Uniacke, James and Zerges, William (2009) Chloroplast protein targeting involves localized translation in Chlamydomonas. Proceedings of the National Academy of Sciences, 106 (5). pp. 1439-1444. ISSN 0027-8424 (In Press)
- Published Version
Official URL: http://dx.doi.org/10.1073/pnas.0811268106
The compartmentalization of eukaryotic cells requires that newly synthesized proteins be targeted to the compartments in which they function. In chloroplasts, a few thousand proteins function in photosynthesis, expression of the chloroplast genome, and other processes. Most chloroplast proteins are synthesized in the cytoplasm, imported, and then targeted to a specific chloroplast compartment. The remainder are encoded by the chloroplast genome, synthesized within the organelle, and targeted by mechanisms that are only beginning to be elucidated. We used fluorescence confocal microscopy to explore the targeting mechanisms used by several chloroplast proteins in the green alga Chlamydomonas. These include the small subunit of ribulose bisphosphate carboxylase (rubisco) and the light-harvesting complex II (LHCII) subunits, which are imported from the cytoplasm, and 2 proteins synthesized in the chloroplast: the D1 subunit of photosystem II and the rubisco large subunit. We determined whether the targeting of each protein involves localized translation of the mRNA that encodes it. When this was the case, we explored whether the targeting sequence was in the nascent polypeptide or in the mRNA, based on whether the localization was translation-dependent or -independent, respectively. The results reveal 2 novel examples of targeting by localized translation, in LHCII subunit import and the targeting of the rubisco large subunit to the pyrenoid. They also demonstrate examples of each of the three known mechanisms—posttranslational, cotranslational (signal recognition particle-mediated), and mRNA-based—in the targeting of specific chloroplast proteins. Our findings can help guide the exploration of these pathways at the biochemical level.
|Divisions:||Concordia University > Faculty of Arts and Science > Biology|
|Authors:||Uniacke, James and Zerges, William|
|Journal or Publication:||Proceedings of the National Academy of Sciences|
|Date:||3 February 2009|
|Keywords:||chloroplast, FISHorganelle, protein target, ingtranslation|
|Deposited By:||DANIELLE DENNIE|
|Deposited On:||16 May 2011 16:15|
|Last Modified:||24 Aug 2016 21:26|
1. St. Johnston D(2005) Moving messages: The intracellular localization of mRNAs. Nat Rev Mol Cell Biol 6:363–375.
2. Sanchirico ME, Fox TD, Mason TL(1998) Accumulation of mitochondrially synthesized Saccharomyces cerevisiae Cox2p and Cox3p depends on targeting information in untranslated portions of their mRNAs. EMBO J 17:5796–5804.
3. Stephens SB, et al.(2008) Analysis of mRNA partitioning between the cytosol and endoplasmic reticulum compartments of mammalian cells. Methods Mol Biol 419:197–214.
4. Inaba T, Schnell DJ(2008) Protein trafficking to plastids: One theme, many variations. Biochem J 413:15–28.
5. Chua NH, Schmidt GW(1979) Transport of proteins into mitochondria and chloroplasts. J Cell Biol 81:461–483.
6. Carde JP, Joyard J, Douce R(1982) Electron microscopic studies of envelope membranes from spinach plastids. Biol Cell 44:315–324.
7. Nanjo Y, et al.(2006) Rice plastidial N-glycosylated nucleotide pyrophosphatase/phosphodiesterase is transported from the ER-Golgi to the chloroplast through the secretory pathway. Plant Cell 18:2582–2592.
8. Villarejo A, et al.(2005) Evidence for a protein transported through the secretory pathway en route to the higher plant chloroplast. Nat Cell Biol 7:1224–1231.
9. Radhamony RN, Theg SM(2006) Evidence for an ER to Golgi to chloroplast protein transport pathway. Trends Cell Biol 16:385–387.
10. Zerges W(2000) Translation in chloroplasts. Biochimie 82:583–601.
11. Borkhsenious ON, Mason CB, Moroney JV(1998) The intracellular localization of ribulose-1,5-bisphosphate carboxylase/oxygenase in Chlamydomonas reinhardtii. Plant Physiol 116:1585–1591.
