Plant Molecular Biology

, Volume 66, Issue 3, pp 221–231 | Cite as

Molecular genetics of puroindolines and related genes: regulation of expression, membrane binding properties and applications



Kernel texture of wheat is a primary determinant of its technological properties. Soft kernel texture phenotype results when the Puroindoline a and Puroindoline b genes are present and encode the wild-type puroindolines PINA and PINB, respectively, and various mutations in either or both gene(s) result in hard phenotypes. A wealth of information is now available that furthers our understanding regarding the spatial and temporal regulation of expression of Puroindoline genes. Through the use of model membranes and synthetic peptides we also have a clearer understanding of the significance of the cysteine backbone, the tryptophan-rich domain (TRD) and the helicoid tertiary structures of PIN proteins in relation to their membrane-active properties. Many studies suggest individual yet co-operative modes of action of the PIN proteins in determining kernel texture, and significant evidence is accumulating that the proteins have in vivo and in vitro antimicrobial activities, shedding light on the biological roles of this unique ensemble of proteins. The puroindolines are now being explored for grain kernel texture modifications as well as antimicrobial activities.


Grain hardness Puroindolines Grain softness protein Kernel texture Wheat × Triticum Gene expression Antimicrobial properties 



Days after flowering (≈days post-anthesis)


Grain softness protein


Near-isogenic lines


Non-specific lipid-transfer proteins


Quantitative trait locus/loci


Single nucleotide polymorphism


Tryptophan-rich domain


  1. Amiour N, Merlino M, Leroy P et al (2003) Chromosome mapping and identification of amphiphilic proteins of hexaploid wheat kernels. Theor Appl Genet 108:62–72PubMedCrossRefGoogle Scholar
  2. Amoroso MG, Longobardo L, Capparelli R (2004) Real time PCR and flow cytometry to investigate wheat kernel hardness: role of puroindoline genes and proteins. Biotechnol Lett 26:1731–1737PubMedCrossRefGoogle Scholar
  3. Baker RJ, Sutherland KA (1991) Inheritance of kernel hardness in five spring wheat crosses. Can J Plant Sci 71:179–181Google Scholar
  4. Beecher B, Bettge A, Smidansky E et al (2002) Expression of wild-type pinB sequence in transgenic wheat complements a hard phenotype. Theor Appl Genet 105:870–877PubMedCrossRefGoogle Scholar
  5. Bhave M, Morris CF (2007) Molecular genetics of puroindolines and related genes: allelic diversity in wheat and other grasses. Plant Mol Biol (in press)Google Scholar
  6. Biswas S, Marion D (2006) Interaction between puroindolines and the major polar lipids of wheat seed endosperm at the air–water interface. Colloids Surf B 53:167–174CrossRefGoogle Scholar
  7. Blein J, Coutos-Thevenot P, Marion D et al (2002) From elicitins to lipid-transfer proteins: a new insight I cell signaling involved in plant defence mechanisms. Trends Plant Sci 7:293–296PubMedCrossRefGoogle Scholar
  8. Bonafede M, Kong L, Tranquilli G et al (2007) Reduction of a Triticum monococcum chromosome segment carrying the softness genes Pina and Pinb translocated to bread wheat. Crop Sci 47:819–826CrossRefGoogle Scholar
  9. Bottley A, Xia GM, Koebner RMD (2006) Homeologous gene silencing in hexapolid wheat. Plant J 47:897–906PubMedCrossRefGoogle Scholar
  10. Boutrot F, Guirao A, Alary R et al (2005) Wheat non-specific lipid transfer protein genes display a complex patterns of expression in developing seeds. Biochim Biophys Acta 1730:114–125PubMedGoogle Scholar
