Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving transferase
Reexamination Certificate
2000-02-14
2001-10-09
Nashed, Nashaat T. (Department: 1652)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving transferase
C435S193000
Reexamination Certificate
active
06300091
ABSTRACT:
FIELD OF THE INVENTION
The invention relates to genes isolated from
Arabidopsis thaliana
that encode proteins essential for plant growth and development. The invention also includes the methods of using these proteins as herbicide targets, based on the essentiality of these genes for normal growth and development. The invention is also useful as a screening assay to identify inhibitors that are potential herbicides. The invention may also be applied to the development of herbicide tolerant plants, plant tissues, plant seeds, and plant cells.
BACKGROUND OF THE INVENTION
The use of herbicides to control undesirable vegetation such as weeds in crop fields has become almost a universal practice. The herbicide market exceeds 15 billion dollars annually. Despite this extensive use, weed control remains a significant and costly problem for farmers.
Effective use of herbicides requires sound management. For instance, the time and method of application and stage of weed plant development are critical to getting good weed control with herbicides. Since various weed species are resistant to herbicides, the production of effective new herbicides becomes increasingly important. Novel herbicides can now be discovered using high-throughput screens that implement recombinant DNA technology. Metabolic enzymes found to be essential to plant growth and development can be recombinantly produced through standard molecular biological techniques and utilized as herbicide targets in screens for novel inhibitors of the enzyme activity. The novel inhibitors discovered through such screens may then be used as herbicides to control undesirable vegetation.
Herbicides that exhibit greater potency, broader weed spectrum, and more rapid degradation in soil can also, unfortunately, have greater crop phytotoxicity. One solution applied to this problem has been to develop crops that are resistant or tolerant to herbicides. Crop hybrids or varieties tolerant to the herbicides allow for the use of the herbicides to kill weeds without attendant risk of damage to the crop. Development of tolerance can allow application of a herbicide to a crop where its use was previously precluded or limited (e.g to pre-emergence use) due to sensitivity of the crop to the herbicide. For example, U.S. Pat. No. 4,761,373 to Anderson et al. is directed to plants resistant to various imidazolinone or sulfonamide herbicides. The resistance is conferred by an altered acetohydroxyacid synthase (AHAS) enzyme. U.S. Pat. No. 4,975,374 to Goodman et al. relates to plant cells and plants containing a gene encoding a mutant glutamine synthetase (GS) resistant to inhibition by herbicides that were known to inhibit GS, e.g. phosphinothricin and methionine sulfoximine. U.S. Pat. No. 5,013,659 to Bedbrook et al. is directed to plants expressing a mutant acetolactate synthase that renders the plants resistant to inhibition by sulfonylurea herbicides. U.S. Pat. No. 5,162,602 to Somers et al. discloses plants tolerant to inhibition by cyclohexanedione and aryloxyphenoxypropanoic acid herbicides. The tolerance is conferred by an altered acetyl coenzyme A carboxylase (ACCase).
Notwithstanding the above-described advancements, there remains a persistent and ongoing problem with unwanted or detrimental vegetation growth (e.g. weeds). Furthermore, as the population continues to grow, there will be increasing food shortages. Therefore, there exists a long felt, yet unfulfilled need, to find new, effective, and economic herbicides.
