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The wheat LEA protein Em functions as an osmoprotective molecule in Saccharomyces cerevisiae

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Abstract

The biased amino acid composition and aperiodic (random coil) configuration of Group 1 late embryogenesis-abundant (LEA) proteins imply that these proteins are capable of binding large amounts of water. While Group 1 LEAs have been predicted to contribute to osmotic stress protection in both embryonic and vegetative tissues, biochemical support has been lacking. We have used Saccharomyces cerevisiae as a model system to test the putative osmoprotective function of a wheat Group 1 LEA protein, Em. We demonstrate that expression of Em protein in yeast cells is not deleterious to growth in media of normal osmolarity and attenuates the growth inhibition normally observed in media of high osmolarity. Enhanced growth is observed in the presence of a variety of osmotically active compounds indicating that Em protein is capable of mitigating the detrimental effect of low water potential in a relatively non-specific manner. These results are the first biochemical demonstration of an osmoprotective function for a Group 1 LEA and suggest that the yeast expression system will be useful in dissecting the mechanism of protection through structure-function studies.

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References

  1. Albertyn J, Hohmann S, Thevelein JM, Prior BA: GPD1, which encodes glycerol-3-phosphate dehydrogenase, is essential for growth under osmotic stress in Saccharomyces cerevisiae, and its expression is regulated by the high-osmolarity glycerol response pathway. Mol Cell Biol 14: 4135–4144 (1994).

    Google Scholar 

  2. Almoguera C, Jordano J: Developmental and environmental concurrent expression of sunflower dry-seed stored molecular weight heat shock protein and Lea mRNA. Plant Mol Biol 19: 781–792 (1992).

    Google Scholar 

  3. Baker JC, Steele C, Dure III L: Sequence and characterization of 6 Lea proteins and their genes in cotton. Plant Mol Biol 11: 277–291 (1988).

    Google Scholar 

  4. Baler R, Welch WJ, Voellmy R: Heat shock gene regulation by nascent polypeptides and denatured proteins: hsp70 as a potential autoregulatory factor. J Cell Biol 117: 1151–1159 (1992).

    Google Scholar 

  5. Bartels D, Singh M, Salamini F: Onset of desiccation tolerance during development of the barley embryo. Planta 175: 485–492 (1988).

    Google Scholar 

  6. Becker DM, Guarente L: High-efficiency transformation of yeast by electroporation. Meth Enzymol 194: 182–187 (1991).

    Google Scholar 

  7. Berge SK, Bartholomew DM, Quatrano RS: Control of the expression of wheat embryo genes by abscisic acid. In: Goldberg RL (ed) Molecular Basis of Plant Development, pp. 193–201. Alan R. Liss, New York (1989).

    Google Scholar 

  8. Blomberg A, Adler L: Physiology of osmotolerance in yeast. Adv Microbiol Physiol 33: 145–212 (1992).

    Google Scholar 

  9. Bostock RM, Quatrano RS: Regulation of Em gene expression in rice. Plant Physiol 98: 1356–1363 (1992).

    Google Scholar 

  10. Bradford M: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254 (1976).

    Google Scholar 

  11. Bray EA: Molecular responses to water deficit. Plant Physiol 103: 1035–1040 (1993).

    Google Scholar 

  12. Cherry JM, Adler C, Ball C, Dwight S, Chervitz S, Jia Y, Juvik G, Weng S, Botstein D: Saccharomyces Genome Database, http://genome-www.stanford.edu/Saccharomyces/, Non-Redundant S. cerevisiae Protein Data Set, May 13, 1997 (1997).

  13. Cherry JM: Codon tables from http://128.252.119.253/ docs/bio/cod.html (1994).

  14. Close TJ, Kortt AA, Chandler PM: A cDNA-based comparison of dehydration-induced proteins (dehydrins) in barley and corn. Plant Mol Biol 13: 95–108 (1989).

    Google Scholar 

  15. Compagno C, Coraggio I, Ranzi BM, Alberghian L, Viotti A, Martegani E: Translational regulation of the expression of zein cloned in yeast under an inducible GAL promoter. Biochem Biophys Res Comm 146: 809–814 (1987).

