cis-analysis of the wound-inducible promoter wun1 in transgenic tobacco plants and histochemical localization of its expression.

Abstract

The 5' region of the wound-inducible gene wun1, derived from potato, has been sequenced and analyzed for cis-acting elements important in controlling gene expression in transgenic tobacco plants. Different 5' deletion fragments were linked to the reporter gene beta-glucuronidase (GUS) as transcriptional fusions, and the expression of these chimeric genes was analyzed in leaf tissue. Sequences 111 base pairs upstream of the transcriptional start site were not able to drive the GUS expression over background levels, whereas sequences between -111 and -571 showed a slightly higher activity with equal levels of transcription in wounded and nonwounded tissue. The addition of further upstream sequences (-571 to -1022) enhanced the level of expression by a factor between 13 and 370. The expression driven by this fragment was inducible by a factor of twofold to ninefold by wounding. Histochemical analysis of different tissue from transgenic plants that contain wun1-GUS fusions demonstrates wound-inducible and cell-specific wun1 promoter activity in plants containing the -1022-base pair fragment. The location of GUS activity appears to be cell-specific, being highest in epidermal cells of leaves and stems and lower in vascular cells. Activity was reduced to levels that could not be detected by histochemical staining in leaves, stems, and roots of plants containing the deleted promoter fragments. Plants that contain the different deletion constructs and plants that carry the -1022-base pair fragment show high expression in anthers and pollen grains that could not be stimulated by wounding.