  11. Breseghello F, Finney PL, Gaines C et al (2005) Genetic loci related to kernel quality differences between a hard and soft wheat cultivar. Crop Sci 45:1685–1695CrossRefGoogle Scholar
  12. Cameron K, Teece M, Smart L (2006) Increased accumulation of cuticular wax and expression of lipid transfer protein in response to periodic drying in leaves of tree tobacco. Plant Physiol 140:176–183PubMedCrossRefGoogle Scholar
  13. Campbell KG, Bergman CJ, Gualberto DG et al (1999) Quantitative trait loci associated with kernel traits in a soft × hard wheat cross. Crop Sci 39:1184–1195CrossRefGoogle Scholar
  14. Campbell JB, Martin JM, Crutcher F et al (2007) Effects on soft wheat (Triticum aestivum L.) quality of increased Puroindoline dosage. Cereal Chem 84:80–87CrossRefGoogle Scholar
  15. Capparelli R, Borriello G, Giroux MJ et al (2003) Puroindoline-a gene expression is involved in association of puroindolines to starch. Theor Appl Genet 107:1463–1468PubMedCrossRefGoogle Scholar
  16. Capparelli R, Amoroso MG, Palumbo D et al (2005) Two plant puroindolines colocalise in wheat seed and in vitro synergistically fight against pathogens. Plant Mol Biol 58:857–867PubMedCrossRefGoogle Scholar
  17. Capparelli R, Palumbo D, Iannaccone M et al (2006) Cloning and expression of two plant proteins: similar antimicrobial activity of native and recombinant form. Biotechnol Lett 28:943–949PubMedCrossRefGoogle Scholar
  18. Capparelli R, Ventimiglia I, Palumbo D et al (2007) Expression of recombinant puroindolines for the treatment of staphylococcal skin infections (Acne vulgaris). J Biotechnol 128:606–614PubMedCrossRefGoogle Scholar
  19. Charnet P, Molle G, Marion D et al (2003) Puroindolines form ion channels in biological membranes. Biophys J 84:2416–2426PubMedCrossRefGoogle Scholar
  20. Chen M, Wilkinson M, Tosi P et al (2005) Novel puroindoline and grain softness protein alleles in Aegilops species with the C, D, S, M and U genomes. Theor Appl Genet 111:1159–1166PubMedCrossRefGoogle Scholar
  21. Chen F, He Z, Xia X et al (2006) Molecular and biochemical characterisation of puroindoline a and b alleles in Chinese landraces and historical cultivars. Theor Appl Genet 112:400–409PubMedCrossRefGoogle Scholar
  22. Clarke B, Rahman S (2005) A microarray analysis of wheat grain hardness. Theor Appl Genet 110:1259–1267PubMedCrossRefGoogle Scholar
  23. Clifton LA, Lad MD, Green RJ et al (2007) Single amino acid substitutions in puroindoline-b mutants influence lipid binding properties. Biochem 46:2260–2266CrossRefGoogle Scholar
  24. Corona V, Gazza L, Boggini G et al (2001) Variation in friabilin composition as determined by A-PAGE fractionation and PCR amplification, and its relationship to grain hardness in bread wheat. J Cereal Sci 34:243–250CrossRefGoogle Scholar
  25. Day L, Bhandari D, Greenwell P et al (2006) Characterisation of wheat puroindoline proteins. FEBS J 273:5358–5373PubMedCrossRefGoogle Scholar
  26. De Planque MRR, Kruijtzer J, Liskamp R et al (1999) Different membrane anchoring positions of tryptophan and lysine in synthetic transmembrane α-helical peptides. J Biol Chem 274:20839–20846PubMedCrossRefGoogle Scholar
  27. Dessalegn T, Labuschagne MT, van Deventer CS (2006) Quality of Ethiopean durum wheat lines in two diverse environments. J Agron Crop Sci 192:147–150CrossRefGoogle Scholar
