Skip to main content
SpringerLink
Log in

Sequence analysis of 22 kDa-like α-coixin genes and their comparison with homologous zein and kafirin genes reveals highly conserved protein structure and regulatory elements

  • Research Articles
  • Published:
Plant Molecular Biology Aims and scope Submit manuscript

Abstract

Several genomic and cDNA clones encoding the 22 kDa-like α-coixin, the α-prolamin of Coix seeds, were isolated and sequenced. Three contiguous 22 kDa-like α-coixin genes designated α-3A, α-3B and α-3C were found in the 15 kb α-3 genomic clone. The α-3A and α-3C genes presented in-frame stop codons at position +652. The two genes with truncated ORFs are flanking the α-3B gene, suggesting that the three α-coixin genes may have arisen by tandem duplication and that the stop codon was introduced before the duplication.

Comparison of the deduced amino acid sequences of α-coixin clones with the published sequences of 22 kDa α-zein and 22 kDa-like α-kafirin revealed a highly conserved protein structure. The protein consists of an N-terminus, containing the signal peptide, followed by ten highly conserved tandem repeats of 15–20 amino acids flanked by polyglutamines, and a short C-terminus. The difference between the 22 kDa-like α-prolamins and the 19 kDa α-zein lies in the fact that the 19 kDa protein is exactly one repeat motif shorter than the 22 kDa proteins.

Several putative regulatory sequences common to the zein and kafirin genes were identified within both the 5′ and 3′ flanking regions of α-3B. Nucleotide sequences that match the consensus TATA, CATC and the ca. −300 prolamin box are present at conserved positions in α-3B relative to zein and kafirin genes. Two putative Opaque-2 boxes are present in α-3B that occupies approximately the same positions as those identified for the 22 kDa α-zein and α-kafirin genes. Southern hybridization, using a fragment of a maize Opaque-2 cDNA clone as a probe, confirmed the presence of Opaque-2 homologous sequences in the Coix and sorghum genomes.

The overall results suggest that the structural and regulatory genes involved in the expression of the 22 kDa-like α-prolamin genes of Coix, sorghum and maize, originated from a common ancestor, and that variations were introduced in the structural and regulatory sequences after species separation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Anderson OD: Characterization of members of a pseudogene subfamily of the wheat α-gliadin storage protein genes. Plant Mol Biol 16: 335–337 (1991).

    Article  PubMed  Google Scholar 

  2. Argos P, Pedersen K, Marks MD, Larkins BA: A structural model for maize zein proteins. J Biol Chem 257: 9984–9990 (1982).

    PubMed  Google Scholar 

  3. Burr B, Burr FA, John TP, Thomas M, Davis RD: Zein storage protein family of maize. J Mol Biol 154: 33–49 (1982).

    PubMed  Google Scholar 

  4. Soave C, Righetti PG, Lorenzoni C, Gentinetta E, Salamini F: Expressivity of the opaque-2 gene at the level of zein molecular components. Maydica 21: 61–75 (1976).

    Google Scholar 

  5. Das OP, Messing JW: Allelic variation and differential expression at the 27-kilodalton zein locus in maize. Mol Cell Biol 7: 4490–4497 (1987).

    PubMed  Google Scholar 

  6. Das OP, Poliak E, Ward K, Messing J: A new allele of the duplicated 27 kDa zein locus of maize generated by homologous recombination. Nucl Acids Res 19: 3325–3330 (1991).

    PubMed  Google Scholar 

  7. DeRose RT, Ma DP, Kwon IJ, Hasmain SE, Klassy RC, Hall TC: Characterization of the kafirin gene family from sorghum reveals extensive homology with zein from maize. Plant Mol Biol 12: 245–256 (1989).

    Google Scholar 

  8. Entwistle J, Knudsen S, Muller M, Cameron-Mills: Amber codon suppression: the in vivo analysis of two C-hordein genes from barley. Plant Mol Biol 17: 1217–1231 (1991).

    PubMed  Google Scholar 

  9. Esen A: Separation of alcohol-soluble protein (zeins) from maize into three fractions by differential solubility. Plant Physiol 80: 623–627 (1986).

    Google Scholar 

  10. Forde BG, Heyworth A, Pywell J, Kreis M: Nucleotide sequence of a B1 hordein gene and the identification of possible upstream regulatory elements in endosperm storage protein genes from barley, wheat and maize. Nucl Acids Res 13: 7327–7339 (1985).