    Google Scholar 

  16. Dure III L, Greenway S, Galau GA: Developmental biochemistry of cottonseed embryogenesis and germination XIV. Changing mRNA populations as shown by in vitro and in vivo protein synthesis. Biochemistry 20: 4162–4168 (1981).

    Google Scholar 

  17. Dure III L, Crouch M, Harada J, Ho T-HD, Mundy J, Quatrano RS, Thomas T, Sung ZR: Common amino acid sequence domains among the LEA proteins of higher plants. Plant Mol Biol 12: 475–486 (1989).

    Google Scholar 

  18. Dure III L: Structural motifs in LEA proteins. In: Close TJ (ed) Plant Responses to Cellular Dehydration during Environmental Stress, pp. 91–103. American Society of Plant Physiologists, Rockville, MN (1993).

    Google Scholar 

  19. Espelund M, Saebøe-Larssen S, Hughes DW, Galau GA, Larsen F, Jakobsen KS: Late embryogenesis-abundant genes encoding proteins with different numbers of hydrophilic repeats are regulated differentially by abscisic acid and osmotic stress. Plant J 2: 241–252 (1992).

    Google Scholar 

  20. Galau GA, Hughes DW, Dure III L: Abscisic acid induction of cloned cotton late embryogenesis-abundant (lea) mRNAs. Plant Mol Biol 7: 155–170 (1986).

    Google Scholar 

  21. Galau GA, Wang HYC, Hughes DW: Cotton Lea4 (D19) and LeaA2 (D132) group 1 Lea genes encoding water stressrelated proteins containing a 20-amino acid motif. Plant Physiol 99: 783–788 (1992).

    Google Scholar 

  22. Gaubier P, Raynal M, Hull G, Huestis GM, Grellet F, Arenas C, Pages M, Delsney M: Two different Em-like genes are expressed in Arabidopsis thaliana seed during maturation. Mol Gen Genet 238: 409–418 (1993).

    Google Scholar 

  23. Gaxiola R, Larrinoa IFD, Villalba MJ, Serrano R: A novel and conserved salt-induced protein is an important determinant of salt tolerance in yeast. EMBO J 11: 3157–3164 (1992).

    Google Scholar 

  24. Goff SA, Goldberg AL: Production of abnormal proteins in E. coli stimulates transcription of lon and other heat shock genes. Cell 41: 587–595 (1985).

    Google Scholar 

  25. Gomez J, Sanchez-Martinez D, Stiefel V, Rigau J, Puigdomenech P, Pages M: A gene induced by the plant hormone abscisic acid in response to water stress encodes a glycine-rich protein. Nature 334: 262–264 (1988).

    Google Scholar 

  26. Grzelczak ZF, Sattolo MH, Hanley-Bowdoin LK, Kennedy TD, Lane BG: Synthesis and turnover of proteins and mRNA in germinating wheat embryos. Can J Biochem 60: 389–397 (1982).

    Google Scholar 

  27. Hanahan D: Studies on transformation of Escherichia coli with plasmids. J Mol Biol 166: 557–580 (1983).

    Google Scholar 

  28. Henson IE: Effects of atmospheric humidity on abscisic acid accumulation and water status in leaves of rice. Ann Bot 54: 569–582 (1984).

    Google Scholar 

  29. Hoffman CS, Winston F: A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformation of Escherichia coli. Gene 57: 267–272 (1987).

    Google Scholar 

  30. Hofmann T, Kells DIC, Lane BG: Partial amino acid sequence of the wheat germ Ec protein: Comparison with another protein very rich in half-cystein and glycine, wheat germ agglutinin. Can J Biochem Cell Biol 62: 908–913 (1984).

    Google Scholar 

  31. Hogue DL, Ellison MJ, Young JD, Cass CE: Identification of a novel membrane transporter associated with intracellular membranes by phenotypic complementation in the yeast Saccharomyces cerevisiae. J Biol Chem 271: 9801–9808 (1996).

    Google Scholar 

  32. Imai R, Chang L, Ohta A, Bray E, Takagi M: A lea-class gene of tomato confers salt and freezing tolerance when expressed in Saccharomyces cerevisiae. Gene 170: 243–248 (1996).

