Invented by Jen Sheen, Yelena V. Kovtun, Wan-Ling Chiu, General Hospital Corp
The market for transgenic plants that express a MAPKKK domain protein kinase is driven by the increasing demand for crops that can withstand harsh environmental conditions, such as drought, salinity, and extreme temperatures. These stresses can significantly reduce crop yields, leading to food shortages and economic losses. Transgenic plants that express a MAPKKK domain protein kinase have been shown to have improved tolerance to these stresses, resulting in higher yields and better quality crops.
Another factor driving the market for transgenic plants that express a MAPKKK domain protein kinase is the growing awareness of the environmental impact of traditional farming practices. The use of pesticides and fertilizers can have negative effects on soil health and water quality, as well as on the health of farmers and consumers. Transgenic plants that express a MAPKKK domain protein kinase can reduce the need for these inputs, as they are naturally resistant to pests and diseases.
The market for transgenic plants that express a MAPKKK domain protein kinase is also supported by advances in genetic engineering technology. The development of new gene editing techniques, such as CRISPR-Cas9, has made it easier and more efficient to introduce specific genes into plant genomes. This has led to the creation of new varieties of transgenic plants that express a MAPKKK domain protein kinase with improved traits, such as higher yields, better nutritional content, and increased resistance to multiple stresses.
However, there are also concerns about the safety and environmental impact of transgenic plants. Critics argue that the long-term effects of these plants on ecosystems and human health are not fully understood, and that the use of genetically modified organisms (GMOs) could have unintended consequences. As a result, there is a need for rigorous testing and regulation of transgenic plants that express a MAPKKK domain protein kinase, to ensure that they are safe for human consumption and do not harm the environment.
In conclusion, the market for transgenic plants that express a MAPKKK domain protein kinase is growing rapidly, driven by the need for crops that can withstand environmental stresses and the development of new genetic engineering technologies. However, there are also concerns about the safety and environmental impact of these plants, highlighting the need for careful regulation and monitoring of their use. As research in this field continues, it is likely that we will see further advances in the development of transgenic plants that can help to address some of the biggest challenges facing agriculture today.
The General Hospital Corp invention works as follows
The invention also includes plants that contain a recombinant transgene capable of expressing a kinase-domain of a mitogen activated protein kinase, kinase, kinase, kinase, kinase, kinase or kinase-domain of said MAPKKK, thus increasing the level stress resistance or tolerability in the transgenic. The invention also features plants that contain a recombinant recombinant expressing the kinase of a mitogen activated protein kinase, kinase-kinase or kinase of said MAPKKKK.
Background for Transgenic plants that express a MAPKKK domain protein kinase
This invention is related to the manipulation and production of transgenic plant genes.
Auxin, an essential plant hormone, regulates diverse processes such as cell division, root and leaves development, apical dominant, tropism and reproduction” (Davies P. J. In: Plant Hormones, edited by Davies P. J. 1-12, Kluwer, Dordrecht, Netherlands, 1995). The auxin response in plants is controlled by a complex network of signaling pathways. This balance is achieved between auxin, other signaling pathways that are synergistic or antagonistic, and the auxin (Bellincampi, Plant Cell, 8: 477-487; Coenen, Trends Plant Sci. 2: 351-356, 1997). The activation of early response genes is a primary event in auxin action. Several auxin-responsive cis-elements, as well as trans-acting factor have been identified in the promoters of early response genes (Abel et.al., Plant Physiol. 111: 9-17, 1996; Ulmasov et al., Science 276: 1865-1868, 1997). While genetic approaches have made significant advances in our understanding of auxin activity (Walden and al., Trends Plant Sci. 1: 335-339, 1996; Leyser, Curr. Biol. 8: R305-R307, 1998; Guilfoyle, Trends Plant Sci. The molecular mechanisms that underlie signal transduction pathways to control auxin-responsive transcription are still largely unknown.