    PubMed  Google Scholar 

  11. Garrat R, Oliva G, Caracelli I, Leite A, Arruda P: Studies of the zein-like α-prolamins based on an analysis of amino acid sequences: Implications to their evolution and three-dimensional structure. Proteins: Structure, Function and Genetics (in press).

  12. Gianazza E, Righetti PG, Pioli F, Galante E, Soave C: Size and charge heterogeneity of zein in normal and opaque-2 maize endosperm. Maydica 21: 1–17 (1976).

    Google Scholar 

  13. Hagen G, Rubenstein I: Complex organization of zein genes in maize. Gene 13: 239–249 (1981).

    PubMed  Google Scholar 

  14. Hartings H, Maddaloni M, Lazzaroni N, DiFonzo N, Motto M, Salamini F, Thompson R: The O2 which regulates zein deposition in maize endosperm encodes a protein with structural homologies to transcriptional activators. EMBO J 8: 2795–2801 (1989).

    PubMed  Google Scholar 

  15. Heidecker G, Chaudhuri S, Messing J: Highly clustered zein gene sequences reveal evolutionary history of the multigene family. Genomics 10: 719–732 (1991).

    PubMed  Google Scholar 

  16. Kreis M, Shewry PR, Forde BG, Forde J, Miflin BJ: Structure and evolution of seed storage proteins and their genes with particular reference to those of wheat, barley and rye. Oxford Surv Plant Mol Cell Biol 2: 253–317 (1985).

    Google Scholar 

  17. Kodrzycki R, Boston RS, Larkins BA: The opaque-2 mutation of maize differentially reduces zein gene transcription. Plant Cell 1: 105–114 (1989).

    PubMed  Google Scholar 

  18. Kriz AL, Boston RS, Larkins BA: Structural and transcriptional analysis of DNA sequences flanking genes that encode 19 kDa zeins. Mol Gen Genet 207: 90–98 (1982).

    Article  Google Scholar 

  19. Laemmli UK: Cleavage of structural proteins during assembly of phage T4 head. Nature 227: 680–685 (1970).

    PubMed  Google Scholar 

  20. Larkins BA, Hurkman WJ: Synthesis and deposition of zein in protein bodies of maize endosperm. Plant Physiol 62: 256–263 (1978).

    Google Scholar 

  21. Leite A, Freitas FA, Yunes JA, Arruda P: Nucleotide sequence of a cDNA clone encoding γ-coixin from Coix lacryma-jobi seeds. Plant Physiol 97: 1604–1605 (1991).

    Google Scholar 

  22. Leite A, Ottoboni LMM, Targor MLPN, Silva MJ, Turcinelli SR, Arruda P: Phylogenetic relationship of zeins and coixins as determined by immunological cross-reactivity and Southern blot analysis. Plant Mol Biol 14: 743–751 (1990).

    PubMed  Google Scholar 

  23. Leite A, Yunes JA, Turcinelli SR, Arruda P: Cloning and characterization of a cDNA encoding a sulfur-rich coixin. Plant Mol Biol 18: 171–174 (1992).

    PubMed  Google Scholar 

  24. Ludevid MD, Martinez-Isquierdo JA, Armengol M, Torrent M, Puigdomenech P, Palau J: Immunological relations between glutelin-2 and low molecular weight zein-2 proteins from maize (Zea mays L.) endosperm. Plant Sci 41: 41–48.

  25. Maiatis T, Fritsch EF, Sambrook J: Molecular Cloning. A Laboratory Manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982).

    Google Scholar 

  26. Marks MD, Larkins BA: Analysis of sequence microheterogeneity among messenger RNAS. J Biol Chem 257: 9976–9983 (1982).

    PubMed  Google Scholar 

  27. Marks MD, Lindell JS, Larkins BA: Nucleotide sequence analysis of zein mRNAs from maize endosperm. J Biol Chem 260: 16451–16459 (1985).

    PubMed  Google Scholar 

  28. Melcher B: Isolation of messenger RNA from membrane bound polysomes. Meth Enzymol 152: 241–248 (1987).

    PubMed  Google Scholar 

  29. Ottoboni LMM, Leite A, Targon MLPN, Crozier A, Arruda P: Characterization of the storage protein in seed of Coix lacryma-jobi var. Adlay. J Agric Food Chem 381: 631–635 (1990).

