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Sigma-like activity from mustard (Sinapis alba L.) chloroplasts conferring DNA-binding and transcription specificity to E. coli core RNA polymerase

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Abstract

A protein fraction which lacks DNA-binding activity itself, but confers enhanced protein-DNA complex formation to E. coli core RNA polymerase, was obtained from mustard chloroplasts by heparin Sepharose chromatography. Gel retardation and competition assays as well as DNase I footprinting experiments with a chloroplast DNA fragment containing the psbA promoter indicate that this reflects sequence-specific binding. Transcription of the psbA template by E. coli core enzyme in the presence of the chloroplast fraction results in enhanced formation of transcripts of the size expected for correct initiation at the in vivo start site. We conclude that the chloroplast fraction reveals sigma-like activity with E. coli RNA polymerase and thus might contain factor(s) of equivalent function in chloroplast transcription.

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References

  1. Berg D, Barrett K, Chamberlin M: Purification of two forms of Escherichia coli RNA polymerase and of sigma component. In: Grossman L, Moldave K (eds) Methods in Enzymology, Vol. 65. Academic Press, New York (1971) pp 499–525.

    Google Scholar 

  2. Bickle TA, Pirrotta V, Imber R: A simple general procedure for purifying restriction endonucleases. Nucleic Acids Res 4: 2561–2572 (1977).

    Google Scholar 

  3. Boyer SK, Mullet JE: Characterization of P. sativum chloroplast psbA transcripts produced in vivo, in vitro and in E. coli. Plant Mol Biol 6: 229–243 (1986).

    Google Scholar 

  4. Bradley D, Gatenby AA: Mutational analysis of the maize chloroplast ATPase-β subunit gene promoter: the isolation of promoter mutants in E. coli and their characterization in a chloroplast in vitro transcription system. EMBO J 4: 3641–3648 (1985).

    Google Scholar 

  5. Briat JF, Dron M, Loiseaux S, Mache R: Structure and transcription of the spinach chloroplast rDNA leader region. Nucleic Acids Res 10: 6865–6878 (1982).

    Google Scholar 

  6. Briat JF, Lescure AM, Mache R: Transcription of the chloroplast DNA: a review. Biochimie 68: 981–990 (1986).

    Google Scholar 

  7. Burgess RR, Travers AA, Dunn JJ, Bautz EKF: Factor stimulating transcription by RNA polymerase. Nature 221: 43–46 (1969).

    Google Scholar 

  8. Carthew RW, Chodosh LA, Sharp PA: An RNA polymerase II transcription factor binds to an upstream element in the adenovirus major late promoter. Cell 43: 439–448 (1985).

    Google Scholar 

  9. Cozens AL, Walker E: Pea chloroplast DNA encodes homologues of Escherichia coli ribosomal subunit S2 and the β′-subunit of RNA polymerase. Biochem J 236: 453–460 (1986).

    Google Scholar 

  10. Duval-Valentin G, Ehrlich R: Interaction between E. coli RNA polymerase and the tetR promoter from pSC101: homologies and differences with other E. coli promoter systems from close contact point studies. Nucleic Acids Res 14: 1967–1983 (1986).

    Google Scholar 

  11. Fried M, Crothers DM: Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res 9: 6505–6525 (1981).

    Google Scholar 

  12. Galas DJ, Schmitz A: DNAase footprinting: a simple method for the detection of protein-DNA binding specificity. Nucleic Acids Res 5: 3157–3168 (1978).

    Google Scholar 

  13. Garner MM, Revzin A: A gel electrophoresis method for quantifying the binding of proteins to specific DNA regions: application to components of the Escherichia coli lactose operon regulatory system. Nucleic Acids Res 9: 3047–3060 (1981).

    Google Scholar 

  14. Garoff H, Ansorge W: Improvements of DNA sequencing gels. Anal Biochem 115: 450–457 (1981).

    Google Scholar 

  15. Greenberg BM, Narita JO, DeLuca-Flaherty CR, Hallick RB: Properties of chloroplast RNA polymerases. In: Steinback KE, Bonitz S, Arntzen C, Bogorad L (eds) Molecular Biology of the Photosynthetic Apparatus. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1985) pp 303–310.

    Google Scholar 

  16. Gruissem W, Zurawski G: Analysis of promoter regions for the spinach chloroplast rbcL, atpB and psbA genes. EMBO J 4: 3375–3383 (1985).

    Google Scholar 

  17. Heumann H, Metzger W, Niehörster M: Visualization of intermediary transcription states in the complex between Escherichia coli DNA-dependent RNA polymerase and a promoter-carrying DNA fragment using the gel retardation method. Eur J Biochem 158: 575–579 (1986).

    Google Scholar 

  18. Hofer B, Müller D, Köster H: The pathway of E. coli RNA polymerase-promoter complex formation as visualized by footprinting. Nucleic Acids Res 13: 5995–6013 (1985).

    Google Scholar 

  19. Jolly SO, Bogorad L: Preferential transcription of cloned maize chloroplast DNA sequences by maize chloroplast RNA polymerase. Proc Natl Acad Sci USA 77: 822–826 (1980).

    Google Scholar 

  20. Kidd GH, Bogorad L: A facile procedure for purifying maize chloroplast RNA polymerase from whole cell homogenates. Biochim Biophys Acta 609: 14–30 (1980).

    Google Scholar 

  21. Kung SD, Lin CM: Chloroplast promoters from higher plants. Nucleic Acids Res 13: 7543–7549 (1985).

    Google Scholar 

  22. Lerbs S, Bräutigam E, Parthier B: Polypeptides of DNA-dependent RNA polymerase of spinach chloroplasts: characterization by antibody-linked polymerase assay and determination of sites of synthesis. EMBO J 4: 1661–1666 (1985).

    Google Scholar 

  23. Lerbs S, Briat JF, Mache R: Chloroplast RNA polymerase from spinach: purification and DNA-binding proteins. Plant Mol Biol 2: 67–74 (1983).

    Google Scholar 

  24. Link G: DNA sequence requirements for the accurate transcription of a protein-coding plastid gene in a plastid in vitro system from mustard (Sinapis alba L.). EMBO J 3: 1697–1704 (1984).

    Google Scholar 

  25. Link G, Langridge U: Structure of the chloroplast gene for the precursor of the Mr 32000 photosystem II protein from mustard (Sinapis alba L.). Nucleic Acids Res 12: 945–957 (1984).

    Google Scholar 

  26. Losick R, Chamberlin MJ: RNA Polymerase. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1976).

    Google Scholar 

  27. Maniatis T, Fritsch EF, Sambrook J: Molecular Cloning, a Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982).

    Google Scholar 

  28. Maxam A, Gilbert W: Sequencing end-labeled DNA with base-specific chemical cleavages. In: Grossman L, Moldave K (eds) Methods in Enzymology, Vol. 65. Academic Press, New York (1980) pp 499–525.

    Google Scholar 

  29. Neuhaus H, Link G: The chloroplast tRNALys(UUU) gene from mustard (Sinapis alba) contains a class II intron potentially coding for a maturase-related polypeptide. Curr Genet 11: 251–257 (1987).

    Google Scholar 

  30. Ohme M, Tanaka M, Chunwongse J, Shinozaki K, Sugiura M: A tobacco chloroplast DNA sequence possibly coding for a polypeptide similar to E. coli RNA polymerase β-subunit. FEBS Lett 200: 87–90 (1986).

    Google Scholar 

  31. Ohyama K, Fukuzawa H, Kohchi T, Shirai H, Sano T, Sano K, Umesono L, Shiki Y, Takeuchi M, Chang Z, Hota S, Inokuchi M, Ozeki H: Chloroplast gene organization deduced from complete sequence of liverwort Marchantia polymorpha chloroplast DNA. Nature 322: 572–574 (1986).

    Google Scholar 

  32. Orozco EM, Mullet JE, Chua NH: An in vitro system for accurate transcription initiation of chloroplast protein genes. Nucleic Acids Res 13: 1283–1302 (1985).

    Google Scholar 

  33. Reiss T, Link G: Characterization of transcriptionally active DNA-protein complexes from chloroplasts and etioplasts of mustard (Sinapis alba L.). Eur J Biochem 148: 207–212 (1985).

    Google Scholar 

  34. Sanger F, Coulson AR: The use of thin acrylamide gels for DNA sequencing. FEBS Lett 87: 107–108 (1978).

    Google Scholar 

  35. Schwarz Z, Kössel H, Schwartz E, Bogorad L: A gene coding for tRNAVal is located near 5′ terminus of 16 S rRNA gene in Zea mays chloroplast genome. Proc Natl Acad Sci USA 78: 4748–4752 (1981).

    Google Scholar 

  36. Shinozaki K, Ohme M, Tanaka M, Wakasugi T, Hayashida N, Matsubayashi T, Zaita N, Chunwongse J, Obokata J, Yamaguchi-Shinozaki K, Ohto C, Torazada K, Meng BY, Sugita M, Deno H, Kamogashira T, Yamada K, Kusuda J, Takaiwa F, Kato A, Tohdoh N, Shimada H, Sugiura M: The complete nucleotide sequence of the tobacco chloroplast genome: its gene organization and expression. EMBO J 5: 2043–2049 (1986).

    Google Scholar 

  37. Sijben-Müller G, Hallick RB, Alt J, Westhoff P, Herrmann RG: Spinach plastid genes coding for initiation factor IF-1, ribosomal protein S11 and RNA polymerase α-subunit. Nucleic Acids Res 14: 1029–1044 (1986).

    Google Scholar 

  38. Smith HJ, Bogorad L: The polypeptide subunit structure of the DNA-dependent RNA polymerase of Zea mays chloroplasts. Proc Natl Acad Sci USA 71: 4839–4842 (1974).

    Google Scholar 

  39. Straney DC, Crothers DM: Intermediates in transcription initiation from the E. coli lac UV5 promoter. Cell 43: 449–459 (1985).

    Google Scholar 

  40. Surzycki SJ, Shellenbarger DL: Purification and characterization of a putative sigma factor from Chlamydomonas reinhardii. Proc Natl Acad Sci USA 73: 3961–3965 (1976).

    Google Scholar 

  41. Tewari KK, Goel A: Solubilization and partial purification of RNA polymerase from pea chloroplasts. Biochemistry 22: 2142–2148 (1983).

    Google Scholar 

  42. Whitfeld PR, Bottomley W: Organization and structure of chloroplast genes. Annu Rev Plant Physiol 34: 279–310 (1983).

    Google Scholar 

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Bülow, S., Link, G. Sigma-like activity from mustard (Sinapis alba L.) chloroplasts conferring DNA-binding and transcription specificity to E. coli core RNA polymerase. Plant Mol Biol 10, 349–357 (1988). https://doi.org/10.1007/BF00029885

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  • DOI: https://doi.org/10.1007/BF00029885

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