In yeast, worms and insects, as well as mammals, the primary response to hormones, growth and stress signals is mediated by a conservation signaling cascade that consists of three proteins kinases: mitogen activated protein kinase kinase kinase kinase kinase kinase kinasase kinase kinase kinase kinase kina MAPKKK phosphorylates MAPKK, which in turn phosphorylates MAPK. The activated MAPK is translocated to the nucleus, where it phosphorylates transcription factor that controls gene expression (Herskowitz Cell 80: 187-197, 1995; Kyriakis et.al., J. Biol. Chem. 271: 24313-24316, 1996). The sequence conservation of MAPK, MAPKK and MAPKKK homologues in plants and their functional complementation with yeast have led to the identification of many, but their exact physiological functions are still unknown. Sci. 2: 11-15, 1997). It is also unclear whether or how these homologues comprise specific MAPK kinase cleavages (Mizoguchi et. al., Trends in Biotech). 15: 15-19, 1997).
Plants are continually exposed to environmental stimuli which influence their growth and developement. Heat, salinity and freezing are just a few of the adverse environmental conditions that can reduce plant productivity. Plant tolerance to stress has been enhanced by genetic approaches that alter osmolytes and osmoprotectants as well as membrane fatty acid, channels, transcription factor, and enzymes which scavenge oxygen-active species. There is a need to develop molecular strategies to enable plants to be resistant or tolerable to adverse environmental conditions.
The invention is based upon the applicants’ discovery that mitogen-activated polypeptides, such as NPK1 from tobacco or the ANPs from Arabidopsis are involved in signaling activation of stress protective genes transcription, repression early auxin response gene transcript, and alteration of seed growth. The invention relates to methods for genetically engineering plants in order to induce altered agronomic or physiological changes, as well as developmental changes, by expressing transgenes encoding DNA encoding the kinase-domain of a MAPKKK. It has been discovered that it is possible for plants to express a recombinant MAPKKK which are resistant to many stresses (e.g. drought, increased saltiness, heat shock and freezing temperatures), have suppressed early auxin gene or altered seed development.
The invention, in one aspect, features a method of increasing stress tolerance or resistance in a plant. The method includes: (a), introducing a transgene containing DNA encoding the kinase of a MAPKKK operably coupled to a plant cell promoter to produce transformed plant cells. (b), generating a plant transgenic from the transformed cells. In preferred embodiments of the invention, the expression DNA encoding for the kinase-domain activates the transcription of a stress inducible gene. The method can be used to provide a plant with resistance or tolerance against an environmental stress. Environmental stresses that are exemplary include, but are not limited to, those resulting from the exposure of a transgenic plant with inadequate or no water (e.g. drought conditions), excessive salt or osmotic condition, extreme temperature conditions (e.g. heat, cold or frost), too much light, pathogens, chemicals (e.g. Metals, herbicides and pollutants, an oxidative state, UV light and wounding are examples of environmental stresses. The plant is protected from multiple stress conditions in preferred embodiments.
The invention also includes a method to reduce the effect of an auxin on a plant. The method comprises the following steps: (a), introducing into plant cell a transgene containing DNA encoding the kinase of a MAPKKK operably coupled to a plant cell promoter to produce transformed plant cells; (b), regenerating a plant transgenic from the transformed cells wherein the MAPKKK kinase is expressed in cells of the plant transgenic, thereby reducing auxin action in the plant transgenic. The expression of DNA encoding a kinase-domain represses an early-auxin (e.g. those that are controlled by a promoter substantially identical to the GH3 or a ER7 element) gene.
The invention also includes a method of altering the development of seeds. The method comprises the following steps: (a), introducing a transgene containing DNA encoding the kinase of a MAPKKK operably coupled to a plant cell promoter to produce transformed plant cells, (b) generating a plant transgenic from the transformed cells wherein the MAPKKK kinase is expressed in cells of the plant transgenic, thus altering the seed development in the plant transgenic. In preferred embodiments the DNA encoding kinase is expressed to enrich endosperm, embryo or seed development. In other preferred embodiments the attenuation results in a plant without seeds (e.g. a fruit or vegetable that is seedless).
The invention also includes a method of increasing the productivity or yield of a transgenic crop. The method includes: (a), introducing into plant cell a DNA encoding the kinase of a MAPKKK operably coupled to a plant cell promoter to produce transformed plant cells. (b), regenerating a plant from transformed cells wherein the MAPKKK kinase is expressed in cells of the plant.
