DEVELOPMENT, HORMONAL, AND PATHOGENESIS-RELATED REGULATION OF THE TOBACCO CLASS-I BETA-1,3-GLUCANASE-B PROMOTER

The class I beta-1,3-glucanases are antifungal vacuolar proteins implicated in plant defense that show developmental, hormonal, and pathogenesis-related regulation. The tobacco enzymes are encoded by a small gene family with members derived from ancestors related to the present-day species Nicotiana sylvestris and N. tomentosiformis, We studied the expression in transgenic tobacco plants of a chimeric beta-glucuronidase (GUS) reporter gene fused to 1.6 kb of upstream sequence of the tobacco class I beta-1,3-glucanase B (GLB) gene, which is of N. tamentosiformis origin. Expression of the GUS reporter gene and the accumulation of class I beta-1,3-glucanase and its mRNA showed very similar patterns of regulation. In young seedlings the reporter gene was expressed in the roots. In mature tobacco plants it was preferentially expressed in lower leaves and roots and was induced in leaves by ethylene treatment and by infection with tobacco mosaic virus (TMV). Furthermore, it was down-regulated in cultured leaf discs by combinations of the hormones auxin and cytokinin. Histological studies of GUS activity showed that the GLB promoter shows highly localized expression in roots of seedlings. It is also expressed in a ring of cells around necrotic lesions induced by TMV infection, but not in cells immediately adjacent to the lesions or in the lesions themselves. The results of deletion analyses suggest that multiple positive and negative elements in the GLB promoter regulate its activity. The region from -1452 to -1193 containing two copies of the heptanucleotide AGCCGCC, which is highly conserved in plant-stress and defense-related genes, is necessary for high level expression in leaves. Additional regions important for organ-specific and regulated expression were: -568 to -402 for ethylene induction of leaves; -402 to -211 for expression in lower leaves and cultured leaf discs and for TMV induction of leaves; and -211 to -60 for expression in roots.