Skip to main content
NCBI home page
Plant Physiology logoLink to Plant Physiology
. 1988 Dec;88(4):1002–1007. doi: 10.1104/pp.88.4.1002

The Synthesis of a 19 Kilodalton Zein Protein in Transgenic Petunia Plants 1

John D Williamson 1,2, Gad Galili 1,2,2, Brian A Larkins 1,2, Stanton B Gelvin 1,2
PMCID: PMC1055705  PMID: 16666411

Abstract

Transcriptional fusions composed of a 19 kilodalton zein cDNA, the 5′ flanking region from a β-phaseolin gene, and 3′ flanking regions from either the phaseolin or a 15-kilodalton zein gene were introduced into Petunia by Agrobacterium-mediated transformation. The expression of both zein mRNA and protein in these transgenic plants was seed-specific and developmentally regulated. Both monocot (zein) and dicot (phaseolin) polyadenylation consensus sequences were recognized in Petunia. Analysis by immunoblotting showed that the Mr of the zein protein corresponded to that of the mature protein, suggesting that recognition and cleavage of the signal sequence had occurred. While zein mRNA accumulated to approximately 1% of the total poly(A)+ RNA in seeds of the transformed plants, zein protein was present at a much lower concentration than expected, at most being 0.005% of the total seed protein. These results suggest that the 19 kilodalton zein gene, in addition to lacking specific sequences required for efficient transcription in dicots, might also lack sequences required for the efficient synthesis, targeting, transport, or stabilization of the protein.

