):8479–8491. doi: 10.1128/mcb.19.12.8479. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kozak M. Initiation of translation in prokaryotes and eukaryotes. Gene. 1999 Jul 8;234(2):187–208. doi: 10.1016/s0378-1119(99)00210-3. [DOI] [PubMed] [Google Scholar]
  18. Leu S., Schlesinger J., Michaels A., Shavit N. Complete DNA sequence of the Chlamydomonas reinhardtii chloroplast atpA gene. Plant Mol Biol. 1992 Feb;18(3):613–616. doi: 10.1007/BF00040681. [DOI] [PubMed] [Google Scholar]
  19. Mayfield S. P., Cohen A., Danon A., Yohn C. B. Translation of the psbA mRNA of Chlamydomonas reinhardtii requires a structured RNA element contained within the 5' untranslated region. J Cell Biol. 1994 Dec;127(6 Pt 1):1537–1545. doi: 10.1083/jcb.127.6.1537. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Nickelsen J., Fleischmann M., Boudreau E., Rahire M., Rochaix J. D. Identification of cis-acting RNA leader elements required for chloroplast psbD gene expression in Chlamydomonas. Plant Cell. 1999 May;11(5):957–970. doi: 10.1105/tpc.11.5.957. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Nickelsen J., van Dillewijn J., Rahire M., Rochaix J. D. Determinants for stability of the chloroplast psbD RNA are located within its short leader region in Chlamydomonas reinhardtii. EMBO J. 1994 Jul 1;13(13):3182–3191. doi: 10.1002/j.1460-2075.1994.tb06617.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Osada Y., Saito R., Tomita M. Analysis of base-pairing potentials between 16S rRNA and 5' UTR for translation initiation in various prokaryotes. Bioinformatics. 1999 Jul-Aug;15(7-8):578–581. doi: 10.1093/bioinformatics/15.7.578. [DOI] [PubMed] [Google Scholar]
  23. Rochaix J. D., Kuchka M., Mayfield S., Schirmer-Rahire M., Girard-Bascou J., Bennoun P. Nuclear and chloroplast mutations affect the synthesis or stability of the chloroplast psbC gene product in Chlamydomonas reinhardtii. EMBO J. 1989 Apr;8(4):1013–1021. doi: 10.1002/j.1460-2075.1989.tb03468.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rochaix J. D. Post-transcriptional regulation of chloroplast gene expression in Chlamydomonas reinhardtii. Plant Mol Biol. 1996 Oct;32(1-2):327–341. doi: 10.1007/BF00039389. [DOI] [PubMed] [Google Scholar]
  25. Rochaix J. D. Restriction endonuclease map of the chloroplast DNA of Chlamydomonas reinhardii. J Mol Biol. 1978 Dec 25;126(4):597–617. doi: 10.1016/0022-2836(78)90011-6. [DOI] [PubMed] [Google Scholar]
  26. Rott R., Levy H., Drager R. G., Stern D. B., Schuster G. 3'-Processed mRNA is preferentially translated in Chlamydomonas reinhardtii chloroplasts. Mol Cell Biol. 1998 Aug;18(8):4605–4611. doi: 10.1128/mcb.18.8.4605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Sakamoto W., Chen X., Kindle K. L., Stern D. B. Function of the Chlamydomonas reinhardtii petd 5' untranslated region in regulating the accumulation of subunit IV of the cytochrome b6/f complex. Plant J. 1994 Oct;6(4):503–512. doi: 10.1046/j.1365-313x.1994.6040503.x. [DOI] [PubMed] [Google Scholar]
  28. Stampacchia O., Girard-Bascou J., Zanasco J. L., Zerges W., Bennoun P., Rochaix J. D. A nuclear-encoded function essential for translation of the chloroplast psaB mRNA in chlamydomonas. Plant Cell. 1997 May;9(5):773–782. doi: 10.1105/tpc.9.5.773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sugiura M., Hirose T., Sugita M. Evolution and mechanism of translation in chloroplasts. Annu Rev Genet. 1998;32:437–459. doi: 10.1146/annurev.genet.32.1.437. [DOI] [PubMed] [Google Scholar]
  30. Woessner J. P., Gillham N. W., Boynton J. E. The sequence of the chloroplast atpB gene and its flanking regions in Chlamydomonas reinhardtii. Gene. 1986;44(1):17–28. doi: 10.1016/0378-1119(86)90038-7. [DOI] [PubMed] [Google Scholar]
  31. Yohn C. B., Cohen A., Danon A., Mayfield S. P. Altered mRNA binding activity and decreased translational initiation in a nuclear mutant lacking translation of the chloroplast psbA mRNA. Mol Cell Biol. 1996 Jul;16(7):3560–3566. doi: 10.1128/mcb.16.7.3560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Yohn C. B., Cohen A., Rosch C., Kuchka M. R., Mayfield S. P. Translation of the chloroplast psbA mRNA requires the nuclear-encoded poly(A)-binding protein, RB47. J Cell Biol. 1998 Jul 27;142(2):435–442. doi: 10.1083/jcb.142.2.435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Zerges W., Girard-Bascou J., Rochaix J. D. Translation of the chloroplast psbC mRNA is controlled by interactions between its 5' leader and the nuclear loci TBC1 and TBC3 in Chlamydomonas reinhardtii. Mol Cell Biol. 1997 Jun;17(6):3440–3448. doi: 10.1128/mcb.17.6.3440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zerges W., Rochaix J. D. The 5' leader of a chloroplast mRNA mediates the translational requirements for two nucleus-encoded functions in Chlamydomonas reinhardtii. Mol Cell Biol. 1994 Aug;14(8):5268–5277. doi: 10.1128/mcb.14.8.5268. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Zerges William. Does complexity constrain organelle evolution? Trends Plant Sci. 2002 Apr;7(4):175–182. doi: 10.1016/s1360-1385(02)02233-1. [DOI] [PubMed] [Google Scholar]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES