Skip to main content
PMC home page
The Plant Cell logoLink to The Plant Cell
. 1990 Jul;2(7):659–671. doi: 10.1105/tpc.2.7.659

Complex RNA maturation pathway for a chloroplast ribosomal protein operon with an internal tRNA cistron.

D A Christopher 1, R B Hallick 1
PMCID: PMC159920  PMID: 2136640

Abstract

We have studied the expression of a large chloroplast ribosomal protein operon from Euglena gracilis that resembles the Escherichia coli S10 and spc ribosomal protein operons. We present evidence that 11 ribosomal protein genes, a tRNA gene, and a new locus, orf214/orf302, are expressed as a single transcription unit. The primary transcript also contains at least 15 group II and group III introns. Gene-specific probes for each ribosomal protein gene, orf214/orf302, and for trnl hybridized to a common pre-mRNA of an estimated size of 8.3 kilobases. This is the RNA size predicted for a full-length transcript of the entire operon after splicing of all 15 introns. Polycistronic ribosomal protein mRNAs accumulated primarily as spliced hepta-, hexa-, penta-, tetra-, tri-, and dicistronic mRNAs, which were presumed to arise by stepwise processing of the 8.3-kilobase pre-mRNA. A novel finding was the cotranscription of the trnl gene as an internal cistron within the ribosomal protein operon. Several combined mRNA/tRNA molecules, such as the pentacistronic rpl5-rps8-rpl36-trnl-rps14, were characterized. The occurrence of the orf214/orf302 is a unique feature of the Euglena operon, distinguishing it from all chloroplast and prokaryotic ribosomal protein operons characterized to date. The orf214/orf302 are not similar to any known genes but are cotranscribed with the ribosomal protein loci and encode stable RNA species of 2.4, 1.8, and 1.4 kilobases.

Full Text

The Full Text of this article is available as a PDF (3.3 MB).

Selected References

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

  1. Barkan A. Proteins encoded by a complex chloroplast transcription unit are each translated from both monocistronic and polycistronic mRNAs. EMBO J. 1988 Sep;7(9):2637–2644. doi: 10.1002/j.1460-2075.1988.tb03116.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Barkan A. Tissue-dependent plastid RNA splicing in maize: transcripts from four plastid genes are predominantly unspliced in leaf meristems and roots. Plant Cell. 1989 Apr;1(4):437–445. doi: 10.1105/tpc.1.4.437. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berends T., Gamble P. E., Mullet J. E. Characterization of the barley chloroplast transcription units containing psaA-psaB and psbD-psbC. Nucleic Acids Res. 1987 Jul 10;15(13):5217–5240. doi: 10.1093/nar/15.13.5217. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cerretti D. P., Dean D., Davis G. R., Bedwell D. M., Nomura M. The spc ribosomal protein operon of Escherichia coli: sequence and cotranscription of the ribosomal protein genes and a protein export gene. Nucleic Acids Res. 1983 May 11;11(9):2599–2616. doi: 10.1093/nar/11.9.2599. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Christopher D. A., Cushman J. C., Price C. A., Hallick R. B. Organization of ribosomal protein genes rpl23, rpl2, rps19, rpl22 and rps3 on the Euglena gracilis chloroplast genome. Curr Genet. 1988 Sep;14(3):275–285. doi: 10.1007/BF00376748. [DOI] [PubMed] [Google Scholar]
  6. Clayton D. A. Transcription of the mammalian mitochondrial genome. Annu Rev Biochem. 1984;53:573–594. doi: 10.1146/annurev.bi.53.070184.003041. [DOI] [PubMed] [Google Scholar]
  7. Cushman J. C., Christopher D. A., Little M. C., Hallick R. B., Price C. A. Organization of the psbE, psbF, orf38, and orf42 gene loci on the Euglena gracilis chloroplast genome. Curr Genet. 1988 Feb;13(2):173–180. doi: 10.1007/BF00365652. [DOI] [PubMed] [Google Scholar]
  8. Cushman J. C., Hallick R. B., Price C. A. The two genes for the P700 chlorophyll a apoproteins on the Euglena gracilis chloroplast genome contain multiple introns. Curr Genet. 1988 Feb;13(2):159–171. doi: 10.1007/BF00365651. [DOI] [PubMed] [Google Scholar]
  9. Deng X. W., Gruissem W. Constitutive transcription and regulation of gene expression in non-photosynthetic plastids of higher plants. EMBO J. 1988 Nov;7(11):3301–3308. doi: 10.1002/j.1460-2075.1988.tb03200.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Deng X. W., Gruissem W. Control of plastid gene expression during development: the limited role of transcriptional regulation. Cell. 1987 May 8;49(3):379–387. doi: 10.1016/0092-8674(87)90290-x. [DOI] [PubMed] [Google Scholar]
  11. Dix D. B., Thompson R. C. Codon choice and gene expression: synonymous codons differ in translational accuracy. Proc Natl Acad Sci U S A. 1989 Sep;86(18):6888–6892. doi: 10.1073/pnas.86.18.6888. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gingrich J. C., Hallick R. B. The Euglena gracilis chloroplast ribulose-1,5-bisphosphate carboxylase gene. I. Complete DNA sequence and analysis of the nine intervening sequences. J Biol Chem. 1985 Dec 25;260(30):16156–16161. [PubMed] [Google Scholar]
  13. Gruissem W., Barkan A., Deng X. W., Stern D. Transcriptional and post-transcriptional control of plastid mRNA levels in higher plants. Trends Genet. 1988 Sep;4(9):258–263. doi: 10.1016/0168-9525(88)90033-9. [DOI] [PubMed] [Google Scholar]
  14. Gruissem W., Zurawski G. Identification and mutational analysis of the promoter for a spinach chloroplast transfer RNA gene. EMBO J. 1985 Jul;4(7):1637–1644. doi: 10.1002/j.1460-2075.1985.tb03831.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hallick R. B., Chelm B. K., Gray P. W., Orozco E. M., Jr Use of aurintricarboxylic acid as an inhibitor of nucleases during nucleic acid isolation. Nucleic Acids Res. 1977 Sep;4(9):3055–3064. doi: 10.1093/nar/4.9.3055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hiratsuka J., Shimada H., Whittier R., Ishibashi T., Sakamoto M., Mori M., Kondo C., Honji Y., Sun C. R., Meng B. Y. The complete sequence of the rice (Oryza sativa) chloroplast genome: intermolecular recombination between distinct tRNA genes accounts for a major plastid DNA inversion during the evolution of the cereals. Mol Gen Genet. 1989 Jun;217(2-3):185–194. doi: 10.1007/BF02464880. [DOI] [PubMed] [Google Scholar]
  17. Hollingsworth M. J., Johanningmeier U., Karabin G. D., Stiegler G. L., Hallick R. B. Detection of multiple, unspliced precursor mRNA transcripts for the Mr 32,000 thylakoid membrane protein from Euglena gracilis chloroplasts. Nucleic Acids Res. 1984 Feb 24;12(4):2001–2017. doi: 10.1093/nar/12.4.2001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hudson G. S., Mason J. G., Holton T. A., Koller B., Cox G. B., Whitfeld P. R., Bottomley W. A gene cluster in the spinach and pea chloroplast genomes encoding one CF1 and three CF0 subunits of the H+-ATP synthase complex and the ribosomal protein S2. J Mol Biol. 1987 Jul 20;196(2):283–298. doi: 10.1016/0022-2836(87)90690-5. [DOI] [PubMed] [Google Scholar]
  19. Karabin G. D., Farley M., Hallick R. B. Chloroplast gene for Mr 32000 polypeptide of photosystem II in Euglena gracilis is interrupted by four introns with conserved boundary sequences. Nucleic Acids Res. 1984 Jul 25;12(14):5801–5812. doi: 10.1093/nar/12.14.5801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kashdan M. A., Dudock B. S. The gene for a spinach chloroplast isoleucine tRNA has a methionine anticodon. J Biol Chem. 1982 Oct 10;257(19):11191–11194. [PubMed] [Google Scholar]
  21. Klein R. R., Mason H. S., Mullet J. E. Light-regulated translation of chloroplast proteins. I. Transcripts of psaA-psaB, psbA, and rbcL are associated with polysomes in dark-grown and illuminated barley seedlings. J Cell Biol. 1988 Feb;106(2):289–301. doi: 10.1083/jcb.106.2.289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lindahl L., Zengel J. M. Ribosomal genes in Escherichia coli. Annu Rev Genet. 1986;20:297–326. doi: 10.1146/annurev.ge.20.120186.001501. [DOI] [PubMed] [Google Scholar]
  23. Mattheakis L. C., Nomura M. Feedback regulation of the spc operon in Escherichia coli: translational coupling and mRNA processing. J Bacteriol. 1988 Oct;170(10):4484–4492. doi: 10.1128/jb.170.10.4484-4492.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 1984 Sep 25;12(18):7035–7056. doi: 10.1093/nar/12.18.7035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Montandon P. E., Stutz E. Nucleotide sequence of a Euglena gracilis chloroplast genome region coding for the elongation factor Tu; evidence for a spliced mRNA. Nucleic Acids Res. 1983 Sep 10;11(17):5877–5892. doi: 10.1093/nar/11.17.5877. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Montandon P. E., Stutz E. The genes for the ribosomal proteins S12 and S7 are clustered with the gene for the EF-Tu protein on the chloroplast genome of Euglena gracilis. Nucleic Acids Res. 1984 Mar 26;12(6):2851–2859. doi: 10.1093/nar/12.6.2851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Nickoloff J. A., Christopher D. A., Drager R. G., Hallick R. B. Nucleotide sequence of the Euglena gracilis chloroplast genes for isoleucine, phenylalanine and cysteine transfer RNAs and ribosomal protein S14. Nucleic Acids Res. 1989 Jun 26;17(12):4882–4882. doi: 10.1093/nar/17.12.4882. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Nomura M., Morgan E. A. Genetics of bacterial ribosomes. Annu Rev Genet. 1977;11:297–347. doi: 10.1146/annurev.ge.11.120177.001501. [DOI] [PubMed] [Google Scholar]
  29. Shinozaki K., Ohme M., Tanaka M., Wakasugi T., Hayashida N., Matsubayashi T., Zaita N., Chunwongse J., Obokata J., Yamaguchi-Shinozaki K. The complete nucleotide sequence of the tobacco chloroplast genome: its gene organization and expression. EMBO J. 1986 Sep;5(9):2043–2049. doi: 10.1002/j.1460-2075.1986.tb04464.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sijben-Müller G., Hallick R. B., Alt J., Westhoff P., Herrmann R. G. Spinach plastid genes coding for initiation factor IF-1, ribosomal protein S11 and RNA polymerase alpha-subunit. Nucleic Acids Res. 1986 Jan 24;14(2):1029–1044. doi: 10.1093/nar/14.2.1029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Tanaka M., Wakasugi T., Sugita M., Shinozaki K., Sugiura M. Genes for the eight ribosomal proteins are clustered on the chloroplast genome of tobacco (Nicotiana tabacum): similarity to the S10 and spc operons of Escherichia coli. Proc Natl Acad Sci U S A. 1986 Aug;83(16):6030–6034. doi: 10.1073/pnas.83.16.6030. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Thomas F., Massenet O., Dorne A. M., Briat J. F., Mache R. Expression of the rpl23, rpl2 and rps19 genes in spinach chloroplasts. Nucleic Acids Res. 1988 Mar 25;16(6):2461–2472. doi: 10.1093/nar/16.6.2461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Westhoff P., Herrmann R. G. Complex RNA maturation in chloroplasts. The psbB operon from spinach. Eur J Biochem. 1988 Feb 1;171(3):551–564. doi: 10.1111/j.1432-1033.1988.tb13824.x. [DOI] [PubMed] [Google Scholar]
  34. Yepiz-Plascencia G. M., Radebaugh C. A., Hallick R. B. The Euglena gracilis chloroplast rpoB gene. Novel gene organization and transcription of the RNA polymerase subunit operon. Nucleic Acids Res. 1990 Apr 11;18(7):1869–1878. doi: 10.1093/nar/18.7.1869. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Zhou D. X., Quigley F., Massenet O., Mache R. Cotranscription of the S10- and spc-like operons in spinach chloroplasts and identification of three of their gene products. Mol Gen Genet. 1989 Apr;216(2-3):439–445. doi: 10.1007/BF00334388. [DOI] [PubMed] [Google Scholar]

Articles from The Plant Cell are provided here courtesy of Oxford University Press

RESOURCES