Pathogenesis-related (PR) promoter
Pathogenesis-related (PR) proteins are a heterogeneous group of proteins induced in plants by pathogen infection and exogenous chemicals. PR proteins take part in the systemic acquired resistance (SAR) that develops in a resistant plant upon infection with a pathogen. As these proteins are responsive not only to pathogens but also to chemicals, the promoter sequences of these proteins have become attractive as inducible expression systems in plants. For this purpose, promoter sequences from diverse PR proteins have been isolated from plants such as Arabidopsis and maize. Recently, these systems have been developed for human cells as well.
Chemicals such as salicylic acid, ethylene, thiamine, benzol (1,2,3) thiadiazole-7-cabothionic acid S-methyl ester (BTH) have been identified as inducers of PR proteins. One of the best studied promoters is the PR-1a promoter from tobacco. The expression of the beta-glucuronidase (gus) gene, (when driven by the PR-1a promoter), increased 5-10 fold after 1-3 days of induction with salicylic acid. The same chemical induced GUS expression levels 10-fold after 8 days of spraying in field conditions. The PR-1a promoter has also been used to induce the expression ofBacillus thuringienesis delta-endotoxin in transgenic plants.
Salicylic acid is a potent inducer but there are crop tolerance problems associated with its use. In contrast, BTH does not have a phytotoxic effect and has a longer-lasting induction response of the PR-1a promoter is achieved when compared with salicylic acid. The compounds also differ in their mechanism of action: salicylic acid induces expression only in the treated tissue, whereas BTH moves systemically through the plant.
Some of the drawbacks of PR gene promoters are their inducibility by common environmental stimuli such as UV-B, ozone, and also oxidative stress. This feature might complicate the control of gene expression by PR promoters in non-laboratory conditions.
The patents covered in this section relate to DNA sequences of isolated PR-plant gene promoters, methods for inducing the transcription of a gene of interest in a plant under the control of a PR promoter, and methods for finding inducers of PR promoters. Novartis and Pioneer Hi-Bred are the main entities having patents related to these aspects.
Broad and in force patents or patent applications directed to any PR promoter or its use as inducible promoters in transformed cells were not found.
A large portfolio comprising two patent families filed by and granted to Novartis cover:
- DNA sequences of PR promoters isolated from Arabidopsis, tobacco and cucumber;
- methods for inducing the transcription of a gene of interest in a plant under the control of a PR promoter; and
- methods of inducing specific tobacco or Arabidopsis PR promoters. The claims of these patents are however, narrow because specific plant PR-1 promoters are covered
Novartis’ portfolio also includes patents granted on isolated PR proteins, methods for isolating chemically-regulatable sequences and methods for screening agrochemicals that have the ability to induce SAR response. Because these areas are outside the scope of this paper, they are not included in the present analysis.
- Pioneer Hi-Bred
Pioneer‘s United States granted patent and European patent application are directed to isolated DNA sequences of maize PR-1 gene promoters, methods of using the promoters, and transformed plants. The isolated promoters include both inducible and constitutive promoters. Some of the stimuli that induce the maize PR-1 promoters are pathogen invasion, externally applied chemicals, and environmental stresses. Plant cells and stably transformed plants containing the promoters are also part of the invention.
See table next page for summary of the related patents.