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KMID : 0903519960390020104
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Journal of the Korean Society of Agricultural Chemistry and Biotechnology
1996 Volume.39 No. 2 p.104 ~ p.111
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Loss of Specific Sequences in a Natural Variant of Potato Proteinase Inhibitor Ⅱ Gene Results in a Loss of Wound - Inducible Gene Expression
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Thornburg, Robert W
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Abstract
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We have isolated several proteinase inhibitor Ⅱ genes pin2 from a Russet Burbank potato DNA library. One of these, pin2T was subcloned and a 1.8 kb XbaI/NsiI insert was sequenced. This fragment contained the complete Inhibitor Ⅱ gene including 965 bp of flanking DNA upstream from the gene and 200 bp of flanking DNA downstream from the gene. The open reading frame encodes a protein that is similar to other reported proteinase Inhibitor II proteins. The DNA sequence of the 5´ flanking region of pin2T from -714 to +1 is highly homologous (91% identity) with that of the previously isolated wound-inducible pin2K. There are, however, four small deletions in the pin2T promoter which are located at -221 to -200, -263 to -254, -523 to -426 and -759 to -708 relative to the transcription start site of the wound-inducible pin2K. Three of these deletions map to a portion of the promoter that controls the wound-inducibility of the proteinase inhibitor genes. Chimeric genes containing the promoter of the pin2T gene linked with the both CAT and GUS were constructed and transfered into tobacco plants. Analysis of these plants indicated that pin2T is not a wound-inducible gene but is expressed at low levels. Thus, wound-inducibility is lost with the concomitant natural deletion of three small regions of the promoter. Comparison of the sequences deleted in pin2T relative to the pin2K with Genebank sequences indicates that the deleted sequences contain a motif (consensus 5´AGTAAA-3´) that is found in many other wound-inducible genes but not easily found in the published promoter sequences of other plant genes. Nuclear proteins from unwounded and wounded potato leaves were bound to the proximal promoter region, downstream of the 5´-AGTAAA-3´, of pin2T. The comparison of the pin2T gene with the pin2K gene indicates that the natural internal promoter deletions are likely responsible for loss of the wound-inducible phenotype in the pin2T gene.
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