12.Chua NH, Blobel G, Siekevitz P, Palade GE(1976) Periodic variations in the ratio of free to thylakoid-bound chloroplast ribosomes during the cell cycle of Chlamydomonas reinhardtii. J Cell Biol 71:497–514.
13.Margulies MM(1983) Synthesis of photosynthetic membrane proteins directed by RNA from rough thylakoids of Chlamydomonas reinhardtii. Eur J Biochem 137:241–248.
14.Eichacker LA, Henry R(2001) Function of a chloroplast SRP in thylakoid protein export. Biochim Biophys Acta 1541:120–134.
15.Gutensohn M, et al.(2006) Toc, Tic, Tat, et al.: Structure and function of protein transport machineries in chloroplasts. J Plant Physiol 163:333–347.
16. Zhang L, Aro EM(2002) Synthesis, membrane insertion and assembly of the chloroplast-encoded D1 protein into photosystem II. FEBS Lett 512:13–18.
17. van Wijk KJ, Knott TG, Robinson C(1995) Evidence for SecA- and delta pH-independent insertion of D1 into thylakoids. FEBS Lett 368:263–266.
18. Amin P, et al.(1999) Arabidopsis mutants lacking the 43- and 54-kilodalton subunits of the chloroplast signal recognition particle have distinct phenotypes. Plant Physiol 121:61–70.
19. Nickelsen J(2003) Chloroplast RNA-binding proteins. Curr Genet 43:392–399.
20. Zerges W, Wang S, Rochaix JD(2002) Light activates binding of membrane proteins to chloroplast RNAs in Chlamydomonas reinhardtii. Plant Mol Biol 50:573–585.
21. Uniacke J, Zerges W(2008) Stress induces the assembly of RNA granules in the chloroplast of Chlamydomonas reinhardtii. J Cell Biol 182:641–646.
22. Zhang L, Paakkarinen V, van Wijk KJ, Aro EM(2000) Biogenesis of the chloroplast-encoded D1 protein: Regulation of translation elongation, insertion, and assembly into photosystem II. Plant Cell 12:1769–1782.
23. Edhofer I, Muhlbauer SK, Eichacker LA(1998) Light regulates the rate of translation elongation of chloroplast reaction center protein D1. Eur J Biochem 257:78–84.
24. Lee J, Herrin DL(2002) Assessing the relative importance of light and the circadian clock in controlling chloroplast translation in Chlamydomonas reinhardtii. Photosynth Res 72:295–306.
25. Herrin DL, Michaels AS, Paul AL(1986) Regulation of genes encoding the large subunit of ribulose-1,5-bisphosphate carboxylase and the photosystem II polypeptides D-1 and D-2 during the cell cycle of Chlamydomonas reinhardtii. J Cell Biol 103:1837–1845.
26. Shapira M, et al.(1997) Differential regulation of chloroplast gene expression in Chlamydomonas reinhardtii during photoacclimation: Light stress transiently suppresses synthesis of the rubisco LSU protein while enhancing synthesis of the PSII D1 protein. Plant Mol Biol 33:1001–1011.
27. Malnoe P, Mayfield SP, Rochaix JD(1988) Comparative analysis of the biogenesis of photosystem II in the wild-type and Y-1 mutant of Chlamydomonas reinhardtii. J Cell Biol 106:609–616.
28. Breidenbach E, Jenni E, Boschetti A(1988) Synthesis of two proteins in chloroplasts and mRNA distribution between thylakoids and stroma during the cell cycle of Chlamydomonas reinhardii. Eur J Biochem 177:225–232.
29. Uniacke J, Zerges W(2007) Photosystem II assembly and repair are differentially localized in Chlamydomonas. Plant Cell 19:3640–3654.
30. Abramoff MD, Magelhaes PJ, Ram SJ(2004) Image processing with Image. J. Biophotonics Int 11:36–42.
31. Adir N, Shochat S, Ohad I(1990) Light-dependent D1 protein synthesis and translocation is regulated by reaction center II: Reaction center II serves as an acceptor for the D1 precursor. J Biol Chem 265:12563–12568.