In related aspects, the invention includes a transgenic plant (or plant cells, plant tissues, organs, or plant components) that contains a recombinant kinase-domain of a MAPKKK. The transgene expressed in the plant is controlled by a promoter which is functional in the plant cell. In preferred embodiments the transgene contains a kinase from a plant. In other preferred embodiments the invention includes a kinase from a fungus, such as a yeast, or animal, such as a mammal. In other preferred embodiments the transgene is essentially the kinase.
In related aspects of the invention, seeds and cells of a plant contain a transgene recombinant capable of expressing a MAPKKK kinase.
In other aspects of the invention, a vector (e.g. an expression vector) includes a promoter that is functional in plant cells and operably linked to the gene encoding the MAPKKK polypeptide, as well as a cell, such a plant or prokaryotic, like Agrobacterium, which contains the vector. In preferred embodiments the gene encodes either a polypeptide essentially consisting of a MAPKKK kinase (e.g. a MAPKKK kinase from a plant MAPKKK, such as NPK1 and an ANP), or a genetically modified chimeric polypeptide including such a domain.
In general the kinase-domain used in the methods of the invention (e.g. transgenic plants, plants bred from a transgenic one) is expressed either by itself or as a MAPKKK or kinase-domain-containing polypeptide, or as part a genetically modified chimeric polypeptide. Some useful kinase regions include those capable of activating genes involved in stress responses, repressing the early auxin gene or altering seed growth. Examples of kinase-domains that can be used as exemplars include those that are substantially the same as the kinase-domains of NPK1 (e.g. ANP1, ANP2, ANP3, or AtMEKK1). The invention’s methods and plants preferably use the kinase of NPK1 (or ANP1). Other preferred embodiments use a MAPKKK polypeptide, or a fragment containing the kinase-domain-containing fragment of it that is substantially identical with any one of NPK1, ANP1, ANP2, ANP3, or ANP3.
The DNA that encodes the kinase is constitutively expressed in most cases.” If desired, however, the kinase is expressed by induction, or in a tissue-specific or organ-specific way. The kinase can be expressed in cycling conditions, such as the cell cycle, or under circadian conditions.
Example plants that are useful for the invention’s methods, and also to generate transgenic plants, including plant cells, tissue, organs or tissues of plants, include dicots (such as sugar cane), monocots (such as rice, maize, corn, sugar beet), mustard, potato, soybean, sorghum, cassava, banana, grape, rye, millet, coconut, orange, rye, cabbage, apple, eggplant
By ?polypeptide? “By?polypeptide?
By ?substantially identical? “By?substantially identical? 11-12, 13,-14-15-16; SEQID NOS: 7-22). The length of the comparison sequences for polypeptides is generally at least 16 amino acid, preferably 20 amino acid, more preferably 25 amino acids and most preferably greater than 35 amino acids. Nucleic acids will have a length of at least 50 nucleotides. This is preferably 60 nucleotides. More preferably 75 nucleotides and more preferably 110 nucleotides.
Sequence identities are typically determined using sequence analysis software, such as the Sequence Analysis Software Package of Genetics Computer Group at University of Wisconsin Biotechnology Center (1710 University Avenue Madison, Wis., 53705, BLAST or FastA programs, PILEUP/PRETTYBOX, etc.). These software programs match identical or similar sequences based on the degree of homology assigned to deletions, substitutions and/or modifications. “Conservative substitutions are usually made within the following groups of amino acids: valine, isoleucine and leucine.
By ?obtained from? “By?obtained from?
By ?recombinant? The term’recombinant’ is used to describe a nucleic acids (e.g. DNA) which is free from the genes that flank the gene in the genome of the organism which is the source of the nucleic acids of the invention. This term includes, for instance, a fragment of a gene that is incorporated in a vector, into a virus or plasmid that can replicate autonomously, or into genomic DNA of an eukaryote. It also includes a fragment of a genome or cDNA that exists independently of other sequences (for example, as a cDNA, genomic or cDNA segment produced by restriction endonuclease digestion or PCR). It also includes nucleic acids that are part of a gene hybrid encoding an additional polypeptide.
By ?transformed cell? “By?transformed cell?
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