  28. Digeon JF, Guiderdoni E, Alary R et al (1999) Cloning of a wheat puroindoline gene promoter by IPCR and analysis of promoter sequences required for tissue-specific expression in transgenic rice seeds. Plant Mol Biol 39:1101–1112PubMedCrossRefGoogle Scholar
  29. Douliez JP, Michon T, Elmorajani K et al (2000) Structure, biological and technological functions of lipid transfer proteins and indolines, the major lipid binding proteins from cereal kernels. J Cereal Sci 32:1–20CrossRefGoogle Scholar
  30. Drea S, Leader DJ, Arnold B et al (2005) Systematic spatial analysis of gene expression during wheat caryopsis development. Plant Cell 17:2172–2185PubMedCrossRefGoogle Scholar
  31. Dubreil L, Compoint JP, Marion D (1997) Interaction of puroindolines with wheat flour polar lipids determines their foaming properties. J Agric Food Chem 45:108–116CrossRefGoogle Scholar
  32. Dubreil L, Gabroit T, Bouchet B et al (1998) Spatial and temporal distribution of the major isoforms of puroindolines (puroindoline-a and puroindoline-b) and non-specific lipid transfer protein (nsLTPe1) of Triticum aestivum seeds. Relationships with their in vitro antifungal properties. Plant Sci 138:121–135CrossRefGoogle Scholar
  33. Elmorjani K, Lurquin V, Lelion A et al (2004) A bacterial expression system revisited for the recombinant production of cysteine-rich plant lipid transfer proteins. Bichem Biophys Res Commun 316:1202–1209CrossRefGoogle Scholar
  34. Faize M, Sourice S, Dupuis F et al (2004) Expression of wheat puroindoline-b reduces scab susceptibility in transgenic apple (Malus X domestica Borkh.). Plant Sci 167:347–354CrossRefGoogle Scholar
  35. Garcia-Olmedo F, Molina A, Alamillo J et al (1998) Plant defence peptides. Biopolymers (Peptide Science) 47:479–491CrossRefGoogle Scholar
  36. Gautier MF, Aleman ME, Guirao A et al (1994) Triticum aestivum puroindolines, two basic cysteine-rich seed proteins: cDNA sequence analysis and developmental gene expression. Plant Mol Biol 25:43–57PubMedCrossRefGoogle Scholar
  37. Gautier MF, Cosson P, Guirao A et al (2000) Puroindoline genes are highly conserved in diploid ancestor wheats and related species but absent in tetraploid Triticum species. Plant Sci 153:81–91CrossRefGoogle Scholar
  38. Gazza L, Niglio A, Vaccino P et al (2003) The long arm of chromosome 5D of bread wheat contains a Pina-D1a-like sequence. In: Pogna NE, Romano M, Pogna E, Galterio G (eds) Proc 10th intl wheat genet symp, vol. 3. Paestum, Italy, pp 1330–1332Google Scholar
  39. Gazza L, Nocente F, Ng PKW et al (2005) Genetic and biochemical analysis of common wheat cultivars lacking puroindoline a. Theor Appl Genet 110:470–478PubMedCrossRefGoogle Scholar
  40. Gazza L, Taddei F, Corbellini M et al (2007) Genetic and environmental factors affecting grain texture in common wheat. J Cereal Sci. doi:10.1016/j.jcs.2007.01.004 Google Scholar
  41. Gedye KR, Morris CF, Bettge AD (2004) Determination and evaluation of the sequence and textural effects of the puroindoline a and puroindoline b genes in a population of synthetic hexaploid wheat. Theor Appl Genet 109:1597–1603PubMedCrossRefGoogle Scholar
  42. Giroux MJ, Morris CF (1997) A glycine to serine change in puroindoline-b is associated with wheat grain hardness and low levels of starch-surface friabilin. Theor Appl Genet 95:857–864CrossRefGoogle Scholar
  43. Giroux MJ, Morris CF (1998) Wheat grain hardness results from highly conserved mutations in the friabilin components puroindoline-a and -b. Proc Natl Acad Sci USA 95:6262–6266PubMedCrossRefGoogle Scholar
  44. Giroux MJ, Talbert L, Habernicht DK et al (2000) Association of puroindoline sequence type and grain hardness in hard red spring wheat. Crop Sci 30:370–374CrossRefGoogle Scholar
