Plant Defense Signal Peptides

Abstract

A 23 amino acid peptide, AtPtpl, plays an important role as a signaling component of the innate immune system of Arabidopsis . The peptide precursor gene is transcribed in response to elicitors generated by pathogens, and AtPep1 is produced to amplify the signaling pathways. Seven paralogs of the AtproPep1 gene have been identified in the Arabidopsis genome, and orthologs have been identified in species of several agriculturally important families. AtPcpl and its paralogs and orthologs play important roles as endogenous signals to amplify innate immunity. The sequence of two AtPep1 receptors from Arabidopsis are also provided.

1 . A composition comprising an isolated defense signal peptide 10 or more amino acid residues in length that has substantial defense signal peptide activity. 
   
   
       2 . The composition of  claim 1  wherein the defense signal peptide is 15 or more amino acid residues in length. 
   
   
       3 . The composition of  claim 1  wherein the defense signal peptide is 20 or more amino acid residues in length. 
   
   
       4 . The composition of  claim 1  wherein the defense signal peptide is 23 or more amino acid residues in length. 
   
   
       5 . The composition of  claim 1  wherein the defense signal peptide is between about 10 and about 50 amino acid residues in length. 
   
   
       6 . The composition of  claim 1  wherein the defense signal peptide is between about 15 and about 50 amino acid residues in length. 
   
   
       7 . The composition of  claim 1  comprising a polypeptide that is processed in a plant cell to produce the defense signal peptide. 
   
   
       8 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having at least 75 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       9 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having at least 80 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       10 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having at least 85 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       11 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having at least 90 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       12 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having complete homology with a member of the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       13 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having at least 90 percent homology with a dicot defense signal peptide consensus sequence. 
   
   
       14 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having complete homology with a dicot defense signal peptide consensus sequence. 
   
   
       15 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having at least 90 percent homology with a monocot defense signal peptide consensus sequence. 
   
   
       16 . The composition of  claim 1  wherein the defense signal peptide comprises a sequence having complete homology with a dicot defense signal peptide consensus sequence. 
   
   
       17 . The composition of  claim 1  comprising a biologically acceptable carrier. 
   
   
       18 . A plant comprising the composition of  claim 17  applied to a surface of said plant. 
   
   
       19 . A seed comprising the composition of  claim 17  applied to a surface of the seed. 
   
   
       20 . An isolated polynucleotide comprising a sequence that encodes a defense signal peptide operably linked to a plant promoter, wherein expression of the polynucleotide in a cell of a plant causes the plant to exhibit an improvement compared to a control plant lacking the polynucleotide that is selected from the group consisting of improved yield of plant product, reduced disease symptoms, and enhanced resistance to disease infestation. 
   
   
       21 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide is 10 or more amino acid residues in length. 
   
   
       22 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide is 15 or more amino acid residues in length. 
   
   
       23 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide is 20 or more amino acid residues in length. 
   
   
       24 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide is 23 or more amino acid residues in length. 
   
   
       25 . The isolated polynucleotide of  claim 20  that comprises a sequence that encodes a polypeptide that is processed in a plant cell to produce the defense signal peptide. 
   
   
       26 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide is between about 10 and about 50 amino acid residues in length. 
   
   
       27 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide is between about 15 and about 50 amino acid residues in length. 
   
   
       28 . The isolated polynucleotide of  claim 20  that encodes a polypeptide that is processed in a plant cell to produce the defense signal peptide. 
   
   
       29 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having at least 75 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       30 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having at least 80 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       31 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having at least 85 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       32 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having at least 90 percent homology with a polypeptide selected from the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       33 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having complete homology with a member of the group consisting of AtPep1, AtPep2, AtPep3, AtPep4, AtPep5, AtPep6, AtPep7, BnPep1, StPep1, PbPep1, GmPep1, MsPep1, VvPep1, OsPep1, OsPep2, TaPep1, TaPep2, ZmPep1, and HvPep2. 
   
   
       34 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having at least 90 percent homology with a dicot defense signal peptide consensus sequence. 
   
   
       35 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having complete homology with a dicot defense signal peptide consensus sequence. 
   
   
       36 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having at least 90 percent homology with a monocot defense signal peptide consensus sequence. 
   
   
       37 . The isolated polynucleotide of  claim 20  wherein the defense signal peptide comprises a sequence having complete homology with a dicot defense signal peptide consensus sequence. 
   
   
       38 . The polynucleotide of  claim 20  wherein the sequence that encodes a defense signal peptide has at least 80 percent sequence similarity to a polynucleotide sequence selected from the group consisting of AtproPep1, AtproPep2, AtproPep3, AtproPep4, AtproPep5, AtproPep6, AtproPep7, BnproPep1, StproPep1, PbproPep1, GmproPep1, MsproPep1, VvproPep1, OsproPep1, OsproPep2, TaproPep1, TaproPep2, ZmproPep1, and HvproPep2. 
   