    Google Scholar 

  33. Jessee J, Bloom F: DH5αMCRTM: New competent cells for cloning methylated DNA. Focus 10: 69–70 (1988).

    Google Scholar 

  34. Johnston M, Davis RW: Sequences that regulate the divergent GAL1–GAL10 promoter in Saccharomyces cerevisiae. Mol Cell Biol 4: 1440–1448 (1984).

    Google Scholar 

  35. Jones H, Leigh RA, Tomos AD, Jones RGW: The effects of abscisic acid on cell turgor pressures, soluble content and growth of wheat roots. Planta 170: 257–262 (1987).

    Google Scholar 

  36. Kelley PM, Schlesinger MJ: The effect of amino acid analogues and heat shock on gene expression in chicken embryo fibroblasts. Cell 15: 1277–1286 (1978).

    Google Scholar 

  37. Kyte J, Doolittle RF: A simple method for displaying the hydropathic character of a protein. J Mol Biol 157: 105–132 (1982).

    Google Scholar 

  38. Lennox ES: Transduction of linked genetic characters of the host by bacteriophage P1. Virology 1: 190–206 (1955).

    Google Scholar 

  39. Litts JC, Colwell GW, Chakerian RL, Quatrano RS: The nucleotide sequence of a cDNA clone encoding the wheat Em protein. Nucl Acids Res 15: 3607–3618 (1987).

    Google Scholar 

  40. Litts JC, Colwell GW, Chakerian RL, Quatrano RS: Sequence analysis of a functional member of the Em gene family from wheat. DNA Sequencing 1: 263–274 (1991).

    Google Scholar 

  41. Litts JC, Erdman MB, Huang N, Karrer EE, Nouiery A, Quatrano RS, Rodriguez RL: Nucleotide sequence of the rice (Oryza sativa) Em protein gene (EMP1). Plant Mol Biol 19: 335–337 (1992).

    Google Scholar 

  42. Manickam A, Van Damme EJM, Kalaiselvi K, Verhaert P, Pumans WJ: Isolation and cDNA cloning of an Em-like protein from mung bean (Vigna radiata) axes. Physiol Plant 97: 524–530 (1996).

    Google Scholar 

  43. Marcotte Jr WR, Bayley CC, Quatrano RS: Regulation of a wheat promoter by abscisic acid in rice protoplasts. Nature 335: 454–457 (1988).

    Google Scholar 

  44. McCubbin WD, Kay CM, Lane BG: Hydrodynamic and optical properties of the wheat germ Em protein. Can J Biochem Cell Biol 63: 803–811 (1985).

    Google Scholar 

  45. Morris PC, Kumar A, Bowles DJ, Cuming AC: Osmotic stress and abscisic acid induce expression of the wheat Em genes. Eur J Biochem 190: 625–630 (1990).

    Google Scholar 

  46. Mundy J, Chua N-H: Abscisic acid and water-stress induce the expression of a novel rice gene. EMBO J 7: 2279–2286 (1988).

    Google Scholar 

  47. Piper PW, Talreja K, Panaretou B, Moradas-Ferreira P, Byrne K, Praekelt UM, Meacock PA, Récnacq M, Boucherie H: Induction of major heat-shock proteins of Saccharomyces cerevisiae, including plasma membrane Hsp30, by ethanol levels above a critical threshold. Microbiology 140: 3031–3038 (1994).

    Google Scholar 

  48. Praekelt UM, Meacock PA: HSP12, a new small heat shock gene of Saccharomyces cerevisiae: analysis of structure, regulation and function. Mol Gen Genet 223: 97–106 (1990).

    Google Scholar 

  49. Pringle JR: Methods for avoiding proteolytic artefacts in studies of enzymes and other proteins from yeasts. Meth Cell Biol 12: 149–184 (1975).

    Google Scholar 

  50. Raynal M, Depigny D, Cooke R, Delsney M: Characterization of a radish nuclear gene expressed during late seed maturation. Plant Physiol 91: 829–836 (1989).