Full text

PDF
1002

Images in this article

1004Fig. 2
on p.1004
1005Fig. 3
on p.1005
1006Fig. 4
on p.1006

Selected References

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

  1. A simple and general method for transferring genes into plants. Science. 1985 Mar 8;227(4691):1229–1231. doi: 10.1126/science.227.4691.1229. [DOI] [PubMed] [Google Scholar]
  2. An G. Development of plant promoter expression vectors and their use for analysis of differential activity of nopaline synthase promoter in transformed tobacco cells. Plant Physiol. 1986 May;81(1):86–91. doi: 10.1104/pp.81.1.86. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Argos P., Pedersen K., Marks M. D., Larkins B. A. A structural model for maize zein proteins. J Biol Chem. 1982 Sep 10;257(17):9984–9990. [PubMed] [Google Scholar]
  4. Beachy R. N., Chen Z. L., Horsch R. B., Rogers S. G., Hoffmann N. J., Fraley R. T. Accumulation and assembly of soybean beta-conglycinin in seeds of transformed petunia plants. EMBO J. 1985 Dec 1;4(12):3047–3053. doi: 10.1002/j.1460-2075.1985.tb04044.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bevan M. Binary Agrobacterium vectors for plant transformation. Nucleic Acids Res. 1984 Nov 26;12(22):8711–8721. doi: 10.1093/nar/12.22.8711. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1016/0003-2697(76)90527-3. [DOI] [PubMed] [Google Scholar]
  7. Das O. P., Messing J. W. Allelic variation and differential expression at the 27-kilodalton zein locus in maize. Mol Cell Biol. 1987 Dec;7(12):4490–4497. doi: 10.1128/mcb.7.12.4490. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ditta G., Stanfield S., Corbin D., Helinski D. R. Broad host range DNA cloning system for gram-negative bacteria: construction of a gene bank of Rhizobium meliloti. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7347–7351. doi: 10.1073/pnas.77.12.7347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Galili G., Kawata E. E., Cuellar R. E., Smith L. D., Larkins B. A. Synthetic oligonucleotide tails inhibit in vitro and in vivo translation of SP6 transcripts of maize zein cDNA clones. Nucleic Acids Res. 1986 Feb 11;14(3):1511–1524. doi: 10.1093/nar/14.3.1511. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Greenwood J. S., Chrispeels M. J. Correct targeting of the bean storage protein phaseolin in the seeds of transformed tobacco. Plant Physiol. 1985 Sep;79(1):65–71. doi: 10.1104/pp.79.1.65. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hagen G., Rubenstein I. Complex organization of zein genes in maize. Gene. 1981 Apr;13(3):239–249. doi: 10.1016/0378-1119(81)90029-9. [DOI] [PubMed] [Google Scholar]
  12. Hoffman L. M., Donaldson D. D., Bookland R., Rashka K., Herman E. M. Synthesis and protein body deposition of maize 15-kd zein in transgenic tobacco seeds. EMBO J. 1987 Nov;6(11):3213–3221. doi: 10.1002/j.1460-2075.1987.tb02638.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jackson R. C., Blobel G. Post-translational cleavage of presecretory proteins with an extract of rough microsomes from dog pancreas containing signal peptidase activity. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5598–5602. doi: 10.1073/pnas.74.12.5598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Keith B., Chua N. H. Monocot and dicot pre-mRNAs are processed with different efficiencies in transgenic tobacco. EMBO J. 1986 Oct;5(10):2419–2425. doi: 10.1002/j.1460-2075.1986.tb04516.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  16. Larkins B. A., Hurkman W. J. Synthesis and deposition of zein in protein bodies of maize endosperm. Plant Physiol. 1978 Aug;62(2):256–263. doi: 10.1104/pp.62.2.256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Marks M. D., Lindell J. S., Larkins B. A. Nucleotide sequence analysis of zein mRNAs from maize endosperm. J Biol Chem. 1985 Dec 25;260(30):16451–16459. [PubMed] [Google Scholar]
  18. Matzke M. A., Susani M., Binns A. N., Lewis E. D., Rubenstein I., Matzke A. J. Transcription of a zein gene introduced into sunflower using a Ti plasmid vector. EMBO J. 1984 Jul;3(7):1525–1531. doi: 10.1002/j.1460-2075.1984.tb02006.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Osborne T. B. OUR PRESENT KNOWLEDGE OF PLANT PROTEINS. Science. 1908 Oct 2;28(718):417–427. doi: 10.1126/science.28.718.417. [DOI] [PubMed] [Google Scholar]
  20. Pedersen K., Argos P., Naravana S. V., Larkins B. A. Sequence analysis and characterization of a maize gene encoding a high-sulfur zein protein of Mr 15,000. J Biol Chem. 1986 May 15;261(14):6279–6284. [PubMed] [Google Scholar]
  21. Pedersen K., Devereux J., Wilson D. R., Sheldon E., Larkins B. A. Cloning and sequence analysis reveal structural variation among related zein genes in maize. Cell. 1982 Jul;29(3):1015–1026. doi: 10.1016/0092-8674(82)90465-2. [DOI] [PubMed] [Google Scholar]
  22. Roussell D. L., Boston R. S., Goldsbrough P. B., Larkins B. A. Deletion of DNA sequences flanking an Mr 19,000 zein gene reduces its transcriptional activity in heterologous plant tissues. Mol Gen Genet. 1988 Feb;211(2):202–209. doi: 10.1007/BF00330595. [DOI] [PubMed] [Google Scholar]
  23. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Schernthaner J. P., Matzke M. A., Matzke A. J. Endosperm-specific activity of a zein gene promoter in transgenic tobacco plants. EMBO J. 1988 May;7(5):1249–1255. doi: 10.1002/j.1460-2075.1988.tb02938.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sengupta-Gopalan C., Reichert N. A., Barker R. F., Hall T. C., Kemp J. D. Developmentally regulated expression of the bean beta-phaseolin gene in tobacco seed. Proc Natl Acad Sci U S A. 1985 May;82(10):3320–3324. doi: 10.1073/pnas.82.10.3320. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  27. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ueng P., Galili G., Sapanara V., Goldsbrough P. B., Dube P., Beachy R. N., Larkins B. A. Expression of a maize storage protein gene in petunia plants is not restricted to seeds. Plant Physiol. 1988 Apr;86(4):1281–1285. doi: 10.1104/pp.86.4.1281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wilson D. R., Larkins B. A. Zein gene organization in maize and related grasses. J Mol Evol. 1984;20(3-4):330–340. doi: 10.1007/BF02104739. [DOI] [PubMed] [Google Scholar]

Articles from Plant Physiology are provided here courtesy of Oxford University Press

RESOURCES