   
       39 . The polynucleotide of  claim 20  wherein the sequence that encodes a defense signal peptide has at least 90 percent sequence similarity to a polynucleotide sequence selected from the group consisting of AtproPep1, AtproPep2, AtproPep3, AtproPep4, AtproPep5, AtproPep6, AtproPep7, BnproPep1, StproPep1, PbproPep1, GmproPep1, MsproPep1, VvproPep1, OsproPep1, OsproPep2, TaproPep1, TaproPep2, ZmproPep1, and HvproPe. 
   
   
       40 . The polynucleotide of  claim 20  wherein the sequence that encodes a defense signal peptide has at least 95 percent sequence similarity to a polynucleotide sequence selected from the group consisting of AtproPep1, AtproPep2, AtproPep3, AtproPep4, AtproPep5, AtproPep6, AtproPep7, BnproPep1, StproPep1, PbproPep1, GmproPep1, MsproPep1, VvproPep1, OsproPep1, OsproPep2, TaproPep1, TaproPep2, ZmproPep1, and HvproPe. 
   
   
       41 . The polynucleotide of  claim 20  wherein the sequence that encodes a defense signal peptide has complete sequence similarity to a polynucleotide sequence selected from the group consisting of AtproPep1, AtproPep2, AtproPep3, AtproPep4, AtproPep5, AtproPep6, AtproPep7, BnproPep1, StproPep1, PbproPep1, GmproPep1, MsproPep1, VvproPep1, OsproPep1, OsproPep2, TaproPep1, TaproPep2, ZmproPep1, and HvproPe. 
   
   
       42 . An isolated polynucleotide comprising a sequence that encodes a defense signal peptide operably linked to a heterologous promoter. 
   
   
       43 . The isolated polynucleotide of  claim 42  wherein expression of the polynucleotide in a cell of a plant causes the plant to exhibit an improvement compared to a control plant lacking the polynucleotide that is selected from the group consisting of improved yield of plant product, reduced disease symptoms, and enhanced resistance to disease infestation. 
   
   
       44 . The isolated polynucleotide of  claim 42  wherein the promoter is a constitutive promoter. 
   
   
       45 . The polynucleotide of  claim 42  wherein the promoter is a non-constitutive promoter. 
   
   
       46 . The polynucleotide of  claim 42  wherein the promoter is an organ- or tissue-specific promoter. 
   
   
       47 . The polynucleotide of  claim 42  wherein the promoter is an inducible promoter. 
   
   
       48 . A cell comprising the isolated polynucleotide of  claim 42 . 
   
   
       49 . The cell of  claim 48  selected from the group consisting of a plant cell, a bacterial cell, a fungal cell and an insect cell. 
   
   
       50 . A plant cell of  claim 48 . 
   
   
       51 . A plant comprising the plant cell of  claim 50 . 
   
   
       52 . A plant of  claim 51  selected from the group consisting of  Acacia , alfalfa, aneth, apple, apricot, artichoke, arugula, asparagus, avocado, banana, barley, beans, beet, blackberry, blueberry, broccoli, brussels sprouts, cabbage, cantaloupe, carrot, cassaya, castorbean, cauliflower, celery, cherry, chicory, cilantro, citrus, clementines, clover, coconut, coffee, corn, cotton, cucumber, Douglas fir, eggplant, endive, escarole, eucalyptus, fennel, figs, garlic, gourd, grape, grapefruit, honey dew, jicama, kiwifruit, lettuce, leeks, lemon, lime, Loblolly pine, linseed, mango, melon, mushroom, nectarine, nut, oat, oil palm, oil seed rape, okra, olive, onion, orange, an ornamental plant, palm, papaya, parsley, parsnip, pea, peach, peanut, pear, pepper, persimmon, pine, pineapple, plantain, plum, pomegranate, poplar, potato, pumpkin, quince, radiata pine, radicchio, radish, rapeseed, raspberry, rice, rye, sorghum, Southern pine, soybean, spinach, squash, strawberry, sugarbeet, sugarcane, sunflower, sweet potato, sweetgum, tangerine, tea, tobacco, tomato, triticale, turf grass, turnip, a vine, watermelon, wheat, yams, and zucchini. 
   
   
       53 . The plant of  claim 51  that exhibits reduced symptoms from, or enhanced resistance to, a disease caused by an organism of a genus selected from the group consisting of  Alternaria, Ascochyta, Aspergillus, Botrytis, Cercospora, Colletotrichum, Diplodia, Erwinia, Erysiphe, Fusarium, Gaeumanomyces, Helminthosporium, Macrophomina, Magnaporthe, Mycosphaerella, Nectria, Peronospora, Phoma, Phymatotrichum, Phytophthora, Plasmopara, Podosphaera, Pseudomonas, Puccinia, Puthium, Pyrenophora, Pyricularia, Pythium, Rhizoctonia, Scerotium, Sclerotinia, Septoria, Thielaviopsis, Uncinula, Venturia, Verticillium , and  Xanthomonas.    
   
   
       54 . A part of the plant of  claim 51  selected from the group consisting of seeds, seed pods, flowers, fruit, tubers, stems, cuttings, and pollen. 
   
   
       55 . A product resulting from processing a plant of  claim 51  or a part thereof. 
   