  45. Hogg AC, Sripo T, Beecher B et al (2004) Wheat puroindolines interact to form friabilin and control wheat grain hardness. Theor Appl Genet 108:1089–1097PubMedCrossRefGoogle Scholar
  46. Hughes P, Dennis E, Whitecross M et al (2000) The cytotoxic plant protein, β-purothionin, forms ion channels in lipid membranes. J Biol Chem 275:823–827PubMedCrossRefGoogle Scholar
  47. Igrejas G, Leroy P, Charmet G et al (2002) Mapping QTLs for grain hardness and puroindoline content in wheat (Triticum aestivum L.). Theor Appl Genet 106:19–27PubMedGoogle Scholar
  48. Ikeda TM, Ohnishi N, Nagamine T et al (2005) Identification of new puroindoline genotypes and their relationship to flour texture among wheat cultivars. J Cereal Sci 41:1–6CrossRefGoogle Scholar
  49. Issaly N, Soisona O, Joudrier P et al (2001) Optimisation of the wheat puroindoline-a production in Pichia pastoris. J Appl Microbiol 90:397–406PubMedCrossRefGoogle Scholar
  50. Jing W, Demcoe A, Vogel HJ (2003) Conformation of a bactericidal domain of puroindoline a: structure and mechanism of action of a 13-residue antimicrobial peptide. J Bacteriol 185:4938–4947PubMedCrossRefGoogle Scholar
  51. Kader J-C (1996) Lipid-transfer proteins in plants. Annu Rev Plant Physiol Plant Mol Biol 47:627–654PubMedCrossRefGoogle Scholar
  52. Kan Y, Wan Y, Beaudoin F et al (2006) Transcriptome analysis reveals differentially expressed storage protein transcripts in seeds of Aegilops and wheat. J Cereal Sci 44:75–85CrossRefGoogle Scholar
  53. Konopka I, Rotkiewicz D, Tanska M (2005) Wheat endosperm hardness. Part II. Relationship to content and composition of flour lipids. Eur Food Res Technol 220:20–24CrossRefGoogle Scholar
  54. Kooijman M, Orsel R, Hessing M et al (1997) Spectroscopic characterisation of the lipid-binding properties of wheat puroindolines. J Cereal Sci 26:145–159CrossRefGoogle Scholar
  55. Kooijman M, Orsel R, Hamer RJ et al (1998) The insertion behaviour of wheat puroindoline-a into diacylgalactosylglycerol films. J Cereal Sci 28:43–51CrossRefGoogle Scholar
  56. Krishnamurthy K, Giroux M (2001) Expression of wheat puroindoline genes in transgenic rice confers grain softness. Nat Biotechnol 19:162–166PubMedCrossRefGoogle Scholar
  57. Krishnamurthy K, Balconi C, Sherwood JE et al (2001) Wheat puroindolines enhance fungal disease resistance in transgenic rice. Mol Plant Microbe Interact 14:1255–1260PubMedCrossRefGoogle Scholar
  58. Laudencia-Chingcuanco DL, Stamova BS et al (2007) Transcriptional profiling of wheat caryopsis development using cDNA microarrays. Plant Mol Biol 63:651–668PubMedCrossRefGoogle Scholar
  59. Le Bihan T, Blochet JE, Desormeaux A et al (1996) Determination of the secondary structure and conformation of puroindolines by infrared and Raman spectroscopy. Biochemistry 35:12712–12722PubMedCrossRefGoogle Scholar
  60. Le Guerneve C, Seigneuret M, Marion D (1998) Interaction of the wheat endosperm lipid-binding protein puroindoline-a with phospholipids. Arch Biochem Biophys 360:179–186PubMedCrossRefGoogle Scholar
  61. Lillemo M, Ringlund K (2002) Impact of puroindoline b alleles on the genetic variation for hardness in soft × hard wheat crosses. Plant Breed 121:210–217CrossRefGoogle Scholar
  62. Lillemo M, Simeone MC, Morris CF (2002) Analysis of puroindoline a and b sequences from Triticum aestivum cv. ‘Penawawa’ and related taxa. Euphytica 126:321–331CrossRefGoogle Scholar
