Plant transcriptional regulators of abiotic stress

Abstract

The invention relates to plant transcription factor polypeptides, polynucleotides that encode them, homologs from a variety of plant species, and methods of using the polynucleotides and polypeptides to produce transgenic plants having advantageous properties, including improved drought and other osmotic stress tolerance, as compared to wild-type or reference plants. Sequence information related to these polynucleotides and polypeptides can also be used in bioinformatic search methods to identify related sequences and is also disclosed.

What is claimed is:  
     
         1 . A recombinant polynucleotide comprising a nucleotide sequence that hybridizes over its full length to the complement of SEQ ID NO 1 under stringent conditions that include two wash steps of 6x SSC and 65° C., each step being 10-30 minutes in duration.  
     
     
         2 . The recombinant polynucleotide of  claim 1 , wherein the polynucleotide is operably linked to at least one regulatory element being effective in controlling expression of said isolated polynucleotide when said isolated polynucleotide is transformed into a plant.  
     
     
         3 . The recombinant polynucleotide of  claim 1 , wherein said recombinant polynucleotide is incorporated into an expression vector.  
     
     
         4 . The recombinant polynucleotide of  claim 3 , wherein said recombinant polynucleotide is incorporated into a cultured host cell.  
     
     
         5 . The recombinant polynucleotide of  claim 1 , wherein the recombinant polynucleotide encodes a polypeptide comprising the AP2 domain, the DML motif, and the B3 domain of SEQ ID NO:2.  
     
     
         6 . The recombinant polynucleotide of  claim 1 , wherein said recombinant polynucleotide encodes a polypeptide comprising SEQ ID NO:2.  
     
     
         7 . The recombinant polynucleotide of  claim 1 , wherein said recombinant polynucleotide comprises SEQ ID NO:1.  
     
     
         8 . An isolated nucleotide sequence that hybridizes over its full length to the complement of a polynucleotide encoding SEQ ID NO 64, which represents the DML motif of SEQ ID NO:1, under stringent conditions that include two wash steps of 6×SSC and 65° C., each step being 10-30 minutes in duration.  
     
     
         9 . The isolated nucleotide sequence of  claim 8 , wherein the isolated nucleotide sequence encodes a polypeptide comprising a DML motif that is substantially identical to SEQ ID NO 64.  
     
     
         10 . The isolated nucleotide sequence of  claim 8 , wherein said isolated nucleotide sequence is incorporated into an expression vector.  
     
     
         11 . The isolated nucleotide sequence of  claim 8 , wherein said isolated nucleotide sequence is incorporated into a cultured host cell.  
     
     
         12 . A transgenic plant that overexpresses the recombinant polynucleotide according to  claim 1 , wherein said transgenic plant has increased abiotic stress tolerance as compared to a non-transformed plant that does not overexpress a polypeptide encoded by the recombinant polynucleotide.  
     
     
         13 . A transgenic plant comprising a recombinant polynucleotide encoding a polypeptide having an AP2 domain and a B3 domain, wherein the polypeptide has the property of SEQ ID NO:2 of regulating abiotic stress tolerance in a plant when said polypeptide is overexpressed, wherein: 
 the AP2 domain is sufficiently homologous to the AP2 domain of SEQ ID NO:2 that the polypeptide binds to a transcription-regulating region comprising the motif CAACA;    the B3 domain is sufficiently homologous to the B3 domain of SEQ ID NO:2 that the polypeptide binds to a transcription regulating region comprising the motif CACCTG; and    wherein said binding cooperatively enhances DNA binding affinity of said polypeptide and overexpression of said polypeptide confers increased abiotic stress tolerance in said transgenic plant as compared to a non-transformed plant that does not overexpress the polypeptide.    
     
     
         14 . The transgenic plant of  claim 13 , wherein said polypeptide comprises SEQ ID NO:2.  
     
     
         15 . The transgenic plant of  claim 13 , wherein said recombinant polynucleotide comprises SEQ ID NO:1.  
     
     
         16 . The transgenic plant of  claim 13 , wherein said recombinant polynucleotide comprises SEQ ID NO:64, which represents the DML motif of SEQ ID NO:1.  
     
     
         17 . The transgenic plant of  claim 13 , wherein said abiotic stress tolerance is selected from the group consisting of drought tolerance, heat tolerance, and salt stress tolerance.  
     
     
         18 . The transgenic plant of  claim 13 , wherein the plant is selected from the group consisting of: soybean, wheat, corn, potato, cotton, rice, oilseed rape, sunflower, alfalfa, clover, sugarcane, turf, banana, blackberry, blueberry, strawberry, raspberry, cantaloupe, carrot, cauliflower, coffee, cucumber, eggplant, grapes, honeydew, lettuce, mango, melon, onion, papaya, peas, peppers, pineapple, pumpkin, spinach, squash, sweet corn, tobacco, tomato, watermelon, mint and other labiates, citrus, fruit trees, rosaceous fruits, and brassicas.  
     
     
         19 . The transgenic plant of  claim 13 , further comprising a constitutive, inducible, or tissue-specific promoter operably linked to said polynucleotide sequence.  
     
     
         20 . The transgenic plant of  claim 13 , wherein said polypeptide is selected from the group consisting of SEQ ID NOs:2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 38, 40, 42, 44, 46, 48, 50, 52, and 53.  
     
     
         21 . A method for producing a transgenic plant having increased tolerance to abiotic stress, the method steps comprising: 
 (a) providing the expression vector according to  claim 3;     (b) introducing the expression vector into a plant cell, and allowing the plant cell to overexpress a polypeptide encoded by the recombinant polynucleotide, said polypeptide having the property of regulating abiotic stress tolerance in a plant as compared to a non-transformed plant that does not overexpress the polypeptide;    (c) growing the plant cell into a plant; and    (d) identifying an abiotic stress tolerant plant so produced with increased abiotic stress tolerance by comparing said abiotic stress tolerant plant with one or more non-transformed plants that do not overexpress the polypeptide.    
     
     
         22 . The method of  claim 21 , the method steps further comprising: 
 (e) selfing or crossing said abiotic stress tolerant plant with itself or another plant, respectively, to produce seed; and    (f) growing a progeny plant from the seed, thus producing a transgenic progeny plant having increased tolerance to abiotic stress.    
     
     
         23 . The method of  claim 22 , wherein: 
 said progeny plant expresses MRNA that encodes a DNA-binding protein having an AP2 domain that binds to a DNA molecule, regulates expression of said DNA molecule, and induces expression of a plant trait gene; and    said mRNA is expressed at a level greater than a non-transformed plant that does not overexpress said DNA-binding protein.    
     
     
         24 . The method of  claim 21 , wherein said abiotic stress tolerance is selected from the group consisting of heat tolerance, chilling tolerance, germination in heat, germination in cold, drought stress tolerance, and salt stress tolerance.  
     
     
         25 . The method of  claim 21 , wherein the polypeptide comprises an AP2 domain, a DML motif, and a B3 domain, in order from N-terminal to C-terminal, respectively.  
     
     
         26 . A method for increasing a plant's tolerance to abiotic stress, said method comprising: 
 (a) providing a vector comprising: 
 (i) regulatory elements flanking the polynucleotide sequence, said regulatory elements being effective to control expression of said polynucleotide sequence in a target plant; and  
 (ii) a polynucleotide sequence that encodes a polypeptide having an AP2 domain that is sufficiently homologous to the AP2 domain of SEQ ID NO:2 that the polypeptide binds to a first transcription regulating region comprising the motif CAACA, and the B3 domain is sufficiently homologous to the B3 domain of SEQ ID NO:2, that the polypeptide binds to a second transcription regulating region comprising the motif CACCTG, and the polypeptide has the property of SEQ ID NO:2 of regulating abiotic stress tolerance in a plant, wherein said binding of said first and second transcription regulating regions confers increased abiotic stress tolerance in said transgenic plant, as compared to a non-transformed plant that does not overexpress the polypeptide; and  
   (b) transforming the target plant with said vector to generate a transformed plant with increased tolerance to abiotic stress.    
     
     
         27 . The method of  claim 26 , wherein said polynucleotide comprises: 
 (i) SEQ ID NO:1;    (ii) a nucleotide sequence that encodes SEQ ID NO:2;    (iii) a nucleotide sequence that hybridizes to the nucleotide sequence of (i) or (ii) under stringent conditions of two washes of 6×SSC and 65° C., each step being 10-30 minutes in duration;    (iv) a nucleotide sequence encoding a polypeptide comprising an AP2 domain and a B3 domain that are substantially identical with the AP2 and B3 domains of SEQ ID NO:2, respectively; or    (v) a nucleotide sequence encoding a polypeptide comprising a DML motif that is substantially identical to SEQ ID NO:64, the DML motif of SEQ ID NO:2.    
     
     
         28 . The method of  claim 26 , wherein said abiotic stress tolerance is selected from the group consisting of heat tolerance, chilling tolerance, germination in heat, germination in cold, drought stress tolerance, and salt stress tolerance.