Abstract
The chloroplast protein CSP41a both binds and cleaves RNA, particularly in stem-loops, and has been found associated with ribosomes. A related protein, CSP41b, co-purifies with CSP41a, ribosomes, and the plastid-encoded RNA polymerase. Here we show that Arabidopsis CSP41a and CSP41b interact in vivo, and that a csp41b null mutant becomes depleted of CSP41a in mature leaves, correlating with a pale green phenotype and reduced accumulation of the ATP synthase and cytochrome b 6 /f complexes. RNA gel blot analyses revealed up to four-fold decreases in accumulation for some chloroplast RNAs, which run-on experiments suggested could tentatively be ascribed to decreased transcription. Depletion of both CSP41a and CSP41b triggered a promoter switch whereby atpBE became predominately transcribed from its nucleus-encoded polymerase promoter as opposed to its plastid-encoded polymerase promoter. Together with published proteomic data, this suggests that CSP41a and/or CSP41b enhances transcription by the plastid-encoded polymerase. Gradient analysis of rRNAs in the mutant suggest a defect in polysome assembly or stability, suggesting that CSP41a and/or CSP41b, which are not present in polysomal fractions, stabilize ribosome assembly intermediates. Although psbA and rbcL mRNAs are normally polysome-associated in the mutant, petD-containing RNAs have diminished association, perhaps accounting for reduced accumulation of its respective multimeric complex. In conclusion, our data suggest that CSP41a and CSP41b stimulate both transcription and translation in the chloroplast.
Similar content being viewed by others
References
Allison LA (2000) The role of sigma factors in plastid transcription. Biochimie 82:537–548. doi:10.1016/S0300-9084(00)00611-8
Alonso JM, Stepanova AN, Leisse TJ, Kim CJ, Chen H, Shinn P, Stevenson DK, Zimmerman J, Barajas P, Cheuk R, Gadrinab C, Heller C, Jeske A, Koesema E, Meyers CC, Parker H, Prednis L, Ansari Y, Choy N, Deen H, Geralt M, Hazari N, Hom E, Karnes M, Mulholland C, Ndubaku R, Schmidt I, Guzman P, Aguilar-Henonin L, Schmid M, Weigel D, Carter DE, Marchand T, Risseeuw E, Brogden D, Zeko A, Crosby WL, Berry CC, Ecker JR (2003) Genome-wide insertional mutagenesis of Arabidopsis thaliana. Science 301:653–657. doi:10.1126/science.1086391
Barkan A (1993) Nuclear mutants of maize with defects in chloroplast polysome assembly have altered chloroplast RNA metabolism. Plant Cell 5:389–402
Barkan A (1998) Approaches to investigating nuclear genes that function in chloroplast biogenesis in land plants. Methods Enzymol 297:38–57. doi:10.1016/S0076-6879(98)97006-9
Baumgartner BJ, Rapp JC, Mullet JE (1993) Plastid genes encoding the transcription-translation apparatus are differentially transcribed early in barley (Hordeum vulgare) chloroplast development: evidence for selective stabilization of psbA mRNA. Plant Physiol 101:781–791
Beligni MV, Mayfield SP (2008) Arabidopsis thaliana mutants reveal a role for CSP41a and CSP41b two ribosome-associated endonucleases in chloroplast ribosomal RNA metabolism. Plant Mol Biol 67:389–401
Bollenbach TJ, Stern DB (2003a) Divalent metal-dependent catalysis and cleavage specificity of CSP41, a chloroplast endoribonuclease belonging to the short chain dehydrogenase/reductase superfamily. Nucleic Acids Res 31:4317–4325. doi:10.1093/nar/gkg640
Bollenbach TJ, Stern DB (2003b) Secondary structures common to chloroplast mRNA 3′-untranslated regions direct cleavage by CSP41, an endoribonuclease belonging to the short chain dehydrogenase/reductase superfamily. J Biol Chem 278:25832–25838. doi:10.1074/jbc.M303559200
Bollenbach TJ, Tatman DA, Stern DB (2003) CSP41a, a multifunctional RNA-binding protein, initiates mRNA turnover in tobacco chloroplasts. Plant J 36:842–852. doi:10.1046/j.1365-313X.2003.01935.x
Bollenbach TJ, Lange H, Gutierrez R, Erhardt M, Stern DB, Gagliardi D (2005) RNR1, a 3′–5′ exoribonuclease belonging to the RNR superfamily, catalyzes 3′ maturation of chloroplast ribosomal RNAs in Arabidopsis thaliana. Nucleic Acids Res 33:2751–2763. doi:10.1093/nar/gki576
Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743. doi:10.1046/j.1365-313x.1998.00343.x
Dohme F, Nierhaus KH (1976) Role of 5S RNA in assembly and function of the 50S subunit from Escherichia coli. Proc Natl Acad Sci USA 73:2221–2225. doi:10.1073/pnas.73.7.2221
Fish RN, Kane CM (2002) Promoting elongation with transcript cleavage stimulatory factors. Biochim Biophys Acta 1577:287–307
Gatenby AA, Rothstein SJ, Nomura M (1989) Translational coupling of the maize chloroplast atpB and atpE genes. Proc Natl Acad Sci USA 86:4066–4070. doi:10.1073/pnas.86.11.4066
Green R, Noller HF (1999) Reconstitution of functional 50S ribosomes from in vitro transcripts of Bacillus stearothermophilus 23S rRNA. Biochemistry 38:1772–1779. doi:10.1021/bi982246a
Hassidim M, Yakir E, Fradkin D, Hilman D, Kron I, Keren N, Harir Y, Yerushalmi S, Green RM (2007) Mutations in CHLOROPLAST RNA BINDING provide evidence for the involvement of the chloroplast in the regulation of the circadian clock in Arabidopsis. Plant J 51:551–562. doi:10.1111/j.1365-313X.2007.03160.x
Hellens RP, Edwards EA, Leyland NR, Bean S, Mullineaux PM (2000) pGreen: a versatile and flexible binary Ti vector for Agrobacterium-mediated plant transformation. Plant Mol Biol 42:819–832. doi:10.1023/A:1006496308160
Horlitz M, Klaff P (2000) Gene-specific trans-regulatory functions of magnesium for chloroplast mRNA stability in higher plants. J Biol Chem 275:35638–35645. doi:10.1074/jbc.M005622200
Kettunen R, Pursiheimo S, Rintamaki E, Van Wijk KJ, Aro EM (1997) Transcriptional and translational adjustments of psbA gene expression in mature chloroplasts during photoinhibition and subsequent repair of photosystem II. Eur J Biochem 247:441–448. doi:10.1111/j.1432-1033.1997.00441.x
Khaitovich P, Tenson T, Kloss P, Mankin AS (1999) Reconstitution of functionally active Thermus aquaticus large ribosomal subunits with in vitro-transcribed rRNA. Biochemistry 38:1780–1788. doi:10.1021/bi9822473
Klaff P, Gruissem W (1991) Changes in chloroplast mRNA stability during leaf development. Plant Cell 3:517–530
Loschelder H, Schweer J, Link B, Link G (2006) Dual temporal role of plastid Sigma Factor 6 in Arabidopsis development. Plant Physiol 142:642–650
Monde RA, Greene JC, Stern DB (2000a) Disruption of the petB-petD intergenic region in tobacco chloroplasts affects petD RNA accumulation and translation. Mol Gen Genet 263:610–618. doi:10.1007/s004380051208
Monde RA, Zito F, Olive J, Wollman FA, Stern DB (2000b) Post-transcriptional defects in tobacco chloroplast mutants lacking the cytochrome b6/f complex. Plant J 21:61–72. doi:10.1046/j.1365-313x.2000.00653.x
Mullet JE, Klein RR (1987) Transcription and RNA stability are important determinants of higher plant chloroplast RNA levels. EMBO J 6:1571–1579
Nakamura T, Ohta M, Sugiura M, Sugita M (2001) Chloroplast ribonucleoproteins function as a stabilizing factor of ribosome-free mRNAs in the stroma. J Biol Chem 276:147–152. doi:10.1074/jbc.M008817200
Peltier JB, Cai Y, Sun Q, Zabrouskov V, Giacomelli L, Rudella A, Ytterberg AJ, Rutschow H, van Wijk KJ (2006) The oligomeric stromal proteome of Arabidopsis thaliana chloroplasts. Mol Cell Proteomics 5:114–133. doi:10.1074/mcp.M500180-MCP200
Pfannschmidt T, Ogrzewalla K, Baginsky S, Sickmann A, Meyer HE, Link G (2000) The multisubunit chloroplast RNA polymerase A from mustard (Sinapis alba L.). Integration of a prokaryotic core into a larger complex with organelle-specific functions. Eur J Biochem 267:253–261. doi:10.1046/j.1432-1327.2000.00991.x
Raab S, Toth Z, de Groot C, Stamminger T, Hoth S (2006) ABA-responsive RNA-binding proteins are involved in chloroplast and stromule function in Arabidopsis seedlings. Planta: 224:900–914. doi:10.1007/s00425-006-0282-4
Schweer J, Loschelder H, Link G (2006) A promoter switch that can rescue a plant sigma factor mutant. FEBS Lett 580:6617–6622. doi:10.1016/j.febslet.2006.11.010
Spierer P, Zimmermann RA (1978) Stoichiometry, cooperativity, and stability of interactions between 5S RNA and proteins L5, L18, and L25 from the 50S ribosomal subunit of Escherichia coli. Biochemistry 17:2474–2479. doi:10.1021/bi00606a002
Spierer P, Bogdanov AA, Zimmermann RA (1978) Parameters for the interaction of ribosomal proteins L5, L18, and L25 with 5S RNA from Escherichia coli. Biochemistry 17:5394–5398. doi:10.1021/bi00618a012
Suzuki JY, Ytterberg AJ, Beardslee TA, Allison LA, Wijk KJ, Maliga P (2004) Affinity purification of the tobacco plastid RNA polymerase and in vitro reconstitution of the holoenzyme. Plant J 40:164–172. doi:10.1111/j.1365-313X.2004.02195.x
Swiatecka-Hagenbruch M, Liere K, Borner T (2007) High diversity of plastidial promoters in Arabidopsis thaliana. Mol Genet Genomics 277:725–734. doi:10.1007/s00438-007-0222-4
Vian A, Henry-Vian C, Davies E (1999) Rapid and systemic accumulation of chloroplast mRNA-binding protein transcripts after flame stimulus in tomato. Plant Physiol 121:517–524. doi:10.1104/pp.121.2.517
Wind M, Reines D (2000) Transcription elongation factor SII. Bioessays 22:327–336. doi:10.1002/(SICI)1521-1878(200004)22:4<327::AID-BIES3>3.0.CO;2-4
Yamaguchi K, Beligni MV, Prieto S, Haynes PA, McDonald WH, Yates JR III, Mayfield SP (2003) Proteomic characterization of the Chlamydomonas reinhardtii chloroplast ribosome. Identification of proteins unique to the 70S ribosome. J Biol Chem 278:33774–33785. doi:10.1074/jbc.M301934200
Yang J, Stern DB (1997) The spinach chloroplast endoribonuclease CSP41 cleaves the 3′ untranslated region of petD mRNA primarily within its terminal stem-loop structure. J Biol Chem 272:12784–12880
Yang J, Schuster G, Stern DB (1996) CSP41, a sequence-specific chloroplast mRNA binding protein, is an endoribonuclease. Plant Cell 8:1409–1420
Acknowledgements
We thank Maureen Hanson’s lab for RpoB antiserum, and the Salk Institute Genomic Analysis Laboratory for T-DNA lines. We also thank Harsh Parikh, a Univ. Waterloo intern, for assistance. This work was supported by DOE Energy Biosciences Program award DE-FG02-90ER20015 to D.B.S.
Author information
Authors and Affiliations
Corresponding author
Additional information
Arabidopsis seed stocks: SALK_107566 (csp41b-1) and SALK_021748 (csp41b-2).
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Bollenbach, T.J., Sharwood, R.E., Gutierrez, R. et al. The RNA-binding proteins CSP41a and CSP41b may regulate transcription and translation of chloroplast-encoded RNAs in Arabidopsis. Plant Mol Biol 69, 541–552 (2009). https://doi.org/10.1007/s11103-008-9436-z
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11103-008-9436-z