  63. Lillemo M, Chen F, Xia X et al (2006) Puroindoline grain hardness alleles in CIMMYT bread wheat germplasm. J Cereal Sci 44:86–92CrossRefGoogle Scholar
  64. Llanos P, Henriquez M, Minic J et al (2006) Puroindoline-a and α1-purothionin form ion channels in giant liposomes but exert different toxic actions on murine cells. FEBS J 273:1710–1722PubMedCrossRefGoogle Scholar
  65. Luo L, Zhang J, Yang G et al (2007) Expression of puroindoline A enhances leaf rust resistance in transgenic tetraploid wheat. Mol Biol Rep. doi:10.1007/s11033-007-9070-x Google Scholar
  66. Marion D, Gautier MF, Joudrier P et al (1994) Structure and function of wheat lipid binding proteins. In: Wheat kernel proteins: molecular and functional aspects, Proc Int’l Mtg. Universita Degli Sudi della Tuscia, Viterbo, pp 175–180Google Scholar
  67. Martin JM, Meyer FD, Smidansky ED et al (2006) Complementation of the pina (null) allele with the wild type Pina sequence restores a soft phenotype in transgenic wheat. Theor Appl Genet 113:1563–1570PubMedCrossRefGoogle Scholar
  68. McIntosh S, Watson L, Bundock P et al (2007) SAGE of the developing wheat caryopsis. Plant Biotechnol J 5:69–83PubMedCrossRefGoogle Scholar
  69. Morris CF (2002) Puroindolines: the molecular genetic basis of wheat grain hardness. Plant Mol Biol 48:633–647PubMedCrossRefGoogle Scholar
  70. Morris CF, Bhave M (2007) Reconciliation of D-genome puroindoline allele designations with current DNA sequence data. J Cereal Sci (in press)Google Scholar
  71. Morris CF, Lillemo M, Simeone MC et al (2001) Prevalence of puroindoline grain hardness genotypes among historically significant North American spring and winter wheats. Crop Sci 41:218–228CrossRefGoogle Scholar
  72. Mourgues F, Brisset M-N, Chevreau E (1998) Activity of different antimicrobial peptides on Erwinia amylovora growth, and evaluation of the phytotoxicity and stability of cecropins. Plant Sci 139:83–91CrossRefGoogle Scholar
  73. Partridge MAK, Appels R, Skerritt JH (2002) Simple ELISA detection of a new polymorphic Ha locus encoded protein. J Cereal Sci 35:189–200CrossRefGoogle Scholar
  74. Perretant MR, Cadalen T, Charnet G et al (2000) QTL analysis of bread-making quality in wheat using a doubled haploid population. Theor Appl Genet 100:1167–1175CrossRefGoogle Scholar
  75. Perrocheau L, Bakan B, Boivin P et al (2006) Stability of barley and malt lipid transfer protein 1 (LTP1) toward heating and reducing agents: relationships with the brewing process. J Agric Food Chem 54:3108–3113PubMedCrossRefGoogle Scholar
  76. Rahman S, Jolly CJ, Skerritt JH et al (1994) Cloning of a wheat 15 kDa grain softness protein (GSP). GSP is a mixture of different puroindoline-like polypeptides. Eur J Biochem 223:917–925PubMedCrossRefGoogle Scholar
  77. Rezansoff AJ, Hunter HN, Jing W et al (2005) Interactions of the antimicrobial peptide Ac-FRWWHR-NH2 with model membrane systems and bacterial cells. J Peptide Res 65:491–501CrossRefGoogle Scholar
  78. Schibli DJ, Epand RF, Vogel HJ et al (2002) Tryptophan-rich antimicrobial peptides: comparative properties and membrane interactions. Biochem Cell Biol 80:667–677PubMedCrossRefGoogle Scholar
  79. See DR, Giroux M, Gill BS (2004) Effect of multiple copies of puroindoline genes on grain softness. Crop Sci 44:1248–1253CrossRefGoogle Scholar
  80. Shewry PR, Jenkins J, Beaudoin F et al (2004) The classification, functions and evolutionary relationships of plant proteins in relation to food allergens. In: Mills ENC, Shewry PR (eds) Plant food allergens. Blackwell Science, Oxford, pp 24–41Google Scholar