   
       56 . A composition comprising the polynucleotide of  claim 42  and a biologically acceptable carrier. 
   
   
       57 . A method of making a defense signal peptide comprising expressing in a cell a polynucleotide of  claim 42 . 
   
   
       58 . The method of  claim 57  wherein the cell is a bacterial cell, a fungal cell, or an insect cell. 
   
   
       59 . The method of  claim 57  further comprising purifying the defense signal peptide. 
   
   
       60 . A method of making a transgenic plant comprising introducing into a cell of a plant a polynucleotide of  claim 42 , thereby producing a transformed cell, and regenerating a transgenic plant from the transformed cell, wherein, compared to a control plant lacking the polynucleotide, the transgenic plant exhibits a characteristic selected from the group consisting of substantially improved yield of plant product, substantially reduced disease symptoms, and substantially enhanced resistance to disease infestation. 
   
   
       61 . A method of making a plant that comprises a transgene comprising a sequence that encodes a defense signal peptide operably linked to a plant promoter, the method comprising sexually crossing a plant that comprises the transgene with a plant that lacks the transgene, thereby producing a plurality of progeny plants, and selecting a progeny plant comprising the transgene. 
   
   
       62 . A method of making a plant that comprises a transgene comprising a sequence that encodes a defense signal peptide operably linked to a plant promoter, the method comprising asexually reproducing a plant that comprises the transgene, thereby producing a plurality of progeny plants, and selecting a progeny plant comprising the transgene. 
   
   
       63 . A method of growing a plant comprising planting a seed that comprises a polynucleotide sequence of  claim 42 , and growing the seed to produce a plant, wherein, compared to a control plant lacking said polynucleotide sequence, the plant grown from the seed exhibits a characteristic selected from the group consisting of substantially improved yield of plant product, substantially reduced disease symptoms, and substantially enhanced resistance to disease infestation. 
   
   
       64 . A method for detecting a plant cell comprising a polynucleotide of  claim 42  in a biological sample, the method comprising contacting the biological sample with a probe that binds specifically to the polynucleotide, and detecting said binding. 
   
   
       65 . The method of  claim 64  wherein the probe is a PCR primer, the method comprising performing PCR on the sample and detecting said binding by detecting an amplification product diagnostic of the presence of the polynucleotide in the sample. 
   
   
       66 . A kit for detecting a plant cell comprising a polynucleotide of  claim 42  in a biological sample, the kit comprising one or more probes that bind specifically to the polynucleotide, and instructions for use. 
   
   
       67 . A method for detecting a plant cell comprising a polynucleotide of  claim 42  in a biological sample, the method comprising contacting the biological sample with a probe that binds specifically to the defense signal peptide, and detecting said binding. 
   
   
       68 . The method of  claim 67  wherein the probe is an antibody. 
   
   
       69 . A kit for detecting a plant cell comprising a polynucleotide of  claim 42  in a biological sample, the kit comprising one or more probes that bind specifically to the defense signal peptide, and instructions for use. 
   
   
       70 . A plant cell comprising an insertion of a foreign promoter upstream of a coding sequence for a defense signal protein, wherein the foreign promoter is operably linked to the coding sequence for the defense signal protein and the plant is characterized by a substantially enhanced resistance to a disease compared to a control plant lacking the insertion of the foreign promoter. 
   
   
       71 . A method of making a transgenic plant comprising (a) introducing into cells of a plant a polynucleotide that comprises a heterologous promoter, thereby producing a cell comprising an insertion of the heterologous promoter upstream of a coding sequence for a defense signal protein, wherein expression of the defense signal protein is controlled by the foreign promoter, and (b) regenerating a transgenic plant from said cell comprising the insertion. 
   
   
       72 . A method of identifying a defense signal peptide comprising: (a) providing a plurality of candidate peptides having a length of at least 10 amino acids; (b) assaying said plurality of candidate peptides for defense signal peptide activity in an alkalinization assay; and (c) selecting a candidate peptide that has substantial defense signal peptide activity. 
   
   
       73 . The method of  claim 72  comprising providing the candidate peptides by chemically synthesizing said plurality of candidate peptides. 
   
   
       74 . The method of  claim 72  comprising administering the candidate peptide to a plant by applying a composition comprising the candidate peptide to the plant. 
   
   
       75 . The method of  claim 72  comprising administering the candidate peptide to a plant by expressing within a cell of the plant a polynucleotide that comprises a sequence that encodes the candidate peptide, thereby producing the candidate peptide within the cell of the plant. 
   
   
       76 . A method of identifying a substance that enhances defense of a plant against a disease comprising (a) contacting an isolated AtPep1 receptor with a plurality of candidate substances; (b) selecting a candidate substance that has a detectable interaction with the isolated AtPep1 receptor; and (c) applying the selected candidate substance to a plant to determine whether the selected candidate substance enhances defense of the plant against a disease. 
   
   
       77 . The method of  claim 76  wherein the candidate substances are peptides. 
   
   
       78 . The method of  claim 76  wherein the candidate substances are non-peptide chemical compounds.