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Theoretical and Applied Genetics

, Volume 82, Issue 5, pp 615–620 | Cite as

Characterization of HSP-70 cognate proteins from wheat

  • S. Giorini
  • G. Galili
Originals

Summary

Animal and plant cells contain a family of constitutively expressed HSP-70 cognate proteins that are localized in different subcellular locations and are presumed to play a role in protein folding and transport. Utilizing antibodies raised against the yeast endoplasmicreticulum-localized HSP-70 cognate termed BiP/GRP-78, as well as antibodies raised against the Escherichia coli HSP-70 protein DnaK, we have identified and characterized a large family of closely related proteins in wheat. One protein band of 78 kDa that is apparently closely related to yeast BiP was localized in the endoplasmic reticulum. This band cross-reacted with the yeast BiP but not with the DnaK-specific antibodies. The yeast BiP antibodies also recognized a cytoplasmic protein of 70 kDa that is probably related to the HSC-70 cognate proteins. These two proteins were further confirmed as HSP-70 cognates by their ability to bind to an ATP-agarose column. Probing of proteins from purified wheat mitochondrial preparations with the yeast BiP and DnaK-specific antibodies showed that this organelle contained a family of HSP-70-related proteins. The yeast BiP antibodies recognized two mitochondrial proteins of 60 and 58 kDa, but failed to detect any protein in the size rang of 70 to 80 kDa. However, the presence of immunologically distinct proteins of 90 and 78 kDa, as well as of lower molecular weight from this family in the mitochondria, was shown by probing with the DnaK-specific antibodies. A new protein of 30 kDa, cross-reacting with anti-yeast BiP antibodies, was detected only in developing seeds, close to their maturity. The evolution of HSP-70 cognate proteins in wheat as shown in this study is discussed.

Key words

HSP-70 BiP/GRP-78 Wheat Triticum aestivum Chaperone 

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References

  1. Amir-Shapira D, Leustek T, Weissbach H, Brot N (1990) HSP-70 proteins, similar to Escherichia coli DnaK, in chloroplasts and mitochondria of Euglena gracilis. Proc Natl Acad Sci; USA 87:1749–1752Google Scholar
  2. Bardwell JC, Craig EA (1984) Major heat shock gene of Drosophila and the Escherichia coli heat inducible DnaK gene are homologous. Proc Natl Acad Sci USA 81:848–852Google Scholar
  3. Bole DG, Hendershot LM, Kearney JF (1986) Posttranslational association of immunoglobulin heavy chain binding protein with nascent chains in nonsecreting and secreting hybridomas. J Cell Biol 102:1558–1566Google Scholar
  4. Bulleid NJ, Fredman RB (1988) Defective co-translational formation of disulfide bonds in protein disulfide-isomerase-deficient microsomes. Nature 335:649–651Google Scholar
  5. Burnette WN (1981) “Western blotting”: electrophoretic transfer of proteins from sodium dodecyl sulfate-polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem 112:195–203Google Scholar
  6. Craig EA, Kramer J, Shilling J, Werner-Washburne M, Holmes S, Kosic-Smithers J, Nicolet CM (1989) SSC1, an essential member of the yeast HSP-70 multigene family, encodes a mitochondrial protein. Mol Cell Biol 9:3000–3008Google Scholar
  7. Dorner AJ, Bole DJ, Kaufman RJ (1987) Reduction of endogenous GRP78 levels improves secretion of a heterologous protein in CHO cells. Mol Cell Biol 8:4063–4070Google Scholar
  8. Flynn G, Chappell TG, Rothman JE (1989) Peptide binding and release by proteins implicated as catalysts of protein assembly. Science 245:385–390Google Scholar
  9. Gething MJ, McCammon K, Sambrook J (1986) Expression of wild type and mutant forms of influenza hemagglutinin: the role of folding in intracellular transport. Cell 46:939–950Google Scholar
  10. Haas IG, Wabl M (1983) Immunoglobulin heavy chain binding protein. Nature 306:387–389Google Scholar
  11. Jindal S, Dudani AK, Singh B, Harley CB, Gupta RS (1989) Primary structure of human mitochondrial protein homologous to the bacterial and plant chaperonins and to the 65-Kda mycobacterial antigen. Mol Cell Biol 9:2279–2283Google Scholar
  12. Kassenbrock CK, Garcia PD, Walter P, Kelly RB (1988) Heavychain binding protein recognizes aberrant polypeptides translocated in vitro. Nature 333:90–93Google Scholar
  13. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 2227:680–685Google Scholar
  14. Leustek T, Dalie B, Amir-Shapira D, Brot N, Weissbach H (1989) A member of the HSP-70 family is localized in mitochondria and resembles Escherichia coli DnaK. Proc Natl Acad Sci USA 86:7805–7808Google Scholar
  15. Marshall JS, DeRocher AE, Keegstra K, Vierling E (1990) Identification of heat shock protein HSP-70 homologous in chloroplasts. Proc Natl Acad Sci USA 87:374–378Google Scholar
  16. Munro S, Pelham HRB (1986) An HSP-70-like protein in the ER: identity with the 78 kDA glucoe regulated protein and immunoglobulin heavy chain binding protein. Cell 46:291–300Google Scholar
  17. Nigam SK, Blobel G (1989) Cyclic AMP-dependent protein kinase in canine pancreatic rough endoplasmic reticulum. J Biol Chem 264:16927–16932Google Scholar
  18. O'Farrell PH (1975) High resolution two-dimensional electrophoresis of proteins. J Biol Chem 250:4007–4021Google Scholar
  19. Pelham HRB (1986) Speculations on the functions of the major heat shock and glucose-regulated proteins. Cell 46:959–961Google Scholar
  20. Rothman JE (1989) Polypeptide chain binding proteins: catalysts of protein folding and related processes in cells. Cell 59:591–601Google Scholar
  21. Rothman JE, Kornberg R (1986) An unfolding story of protein translocation. Nature 332:209–210Google Scholar
  22. Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets, procedures and some applications. Proc Natl Acad Sci USA 76:4350–4354Google Scholar
  23. Vogel JP, Misra LM, Rose MD (1990) Loss of BiP/GRP78 function blocks translocation of secretory proteins in yeast. J Cell Biol 110:1885–1895Google Scholar
  24. Welch JW, Feramisco JR (1985) Rapid purification of mammalian 70,000-Dalton proteins: affinity of the proteins for nucleotides. Mol Cell Biol 5:1229–1237Google Scholar

Copyright information

© Springer-Verlag 1991

Authors and Affiliations

  • S. Giorini
    • 1
  • G. Galili
    • 1
  1. 1.Department of Plant GeneticsThe Weizmann Institute of ScienceRehovotIsrael

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