Method of identifying an agent that modulates the activity...

Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...

Reexamination Certificate

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C435S007200, C435S007210

Reexamination Certificate

active

06780601

ABSTRACT:

BACKGROUND OF THE INVENTION
Glutamate-gated chloride channels are a family of ligand-gated chloride channels unique to invertebrates. Glutamate-gated chloride channels have been cloned from
Caenorhabditis elegans
(Cully et al. (1994) Nature 20:371; U.S. Pat. No. 5,527,703),
Drosophila melanogaster
(Cully et al. (1996) J. Biol. Chem. 271:20187 and U.S. Pat. No. 5,693,492),
Haemonchus contortus
(Delany et al. (1998) Mol. Biochem. Parasit. 97:177),
Lucilia cuprina
(GenBank Accession No. AAC31949) and
Schistocerca americana
(Cohen et al. (1999) 29th Annual Neuroscience Meeting, p. 199). The clones isolated from
C. elegans, D. melanogaster
and
S. americana
have been functionally expressed in Xenopus oocytes, and shown to be activated by glutamate and avermectin. (Arena et al. (1991) Molecular Pharm. 40:368; Arena et al. (1992) Molecular Brain Research 15:339; U.S. Pat. No. 5,693,492; U.S. Pat. No. 5,527,703; Cohen et al., supra).
Because glutamate-gated chloride channels are specific to invertebrates, the channels provide a target for insecticides. In particular, the glutamate-gated chloride channels are the target of the avermectin class of insecticides. Avermectins are naturally occurring and synthetic macrocylic lactones that are widely used in the treatment of parasites and insects.
Insects of the order lepidoptera are significant pests, and in particular the larvae are destructive defoliaters. Further, lepidopteran pests are typically harder to control than diptera. Accordingly, there is a need to identify and develop safe and specific insecticides against lepidopteran pests. The present invention addresses this need by providing isolated nucleic acids encoding a lepidopteran glutamate-gated chloride channel, recombinant lepidopteran glutamate-gated chloride channels, and a method of identifying agents that modulate the activity of the channel.
SUMMARY OF THE INVENTION
The present invention is directed to an isolated nucleic acid encoding a lepidopteran glutamate-gated chloride channel. In a preferred embodiment the nucleic acid is isolatable from
Heliothis virescens
. In another preferred embodiment the nucleic acid comprises a sequence encoding the amino acid sequence of SEQ ID NO. 14.
The present invention further comprises vectors comprising a nucleic acid encoding a lepidopteran glutamate-gated chloride channel, and host cells comprising the vectors.
Another aspect of the present invention provides a recombinant lepidopteran glutamate-gated chloride channel, and kits and compositions comprising a recombinant lepidopteran glutamate-gated chloride channel. A method for preparing a lepidopteran glutamate-gated chloride channel is also provided.
In yet another embodiment, the present invention provides a Xenopus oocyte comprising a nucleic acid encoding a lepidopteran glutamate-gated chloride channel, and a Xenopus occyte expressing a functional lepidopteran glutamate-gated chloride channel.
The present invention further provides a method of identifying agents that modulate the activity of the lepidopteran glutamate-gated chloride channel, and agents identified by the method.


REFERENCES:
patent: 5527703 (1996-06-01), Cully et al.
patent: 5693492 (1997-12-01), Cully et al.
patent: 9849185 (1998-05-01), None
patent: 9907828 (1999-02-01), None
Arena et al. (1991) “Avermectin-Sensitive Chloride Currents Induced byCaenorhabditis ElegansRNA in Xenopus Oocytes,” Molecular Pharm. 40:368.
Arena et al. (1992) “Expression of a Glutamate-Activated Chloride Current inXenopusOocytes Injected withCaenorhabditis ElegansRNA: Evidence for Modulation by Avermectin,” Molecular-Brain Research 15:339.
Cully et al. (1994) “Cloning of an Avermectin-Sensitive Glutamate-Gated Cholride Channel fromCaenorhabditis Elegans,” Nature 371:707.
Cully et al. (1996) “Identification of aDrosophila MelanogasterGlutamate-Gated Chloride Channel Sensitive to the Antiparasitic Agent Avermectin,” J. Biol. Chem. 271:20187.
Delany et al. (1998) “Cloning and Localisation of an Avermectin Receptor-Related Subunit fromHaemonchus Contortus,” Mol. Biochem. Parasit., 97:177.
Mikayama et al. (1993) “molecular Cloning and Functional Expression of a cDNA Encoding Glycosylation-Inhibitin Factor,” Proc. Natl. Acad. Sci. USA 90:10056.
Voet et al. (1990) Biochemistry, John Wiley & Sons, Inc., 126-128 and 228-234.
Smith MH, Warren VA, Thomas BS, Brochu RM, Ertel EA, Rohrer S, Schaeffer J, Schmatz D, Petuch BR, Tang YS, Meinke PT, Kaczorowski GJ, Cohen CJ. Nodulisporic acid opens insect glutamate-gated chloride channels: identification of a new high affinity modulator. Biochemistry 2000;39:5543-5554.
Pomes A, Kempner E, Rohrer S. Target size analysis of an avermectin binding site fromDrosophila melanogaster.Biochimica et Biophysica Acta 1997;1339:233-238.
Yu SJ, Nguyen SN. Insecticide susceptibility and detoxification enzyme activities in permethrin-selected diamondback moths. Peticide Biochemistry and Physiology 1996;56:69-77.

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