32. Minagawa J, Takahashi Y(2004) Structure, function and assembly of photosystem II and its light-harvesting proteins. Photosynth Res 82:241–263.
33. Colon-Ramos DA, et al.(2003) Asymmetric distribution of nuclear pore complexes and the cytoplasmic localization of beta2-tubulin mRNA in Chlamydomonas reinhardtii. Dev Cell 4:941–952.
34. Fleming GH, Boynton JE, Gillham NW(1987) Cytoplasmic ribosomal proteins from Chlamydomonas reinhardtii: Characterization and immunological comparisons. Mol Gen Genet 206:226–237.
35. Durnford DG, Price JA, McKim SM, Sarchfield ML(2003) Light-harvesting complex gene expression is controlled by both transcriptional and post-transcriptional mechanisms during photoacclimation in Chlamydomonas reinhardtii. Physiol Plant 118:193–205.
36. Khrebtukova I, Spreitzer RJ(1996) Elimination of the Chlamydomonas gene family that encodes the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase. Proc Natl Acad Sci U S A 93:13689–13693.
37. Reinbothe S, Reinbothe C, Parthier B(1993) Methyl jasmonate–regulated translation of nuclear-encoded chloroplast proteins in barley (Hordeum vulgare L. cv. Salome) J Biol Chem 268:10606–10611.
38. Emanuelsson O, Nielsen H, Brunak S, von Heijne G(2000) Predicting subcellular localization of proteins based on their N-terminal amino acid sequence. J Mol Biol 300:1005–1016.
39. Gibbs S(1979) The route of entry of cytoplasmically synthesized proteins into chloroplasts of algae possessing chloroplast ER. J Cell Sci 35:253–266.
40. Chua NH, Blobel G, Siekevitz P, Palade GE(1973) Attachment of chloroplast polysomes to thylakoid membranes in Chlamydomonas reinhardtii. Proc Natl Acad Sci U S A 70:1554–1558.
41. Suss KH, Prokhorenko I, Adler K(1995) In situ association of Calvin cycle enzymes, ribulose-1,5-bisphosphate carboxylase/oxygenase activase, ferredoxin-NADP+ reductase, and nitrite reductase with thylakoid and pyrenoid membranes of Chlamydomonas reinhardtii chloroplasts as revealed by immunoelectron microscopy. Plant Physiol 107:1387–1397.
42. Pyhtila B, et al.(2008) Signal sequence– and translation-independent mRNA localization to the endoplasmic reticulum. RNA 14:445–453.
43. Margulies MM, Michaels A(1974) Ribosomes bound to chloroplast membranes in Chlamydomonas reinhardtii. J Cell Biol 60:65–77.
44. Yamamoto T, Burke J, Autz G, Jagendorf AT(1981) Bound ribosomes of pea chloroplast thylakoid membranes: Location and release in vitro by high salt, puromycin, and RNase. Plant Physiol 67:940–949.
45. Green-Willms NS, Fox TD, Costanzo MC(1998) Functional interactions between yeast mitochondrial ribosomes and mRNA 5′ untranslated leaders. Mol Cell Biol 18:1826–1834.
46. Schmidt RJ, Myers AM, Gillham NW, Boynton JE(1984) Immunological similarities between specific chloroplast ribosomal proteins from Chlamydomonas reinhardtii and ribosomal proteins from Escherichia coli. Mol Biol Evol 1:317–334.
47. Randolph-Anderson BL, Gillham NW, Boynton JE(1989) Electrophoretic and immunological comparisons of chloroplast and prokaryotic ribosomal proteins reveal that certain families of large subunit proteins are evolutionarily conserved. J Mol Evol 29:68–88.
48. Schmidt RJ, Richardson CB, Gillham NW, Boynton JE(1983) Sites of synthesis of chloroplast ribosomal proteins in Chlamydomonas. J Cell Biol 96:1451–1463.
49. Fleming GH, Boynton JE, Gillham NW(1987) The cytoplasmic ribosomes of Chlamydomonas reinhardtii: Characterization of antibiotic sensitivity and cycloheximide-resistant mutants. Mol Gen Genet 210:419–428.
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