Method for improving plant tolerance to environmental stress

Abstract

Provided are DNA sequences encoding a novel type of AP2 domain-containing transcription factor as well as methods for obtaining similar sequences. Also described are methods for obtaining plants with improved growth and enhanced stress tolerance, particularly tolerance to osmotic and dehydration stress, such methods comprising expression of such DNA sequences in a plant or parts thereof. Further described are diagnostic compositions comprising the aforementioned DNA sequences and the use of such sequences in plant breeding and/or agriculture.

1 . An isolated nucleic acid encoding an AP2/EREBP domain-containing transcription factor or encoding an immunologically active and/or functional fragment of such a protein selected from the group of:
 (a) nucleic acid comprising at least part of the DNA sequence as given in SEQ ID NO 2,   (b) nucleic acid comprising the RNA sequence corresponding to at least part of SEQ ID NO 2,   (c) nucleic acid specifically hybridizing with the nucleic acid as defined in (a) or (b),   (d) nucleic acid encoding a protein having an amino acid sequence which is at least 35%, preferably 37% or 40%, and mare preferably 45% or more identical to the amino acid sequence as given is SEQ ID NO 3,   (e) nucleic acid encoding a protein comprising the amino acid sequence as given in SEQ ID NO 3 or an immunologically active or functional fragment thereof,   (f) nucleic acid encoding a protein comprising the amino acid sequence as given in SEQ ID NO 14 or comprising a sequence which is at least 75% identical to the sequence as represented in SEQ ID NO 14, or an immunologically active or functional fragment of said protein,   (g) nucleic acid which is degenerated as a result of the genetic code to a nucleic acid sequence encoding a protein as given in SEQ ID NO 3 or to a nucleic acid as defined in (a) to (f),   (h) nucleic acid which is diverging due to the differences in codon usage between the organisms to a nucleic acid sequence encoding a protein as given in SEQ ID. NO 3 or the nucleic acid as defined in (a) to (f),   (i) nucleic acid encoding a protein as given in SEQ ID. NO. 3 or as defined in (a) to (f), which are diverging due to the differences between alleles,   (j) nucleic acid encoding a protein as defined in SEQ ID NO. 3 interrupted by intervening DNA sequences or nucleic acid as defined in any one of (a) to (i) interrupted by intervening DNA sequences, and   (k) nucleic acid representing the complement of any of said nucleic acids as defined in (a) tot (j),   provided that said nucleic acid is not the nucleic acid as deposited under the GenBank accession numbers AW438153 or BF728697.   
     
     
         2 . An isolated nucleic acid according to  claim 1  encoding a polypeptide or a fragment thereof which binds to a DRE cis regulatory DNA sequence. 
     
     
         3 . An isolated nucleic acid sequence encoding an AP2/EREBP domain-containing transcription factor or fragment thereof which binds to a cis regulatory DNA sequence and for which the intensity of such binding is regulated by ABA. 
     
     
         4 . An isolated nucleic acid encoding an AP2/EREBP domain-containing transcription factor or fragment thereof which binds to a cis regulatory DNA sequence and which activates the transcription of the sequences regulated by said cis regulatory DNA sequence in response to ABA. 
     
     
         5 . An isolated nucleic acid according to  claim 3  or  4  wherein said cis regulatory DNA sequence is a DRE element. 
     
     
         6 . An isolated nucleic acid encoding an AP2/EREBP domain-containing transcription factor or an immunologically active or functional fragment thereof comprising a sequence as represented in SEQ ID NO 14, or comprising a sequence which is at least 75% identical to the sequence as represented in SEQ ID NO 14. 
     
     
         7 . An isolated nucleic acid according to any of  claims 1  to  6  which is cDNA, DNA, genomic DNA or synthetic DNA, or RNA wherein T is replaced by U. 
     
     
         8 . An isolated nucleic acid according to any of  claims 1  to  7  which is derived from a monocotyledonous plant. 
     
     
         9 . A nucleic acid molecule of at least 15 nucleotides in length hybridizing specifically with a nucleic acid of any of  claims 1  to  8 . 
     
     
         10 . A nucleic acid molecule of at least 15 nucleotides in length specifically amplifying a nucleic acid of any of  claims 1  to  8 . 
     
     
         11 . A polypeptide encoded by a nucleic acid as defined in any of  claims 1  to  8 . 
     
     
         12 . A vector comprising a nucleic acid sequence according to any of  claims 1  to  8 . 
     
     
         13 . The vector of  claim 12  which is an expression vector wherein said nucleic acid sequence is operably linked to one or more control sequences allowing the expression in prokaryotic and/or eukaryotic host cells. 
     
     
         14 . A host cell containing a nucleic acid molecule according to any of  claims 1  to  8  or a vector according to  claim 12  or  13 . 
     
     
         15 . A host cell according to  claim 14  selected from the group consisting of a bacterial, insect, fungal, plant or animal cell. 
     
     
         16 . A method for the production of transgenic plants, plant cells or plant tissue comprising the introduction of a nucleic acid of any of  claims 1  to  8  or a vector of  claim 12  or  13  into a plant, plant cell or plant tissue. 
     
     
         17 . The method of  claim 16  further comprising regenerating a plant from said plant tissue or plant cell. 
     
     
         18 . A transgenic plant cell comprising a nucleic acid of any of  claims 1  to  8  which is operably linked to regulatory elements allowing transcription and/or expression of the nucleic acid sequence in plant cells or obtainable according to the method of  claim 16  or  17 . 
     
     
         19 . The transgenic plant cell of  claim 18  wherein said nucleic acid sequence or said vector is stably integrated into the genome of the plant cell. 
     
     
         20 . A transgenic plant or a plant tissue comprising plant cells of  claim 18  or  19 . 
     