  81. Simeone MC, Lafiandra D (2005) Isolation and characterization of friabilin genes in rye. J Cereal Sci 41:115–122CrossRefGoogle Scholar
  82. Simeone MC, Gedye KR, Mason-Gammer R et al (2006) Conserved regulatory elements identified from a comparative puroindoline gene sequence survey of Triticum and Aegilops diploid taxa. J Cereal Sci 44:21–33CrossRefGoogle Scholar
  83. Swan CG, Meyer FD, Hogg AC et al (2006) Puroindoline B limits binding of puroindoline A to starch and grain softness. Crop Sci 46:1656–1665CrossRefGoogle Scholar
  84. Tassin S, Broekaert WF, Marion D et al (1998) Solution structure of Ace-AMP1, a potent antimicrobial protein extracted from onion seeds. Structural analogies with plant non-specific lipd transfer proteins. Biochemistry 27:3623–3637CrossRefGoogle Scholar
  85. Tranquilli G, Heaton J, Chicaiza O et al (2002) Substitutions and deletions of genes related to grain hardness and their effect on grain texture. Crop Sci 42:1812–1817CrossRefGoogle Scholar
  86. Turnbull KM, Rahman S (2002) Endosperm texture in wheat. J Cereal Sci 36:327–337CrossRefGoogle Scholar
  87. Turnbull KM, Gaborit T, Marion D et al (2000) Variation in puroindoline polypeptides in Australian wheat cultivars in relation to grain hardness. Aus J Plant Physiol 55:89–95Google Scholar
  88. Turnbull KM, Marion D, Gaborit T et al (2003a) Early expression of grain hardness in the developing wheat endosperm. Plants 216:699–706Google Scholar
  89. Turnbull KM, Turner M, Mukai Y et al (2003b) The organization of genes tightly linked to the Ha locus in Aegilops tauschii, the D-genome donor to wheat. Genome 46:330–338PubMedCrossRefGoogle Scholar
  90. Turner M, Mukai Y, Leroy P et al (1999) The Ha locus of wheat: identification of a polymorphic region for tracing grain hardness in crosses. Genome 42:1242–1250PubMedCrossRefGoogle Scholar
  91. Turner AS, Bradburne RP, Fish L et al (2004) New quantitative trait loci influencing grain texture and protein content in bread wheat. J Cereal Sci 40:51–60CrossRefGoogle Scholar
  92. Van den Bulck K, Loosveld A, Courtin CM et al (2002) Amino acid sequence of wheat flour arabinogalactan peptide, identical to part of grain softness protein GSP-1, leads to improved structural model. Cereal Chem 79:329–331CrossRefGoogle Scholar
  93. Vila-Perello M, Tognon S, Sanchez-Vallet A et al (2006) A minimalist design approach to antimicrobial agents based on a thionin template. J Med Chem 49:448–451PubMedCrossRefGoogle Scholar
  94. Wanguji HW, Hogg AC, Martin JM et al (2007) The role of puroindoline A and B individually and in combination on grain hardness and starch association. Crop Sci 47:67–76CrossRefGoogle Scholar
  95. Xia L, Geng H, Chen X et al (2007) Silencing of puroindoline a alters the kernel texture in transgenic bread wheat. J Cereal Sci. doi:10.1016/j.jcs.2007.04.016 Google Scholar
  96. Yau WM, Wimley WC, Gawrisch K et al (1998) The preference of tryptophan for membrane interfaces. Biochemistry 37:14713–14718PubMedCrossRefGoogle Scholar
  97. Zhang J (2003) Evolution by gene duplication: an update. Trends Ecol Evol 18:292–298CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2007

Authors and Affiliations

  1. 1.Environment and Biotechnology Centre, Faculty of Life and Social SciencesSwinburne University of TechnologyMelbourneAustralia
  2. 2.U.S. Department of Agriculture, Agricultural Research Service, Western Wheat Quality Laboratory, E-202 Food Science & Human Nutrition Facility EastWashington State UniversityPullmanUSA

Personalised recommendations