Multicellular living organisms and unmodified parts thereof and – Method of using a plant or plant part in a breeding process...
Reexamination Certificate
1998-11-23
2002-07-09
Riley, Jezia (Department: 1656)
Multicellular living organisms and unmodified parts thereof and
Method of using a plant or plant part in a breeding process...
C435S006120, C536S022100, C536S023100
Reexamination Certificate
active
06417428
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to the regulatory response of plants to environmental stresses such as cold and to drought. More specifically, the present invention relates to genes which regulate the response of a plant to environmental stresses such as cold or drought and their use to enhance the stress tolerance of recombinant plants into which these genes are introduced.
BACKGROUND OF THE INVENTION
Environmental factors serve as cues to trigger a number of specific changes in plant growth and development. One such factor is low temperature. Prominent examples of cold-regulated processes include cold acclimation, the increase in freezing tolerance that occurs in response to low non-freezing temperatures (Guy, C. L., Annu. Rev. Plant Physiol. Plant Mol. Biol. 41:187-223 (1990)); vernalization, the shortening of time to flowering induced by low temperature (Lang, A., in Encyclopedia of Plant Physiology, Vol.15-1, ed. Ruhland, W. (Springer, Berlin), pp. 1489-1536 (1965)); and stratification, the breaking of seed dormancy by low temperature (Berry, J. A. and J. K. Raison, in Encyclopedia of Plant Physiology, Vol. 12A, eds. Lange, O. L., Nobel, P. S., Osmond, C. B. and Ziegler, H. (Springer, Berlin), pp. 277-338 (1981)). Due to the fundamental nature and agronomic importance of these processes, there is interest in understanding how plants sense and respond to low temperature. One approach being taken is to determine the signal transduction pathways and regulatory mechanisms involved in cold-regulated gene expression.
Strong evidence exists for calcium having a role in low temperature signal transduction and regulation of at least some COR (cold-regulated) genes. Dhindsa and colleagues (Monroy, A. F., et al, Plant Physiol. 102:1227-1235 (1993); Monroy, A. F., and R. S., The Plant Cell, 7:321-331 (1995)) have shown that, in alfalfa, calcium chelators and calcium channel blockers prevent low temperature induction of COR genes and that calcium ionophores and calcium channel agonists induce expression of COR genes at normal growth temperatures. Similarly, Knight et al (The Plant Cell 8:489-503 (1996)) have shown that cold-induced expression of the
Arabidopsis thaliana
COR gene KIN1 is inhibited by calcium chelators and calcium channel blockers. These results suggest that low temperature triggers an influx of extracellular calcium that activates a signal transduction pathway that induces the expression of COR genes. Consistent with this notion is the finding that low temperature evokes transient increases in cytosolic calcium levels in plants (Knight, M. R. et al, Nature 352:524-526 (1991); Knight, H., et al., The Plant Cell 8:489-503 (1996)). In addition, low temperatures have been shown to stimulate the activity of mechanosensitive calcium-selective cation channels in plants (Ding, J. P. and B. G. Pickard, Plant J. 3:713-720 (1993)).