    Google Scholar 

  51. Roberts JK, Noelle AD, Wilma LL, Dure III L: Cellular concentrations and uniformity of cell-type accumulation of two lea proteins in cotton embryos. Plant Cell 5: 769–780 (1993).

    Google Scholar 

  52. Sambrook J, Fritsch EF, Maniatis T: Molecular Cloning: A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY (1989).

    Google Scholar 

  53. Schmitt ME, Brown TA, Trumpower BL: A rapid and simple method for preparation of RNA from Saccharomyces cerevisiae. Nucl Acids Res 18: 3091–3092 (1990).

    Google Scholar 

  54. Schneider JC, Guarente L: Vectors for expression of cloned genes in yeast: regulation, overproduction, and underproduction. Meth Enzymol 194: 373–388 (1991).

    Google Scholar 

  55. Selden RF: Analysis of RNA by norhern hybridization. In: Ausubel FA et al. (eds) Current Protocols in Molecular Biology, pp. 4.9.4.–4.9.8. Greene Publishing/Wiley-Interscience, New York (1987).

    Google Scholar 

  56. Serrano R, Márquez JA, Ríos G: Crucial factors in salt stress tolerance. In: Hohmann S, Mager W (eds) Yeast Stress Responses, pp. 147–169. RG Landes, Austin, TX (1997).

    Google Scholar 

  57. Sherman F: Getting started with yeast. Meth Enzymol 194: 3–21 (1991).

    Google Scholar 

  58. Steponkus PL, Webb MS: Freeze-induced dehydration and membrane destabilization in plants. In: Somero GN, Osmond CB, Bolis CL (eds) Water and Life: Comparative Analysis of Water Relationships at the Organismic, Cellular and Molecular Levels, pp. 338–362. Springer-Verlag, Berlin (1992).

    Google Scholar 

  59. Stone RL, Matarese V, Magee B, Magee PT, Bernlohr DA: Cloning, sequencing and chromosomal assignment of a gene from Saccharomyces cerevisiae which is regulated negatively by glucose and positively by lipids. Gene 96: 171–176 (1990).

    Google Scholar 

  60. Tartof KD, Hobbs CA: Improved media for growing plasmid and cosmid clones. Focus 9: 12 (1987).

    Google Scholar 

  61. Ulrich TU, Wurtele ES, Nikolau BL: Sequence of EMB-1, an mRNA accumulating specifically in embryos of carrot. Nucl Acids Res 18: 2826 (1990).

    Google Scholar 

  62. Varela JCS, van Beekvelt CA, Planta RJ, Mager WH: Osmostress-induced changes in yeast gene expression. Mol Microbiol 6: 2183–2190 (1992).

    Google Scholar 

  63. Varela JCS, Mager WH: Response of Saccharomyces cerevisiae to changes in external osmoticum. Microbiology 142: 721–731 (1996).

    Google Scholar 

  64. Villardell J, Goday A, Freire MA, Torrent M, Martinez C, Torne H, Pages M: Gene sequence, developmental expression and protein phosphorylation of RAB17 in maize. Plant Mol Biol 14: 423–432 (1990).

    Google Scholar 

  65. Williams B, Tsang A: A maize gene expressed during embryogenesis is abscisic acid-inducible and highly conserved. Plant Mol Biol 16: 916–923 (1991).

    Google Scholar 

  66. Xu D, Duan X, Wang B, Hong B, Ho T-HD, Wu R: Expression of a late embryogenesis abundant protein gene, HVA1, from barley confers tolerance to water deficit and salt stress in transgenic rice. Plant Physiol 110: 249–257 (1996).

    Google Scholar 

  67. Zaret KS, Sherman F: DNA sequence required for efficient transcription termination in yeast. Cell 28: 563–573 (1982).

    Google Scholar 

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  1. Clemson University, 132 Long Hall, Clemson, SC, 29634-1903, USA

    Ginger A. Swire-Clark & William R. Marcotte Jr.

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Swire-Clark, G.A., Marcotte, W.R. The wheat LEA protein Em functions as an osmoprotective molecule in Saccharomyces cerevisiae. Plant Mol Biol 39, 117–128 (1999). https://doi.org/10.1023/A:1006106906345

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