     
         21 . A method for the production of transgenic plants, said plants characterized as having increased growth and/or yield under stressed and/or non-stressed conditions, said method comprising:
 (a) introducing a nucleic acid of any of  claims 1  to  8  or a vector of  claim 12  or  13  into a plant cell or plant tissue, or,   (b) introducing a nucleic acid comprising a sequence as represented in SEQ ID NO 15, or a homologue thereof, or a vector comprising the sequence as represented in SEQ ID NO 15 into a plant cell or plant tissue,   (c) regenerating plants from said transformed plant cells or plant tissues, and,   (d) selecting a plant exhibiting said increased growth and/or yield.   
     
     
         22 . A method for the production of transgenic plants, said plants characterized as having increased expression of at least one stress-related protein, said method comprising:
 (a) introducing a nucleic acid of any of  claims 1  to  6  on a vector of  claim 12  or  13  into a plant cell or plant tissue, or   (b) introducing a nucleic add comprising a sequence as represented in SEQ ID NO 15, or a homologue thereof, or a vector comprising the sequence as represented in SEQ ID NO 15 into a plant dell or plant tissue, or   (c) regenerating plants front said transformed plant cells or plant tissues, and,   (d) selecting a plant exhibiting said increased expression of said stress-related protein.   
     
     
         23 . A transgenic plant obtainable by any of the methods of  claim 16 ,  17 ,  21  or  22 . 
     
     
         24 . A transgenic plant according to  claim 20  or obtainable by any of the methods of  claim 16 ,  17 ,  21  or  22  wherein said plant is a monocotyledonous plant. 
     
     
         25 . Plant tissue derived from a plant of  claim 20  or from a plant obtainable by a method of any of  claim 16 ,  17 ,  21  or  22 . 
     
     
         26 . A harvestable part or propagule derived from a plant of  claim 23  or  24 . 
     
     
         27 . The harvestable part of  claim 26  which is selected from the group consisting of seeds, leaves, roots, flowers; fruits, stems, rhizomes, tubers and bulbs. 
     
     
         28 . The progeny derived from any of the plants, plant tissues or plant parts of any of  claims 23  to  26 . 
     
     
         29 . A method for diagnosing changes in mRNA expression in different cells, tissues, or organs of plants and under different environmental conditions wherein a nucleic acid of any of  claims 1  to  8 , or a nucleic acid as represented in SEQ ID NO 15, or a homologue thereof, is used. 
     
     
         30 . A method for diagnosing inheritance of growth and/or yield-related traits in crop breeding programs wherein a nucleic acid of any of  claims 1  to  8 , or a nucleic acid as represented in SEQ ID NO 15, or a homologue thereof, is used as a molecular marker. 
     
     
         31 . A method for identifying allelic variants of such sequences in crop breeding programs wherein a nucleic acid of any of  claim 1  to  8 , or a nucleic acid as represented in SEQ ID NO 15, or a homologue thereof, is used. 
     
     
         32 . A method for identifying chemical compounds that modify growth and/or stress tolerance of plants, comprising:
 (a) combining a polypeptide according to  claim 11  or a polypeptide comprising a sequence as represented in SEQ ID NO 16 with said compound or mixtures of compounds under conditions suitable to allow complex formation, and,   (b) detecting complex formation, wherein the presence of a complex identifies a compound which specifically binds said polypeptide.   
     
     
         33 . Use of a compound or a mixture of compounds obtainable by a method of  claim 32  as growth regulator and/or herbicide. 
     
     
         34 . A method for identifying polypeptides that interact with a polypeptide of  claim 11  or a polypeptide comprising a sequence as represented in SEQ ID NO 16, comprising a two-hybrid screening assay wherein at least one polypeptide of  claim 11  or a polypeptide comprising a sequence as represented in SEQ ID NO 16, as a bait and a cDNA library of a plant or plant part as prey are expressed. 
     
     
         35 . A method for identifying and obtaining nucleic acids encoding DRE-binding polypeptides comprising a one hybrid screening assay wherein a DRE element, such as represented in SEQ ID NO 1 as a bait and a cDNA library of a monocotyledonous plant or plant part as prey are expressed. 
     
     
         36 . A method for identifying and obtaining nucleic acids encoding DNA-binding polypeptides that modify ABA- and stress-related responses in plants comprising a one hybrid screening assay wherein a DRE element, such as represented in SEQ ID NO 1 or a sequence which is at least 70% identical to SEQ ID NO 1 is used as a bait and a cDNA library of a monocotyledonous plant or plant part as prey are expressed. 
     
     
         37 . An isolated polypeptide obtainable by any of the methods of  claims 34  to  36 . 
     
     
         38 . An antibody specifically recognizing the nucleic add sequence as given in SEQ ID NO 1. 
     
     
         39 . An antibody specifically recognizing a polypeptide of  claim 11  or a specific epitope thereof. 
     
     
         40 . Nucleic acid encoding a polypeptide of  claim 38  or  39 . 
     
     
         41 . A vector comprising a nucleic acid of  claim 40 . 
     
     
         42 . The vector of  claim 41  which is an expression vector wherein said nucleic acid is operably linked to one or more control sequences allowing the expression in prokaryotic and/or eukaryotic host cells. 
     
     
         43 . A method for the production of transgenic monocotyledonous plants, said plants characterized as having, increased expression of at least one stress-related protein, and/or showing improved stress tolerance, and/or giving higher yield and better growth, said method comprising:
 (a) introducing a nucleic acid of  claim 40  or a vector of  claim 41  or  42  into the genome of a plant cell or plant tissue,   (b) regenerating plants from said transformed plant cells or plant tissues, and   (c) selecting a plant exhibiting said increased expression.