Methods and compositions to enhance activity of cry endotoxins

Abstract

Methods and compositions for enhancing the resistance of plants to plant pests are provided. Chimeric pesticidal polypeptides and nucleic acid molecules encoding the chimeric pesticidal polypeptides are provided. The chimeric pesticidal polypeptides comprising a solubility-enhancing polypeptide operably linked to a polypeptide comprising pesticidal activity. The nucleic acid molecules can be used in expression cassettes for making transformed plants with enhanced resistance to plant pests. Further provided are transformed plants, plant tissues, plant cells, other host cells, and seeds as well as pesticidal compositions.

That which is claimed: 
     
         1 . A method of enhancing pesticidal activity of a Cry endotoxin, the method comprising operably linking a first amino acid sequence of a solubility-enhancing polypeptide to a second amino acid sequence of a Cry endotoxin, whereby a chimeric pesticidal polypeptide is produced, the chimeric pesticidal polypeptide comprising the first amino acid sequence operably linked to second amino acid sequence. 
     
     
         2 . The method of  claim 1 ; wherein the solubility-enhancing polypeptide is selected from the group consisting of maltose-binding protein (MBP), thioredoxin, and transcription elongation factor NusA. 
     
     
         3 . The method of  claim 2 , wherein the solubility-enhancing polypeptide comprises an amino acid sequence selected from the group consisting of the amino acid sequences set forth in SEQ ID NOS: 4, 6, 34, 35 and 36. 
     
     
         4 . The method of  claim 1 ,  2  or  3 , wherein the Cry endotoxin comprises an amino acid sequence selected from the group consisting of SEQ ID NOS: 8, 10, 12 and 14. 
     
     
         5 . The method of  claim 1 ,  2 ,  3  or  4 , wherein the chimeric pesticidal polypeptide comprises an amino acid sequence selected from the group consisting of the amino acid sequences set forth in SEQ ID NOS: 2, 16, 18, 21, 23, 25, 27 and 33. 
     
     
         6 . A chimeric pesticidal polypeptide comprising a first amino acid sequence of a solubility-enhancing polypeptide operably linked to a second amino acid sequence of a Cry endotoxin, wherein the operably linked first and second amino sequences comprise an amino acid sequence selected from the group consisting of:
 (a) the amino acid sequence set forth in SEQ ID NO: 2, 16, 18, 21, 23, 25, 27 or 33; and   (b) an amino acid sequence encoded by the nucleotide sequence set forth in SEQ ID NO: 1, 15, 17, 20, 22, 24, 26 or 32.   
     
     
         7 . A nucleic acid molecule encoding the chimeric pesticidal polypeptide of  claim 6 . 
     
     
         8 . An expression cassette comprising a promoter that drives expression in a host cell operably linked to the nucleic acid molecule of  claim 7 . 
     
     
         9 . A vector comprising the expression cassette of  claim 8 . 
     
     
         10 . A transformed plant, plant part, plant cell or seed comprising in its genome the expression cassette of  claim 8 . 
     
     
         11 . A pesticidal composition comprising an effective amount of a chimeric pesticidal polypeptide of  claim 6  or an active variant or fragment thereof having pesticidal activity. 
     
     
         12 . A method of protecting a plant from an insect pest, the method comprising providing an effective amount of the pesticidal composition of  claim 10  to reduce insect pest-related damage to the plant. 
     
     
         13 . A plant comprising a polynucleotide construct stably incorporated in its genome, the polynucleotide construct comprising a nucleotide sequence operably linked to a promoter that drives expression in the plant, wherein the nucleotide sequence encodes a chimeric pesticidal polypeptide comprising a first amino acid sequence of a solubility-enhancing polypeptide operably linked to a second amino acid sequence of a Cry endotoxin, wherein the solubility-enhancing polypeptide is selected from the group consisting of maltose-binding protein (MBP), thioredoxin, and transcription elongation factor NusA. 
     
     
         14 . The plant of  claim 13 , wherein the nucleotide sequence is selected from the group consisting of:
 (a) the nucleotide sequence set forth in SEQ ID NO: 1, 15, 17, 20, 22, 24, 26 or 32; and   (b) a nucleotide sequence encoding the amino acid sequence set forth in SEQ ID NO: 2, 16, 18, 21, 23, 25, 27 or 33.   
     
     
         15 . A method of protecting a plant, plant part or plant host cell from an insect pest, the method comprising the steps of:
 (a) introducing into the plant, plant part or plant host cell an expression cassette of  claim 7 ; and   (b) regenerating the plant, plant part or plant host cell into a morphologically normal fertile plant, wherein the plant or part thereof comprises a chimeric pesticidal polypeptide.   
     
     
         16 . A method of enhancing the resistance of a plant to at least one pest, the method comprising introducing into a plant or at least one plant cell a polynucleotide construct comprising a nucleotide sequence operably linked to a promoter that drives expression in the plant, wherein the nucleotide sequence encodes a chimeric pesticidal polypeptide comprising a first amino acid sequence of a solubility-enhancing polypeptide operably linked to a second amino acid sequence of a Cry endotoxin. 
     
     
         17 . The method of  claim 16 , wherein the solubility-enhancing polypeptide is selected from the group consisting of maltose-binding protein (MBP), thioredoxin, and transcription elongation factor NusA. 
     
     
         18 . The method of  claim 17 , wherein the nucleotide sequence comprises a nucleotide sequence selected from the group consisting of:
 (a) the nucleotide sequence set forth in SEQ ID NO: 1, 15, 17, 20, 22, 24, 26 or 32; and   (b) a nucleotide sequence encoding an amino acid sequence comprising the amino acid sequence set forth in SEQ ID NO: 2, 16, 18, 21, 23, 25, 27 or 33.