SUMMARY OF THE INVENTION
It is an object of the invention to provide an effective and beneficial method to identify novel herbicides. A feature of the invention is the identification of a gene in
A. thaliana
, herein referred to as the 8388 gene, which shows sequence similarity to DEAD box RNA helicase (Luking et al. (1998) Critical Reviews in Biochemistry and Molecular Biology, 33(4): 259-296). A feature of the invention is the identification of a gene in
A. thaliana
, herein referred to as the 18048 gene, which shows sequence similarity to ADP-ribosylation factor (Art) genes (Regad et al. (1993) FEBS Lett. 25: 133-136; Bar-Peled et al. (1995) The Plant Cell, 7: 667-676). A feature of the invention is the identification of a gene in
A. thaliana
, herein referred to as the 16713 gene, which shows sequence similarity to acetoacetyl coA thiolases (Vollack and Bach (1996) Plant Physiol. 111: 1097-1107; Hiser et al. (1994) J. Biol. Chem. 269: 31383-31389; Fukao et al. (1990) J. Clin. Invest. 86: 2086-2092; Fukao et al. (1989) J. Biochem. 106: 197-204; Wilson et al. (1994) Nature 368: 32-38). A feature of the invention is the identification of a gene in Arabidopsis, herein referred to as the 4144 gene, which encodes a protein with sequence similarity to chloroplast ATP synthase delta chain (Hermans et al. (1988) Plant Mol. Biol. 10: 323-330; Hoesche and Berzborn (1992) Biochimica et Biophysica Acta, 1171: 201-204; Hoesche and Berzbom (1993) Biochimica et Biophysica Acta, 1142: 293-305; Napier et al. (1992) Plant Mol. Biol. 20: 549-554). Another feature of the invention is the discovery that the 8388, 18048, 16713, and 4144 genes are essential for normal growth and development. An advantage of the present invention is that the newly discovered essential genes provide the basis for identity of a novel herbicidal mode of action which enables one skilled in the art to easily and rapidly discover novel inhibitors of gene function useful as herbicides.
One object of the present invention is to provide essential genes in plants for assay development for inhibitory compounds with herbicidal activity. Genetic results show that when any one of the 8388, 18048, 16713, or 4144 genes is mutated in
Arabidopsis thaliana
, the resulting phenotype is lethal in the homozygous state. This suggests a critical role for the gene products encoded by the 8388, 18048, 16713, and 4144 genes.
Using T-DNA insertion mutagenesis, the inventors of the present invention have demonstrated that the activity of any one of the 8388, 18048, 16713, or 4144 gene products is essential for
A. thaliana
growth. This implies that chemicals, which inhibit the function of the 8388, 18048, 16713, or 4144-encoded protein in plants, are likely to have detrimental effects on plants and are potentially good herbicide candidates. The present invention therefore provides methods of using a purified protein encoded by the 8388, 18048, 16713, or 4144 gene sequence described below to identify inhibitors thereof, which can then be used as herbicides to suppress the growth of undesirable vegetation, e.g. in fields where crops are grown, particularly agronomically important crops such as maize and other cereal crops such as wheat, oats, rye, sorghum, rice, barley, millet, turf and forage grasses, and the like, as well as cotton, sugar cane, sugar beet, oilseed rape, and soybeans.
The present invention discloses novel nucleotide sequences derived from
A. thaliana
, designated the 8388, 18048, 16713, or 4144 genes. The nucleotide sequences of the coding regions for the cDNA clones are set forth in SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:7, and SEQ ID NO:21, respectively, and the corresponding amino acid sequences of the 8388, 18048, 16713, or 4144-encoded protein are set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:8, and SEQ ID NO:22, respectively. The present invention also includes nucleotide sequences substantially similar to those set forth in SEQ ID NO:1, SEQ ID NO:5, SEQ ID NO:7, and SEQ ID NO:21, respectively. The present invention also encompasses plant proteins whose amino acid sequence are substantially similar to the amino acid sequences set forth in SEQ ID NO:2, SEQ ID NO:6, SEQ ID NO:8, and SEQ ID NO:22, respectively. The present invention also includes methods of using the 8388, 18048, 16713, or 4144 gene products as herbicide targets, based on the essentiality of these genes for normal growth and development. Furthermore, the invention can be used in a screening assay to identify inhibitors of 8388, 18048, 16713, or 4144 gene function that are potential herbicides.
In a pr
Ashby Carl S.
McElver John A.
Patton David A.
Fronda Christian L.
Lebel Edouard G.
Meigs J. Timothy
Nashed Nashaat T.
Stults Larry W.
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