Recent efforts have led to the identification of a cis-acting cold-regulatory element in plants, the C-repeat/DRE (Yamaguchi-Shinozaki, et al., The Plant Cell 6:251-264 (1994); Baker, S. S., et al., Plant. Mol. Biol. 24:701-713 (1994); Jiang, C., et al., Plant Mol. Biol. 30:679-684 (1996)). The element, which has a 5 base pair core sequence for CCGAC, is present once to multiple times in all plant cold-regulated promoters that have been described to date; these include the promoters of the COR15a (Baker, S. S., et al, Plant. Mol. Biol. 24:701-713 (1994)), COR78/RD29A (Horvath, D. P., et al, Plant Physiol. 103:1047-1053 (1993); Yamaguchi-Shinozaki, K., et al., The Plant Cell 6:251-264 (1994)), COR6.6 (Wang, H., et al., Plant Mol. Biol. 28:605-617 (1995)) and KIN1 (Wang, H., et al, Plant Mol. Biol. 28:605-617 (1995)) genes of Arabidopsis and the BN115 gene of
Brassica napus
(White, T. C., et al, Plant Physiol. 106:917-928 (1994)). Deletion analysis of the Arabidopsis COR15a gene suggested that the CCGAC sequence, designated the C-repeat, might be part of a cis-acting cold-regulatory element (Baker, S. S., et al., Plant Mol. Biol. 24:701-713 (1994)). That this was the case was first demonstrated by Yamaguchi-Shinozaki and Shinozaki (Yamaguchi-Shinozaki, K., et al., The Plant Cell 6:251-264 (1994)) who showed that two of the C-repeat sequences present in the promoter of COR78/RD29A induced cold-regulated gene expression when fused to a reporter gene. It was also found that these two elements stimulate transcription in response to dehydration and high salinity and thus, was designated the DRE (dehydration, low temperature and high salt regulatory element). Recent studies by Jiang et al (Jiang, C., et al., Plant Mol. Biol. 30:679-684 (1996)) indicate that the C-repeats (referred to as low temperature response elements) present in the promoter of the
B. napus
BN115 gene also impart cold-regulated gene expression.
U.S. Pat. Nos. 5,296,462 and 5,356,816 to Thomashow describe the genes encoding the proteins involved in cold adaptation in
Arabidopsis thaliana
. In particular the DNA encoding the COR15 proteins is described. These proteins are significant in promoting cold tolerance in plants.
A need exists for the identification of genes which regulate the expression of cold tolerance genes and drought tolerance genes. A further need exists for DNA constructs useful for introducing these regulatory genes into a plant in order to cause the plant to begin expressing or enhance their expression of native or non-native cold tolerance genes and drought tolerance genes. These and other needs are provided by the present invention.
SUMMARY OF THE INVENTION
DNA in isolated form is provided which includes a sequence encoding a binding protein capable of selectively binding to a DNA regulatory sequence which regulates expression of one or more environmental stress tolerance genes in a plant. The binding protein is preferably capable of regulating expression of one or more environmental stress tolerance genes in a plant by selectively binding to a DNA regulatory sequence which regulates the one or more environmental stress tolerance genes. In one embodiment, the binding protein is a non-naturally occurring protein formed by combining an amino acid sequence capable of binding to a CCG regulatory sequence, preferably a CCGAC regulatory sequence with an amino acid sequence which forms a transcription activation region which regulates expression of one or more environmental stress tolerance genes in a plant by regulating expression of one or more environmental stress tolerance genes when the binding protein binds to the regulatory region.
DNA in isolated form is also provided which includes a promoter and the sequence encoding the binding protein. In one variation, the promoter causes expression of the binding protein in a manner which is different than how the binding protein is expressed in its native state. For example, the promoter may increase the level at which the binding protein is expressed, express the binding protein without being induced by an environmental stress and/or express the binding protein in response to a different form or degree of environmental stress than would otherwise be needed to induce expression of the binding protein. The promoter may also be inducible by an exogenous agent. The promoter can also be selected with regard to the type or types of plant tissues that the binding protein will be expressed as well as when in the plant's life the promoter will function to regulate expression of the binding protein.
A nucleic acid construct capable of transforming a plant is also provided which includes a sequence encoding a binding protein capable of selectively binding to a DNA regulatory sequence which regulates expression of one or more environmental stress tolerance genes in a plant. The binding protein is preferably capable of regulating expression of one or more environmental stress tolerance genes in a plant by selectively binding to a DNA regulatory sequence which regulates the one or more environmental stress tolerance genes. The nucleic acid construct may be an RNA or DNA construct. Examples of types of construc
Gilmour Sarah Jane
Jaglo-Ottosen Kirsten
Jiang Cai-Zhong
Stockinger Eric J.
Thomashow Michael F.
Guerrero Karen
Riley Jezia
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