    Google Scholar 

  30. Ottoboni LMM, Leite A, Targon MLPN, Silva MJ, Arruda P: Heterogeneity of Coix, maize and teosinte prolamins detected by isoelectric focusing. Rev Brass Genet 13: 313–322 (1990).

    Google Scholar 

  31. Pederson K, Argos P, Naravana SVL, Larkins BA: Sequence analysis and characterization of a maize gene encoding a high-sulfur zein protein Mr 15,000. J Biol Chem 261: 6279–6284 (1986).

    PubMed  Google Scholar 

  32. Righetti PG, Gianazza E, Viotti A, Soave C: Heterogeneity of storage proteins in maize. Planta 136: 115–123 (1977).

    Google Scholar 

  33. Rivin CJ, Zimmer EA, Walbot V: Isolation of DNA and DNA recombinants from maize. In: Sheridan WF (ed) Maize for Biological Research, pp. 161–164. University Press, University of North Dakota, Grand Forks (1982).

    Google Scholar 

  34. Schmidt RJ, Burr FA, Aukerman MJ, Burr B: Maize regulatory gene opaque-2 encodes a protein with a ‘leucine-zipper’ motif that binds to zein DNA. Proc Natl Acad Sci USA 87: 46–50 (1990).

    PubMed  Google Scholar 

  35. Schmidt RJ, Ketudat M, Aukerman MJ, Hoscheck G: Opaque-2 is a transcriptional activator that recognizes a specific target site in 22-kD zein gene. Plant Cell 4: 689–700 (1992).

    Article  PubMed  Google Scholar 

  36. Soave C, Reggiani R, DiFonzo N, Salamini F: Clustering of genes for 20 K zein subunits in the short arm of maize chromosome 7. Genetics 97: 363–377 (1981).

    Google Scholar 

  37. Soave C, Reggiani R, DiFonzo N, Salamini F: Genes for zein subunits on maize chromosome 4. Biochem Genet 11: 1027–1037 (1982).

    Google Scholar 

  38. Shewry PR, Tatham AS: The prolamin storage proteins of cereal seeds: structure and evolution. Biochem J 267: 1–12 (1990).

    PubMed  Google Scholar 

  39. Targon MLPN, Ottoboni LMM, Leite A, Ludevid D, Puigdomenech P, Arruda P: Synthesis and deposition of coixin in seeds of Coix lacryma-jobi. Plant Sci 83: 169–180.

  40. Viotti A, Sala E, Marotta R, Alberti P, Paducci C, Soave C: Genes and mRNAs coding for zein polypeptides in Zea mays L. Eur J Biochem 102: 211–222 (1979).

    PubMed  Google Scholar 

  41. Wandelt C, Feix G: Sequence of a 21 kDa zein gene from maize containing an in frame stop codon. Nucl Acids Res 17: 2354 (1989).

    PubMed  Google Scholar 

  42. Wilson CM: Seed protein fractions of maize, sorghum and related cereals. In: Gottschal KW, Muller HP (eds) Seed Proteins: Biochemical, Genetics, Nutritive Value, pp. 271–307. Martinus Nijhoff/Junk, The Hague, Netherlands (1983).

    Google Scholar 

  43. Wilson CM: Serial analysis of zein by isoelectric focusing and sodium dodecil sulfate gel electrophoresis. Plant Physiol 82: 196–202 (1986).

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Centro de Biologia Molecular e Engenharia Genética, Universidade Estadual de Campinas, 13081, Campinas, SP, Brazil

    Laura M. M. Ottoboni, Adilson Leite, Jose A. Yunes, Targon Maria Luiza P. N., Gonçalo A. de Souza Filho & Paulo Arruda

  2. Departamento de Genética e Evoluçao, IB, Universidade Estadual de Campinas, 13081, Campinas, SP, Brazil

    Paulo Arruda

Authors
  1. Laura M. M. Ottoboni
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Adilson Leite
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jose A. Yunes
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Targon Maria Luiza P. N.
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Gonçalo A. de Souza Filho
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Paulo Arruda
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ottoboni, L.M.M., Leite, A., Yunes, J.A. et al. Sequence analysis of 22 kDa-like α-coixin genes and their comparison with homologous zein and kafirin genes reveals highly conserved protein structure and regulatory elements. Plant Mol Biol 21, 765–778 (1993). https://doi.org/10.1007/BF00027110

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00027110

Key words

Search

Navigation