Glutamate receptors in nutrient sensing, metabolism, growth and development

Abstract

The present invention relates to methods that may be used to improve or modify nutrient sensing, absorption, metabolism, root growth, stomatal conductance, N use efficiency, C and N metabolism, plant biomass production and seed yield. More specifically, this invention is related to the glutamate receptors (GLRs) and their role(s) in nutrient sensing, metabolism, regulation of growth, development, and yield.

We claim: 
     
         1 . Glutamate receptors of the sequences SEQ ID:1 and/or SEQ ID: 2, wherein they regulate amino acid sensing, metabolism, growth and development. 
     
     
         2 . The method of  claim 1  wherein the glutamate receptor gene product is at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% similar to SEQ. ID:1 and/or SEQ ID:2. 
     
     
         3 . The method of  claim 1  wherein the activity of the protein is modulated by chemicals such as agonists, antagonists or modulators or interacting proteins. 
     
     
         4 . A method for developing plant or cell line or clone that are efficient in N use, metabolism, growth and yield, comprising: obtaining and/or selecting a plant or cell line or clone that have modified levels or efficiency of glutamate receptor(s) compared to a progenitor plant. 
     
     
         5 . The method of  claim 4  wherein the plant or cell line or clone demonstrates improved N sensing responses. 
     
     
         6 . The method of  claim 4  wherein the glutamate receptor gene product is at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ. ID: 1. 
     
     
         7 . The method of  claim 4  wherein the glutamate receptor gene product is at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ. ID: 2. 
     
     
         8 . The method of  claim 4  wherein the glutamate receptor gene product is AtGLR1.1 polypeptide (SEQ. ID: 1) 
     
     
         9 . The method of  claim 4  wherein the glutamate receptor gene product is AtGLR3.2 polypeptide (SEQ. ID: 2) 
     
     
         10 . The method of  claim 4 , comprising introducing into a plant or cell line or clone an expression cassette, the expression cassette comprising a promoter operably linked to the polynucleotide, or its complement, encoding glutamate receptor group gene product. 
     
     
         11 . The method of  claim 4 , comprising introducing into a plant or cell line or clone an expression cassette, the expression cassette comprising a promoter operably linked to the polynucleotide, or its complement, encoding glutamate receptor group gene product at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ. ID. 1. 
     
     
         12 . The method of  claim 4 , comprising introducing into a plant or cell line or clone an expression cassette, the expression cassette comprising a promoter operably linked to the polynucleotide, or its complement, encoding glutamate receptor group gene product at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ. ID. 2. 
     
     
         13 . The method of  claim 4 , comprising introducing into a plant or cell line or clone an expression cassette, the expression cassette comprising a promoter operably linked to the polynucleotide, or its complement, encoding glutamate receptor group gene product, wherein the gene product is AtGLR1.1 (SEQ ID 1). 
     
     
         14 . The method of  claim 4 , comprising introducing into a plant or cell line or clone an expression cassette, the expression cassette comprising a promoter operably linked to the polynucleotide, or its complement, encoding glutamate receptor group gene product, wherein the gene product is AtGLR3.2 (SEQ ID 2). 
     
     
         15 . The method of  claim 10 , wherein the expression cassette is an antisense expression cassette, the antisense expression cassette comprising a promoter operably linked to the complement of the polynucleotide encoding a polypeptide identical to SEQ ID NO:1 or SEQ ID NO:2. 
     
     
         16 . The method of  claim 10 , wherein the expression cassette is an antisense expression cassette, the antisense expression cassette comprising a promoter operably linked to the complement of the polynucleotide encoding a polypeptide at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ ID NO:1 or SEQ ID NO:2. 
     
     
         17 . The method of  claim 10 , wherein the promoter is tissue-specific. 
     
     
         18 . The method of  claim 10 , wherein the promoter is inducible. 
     
     
         19 . The method of  claim 10 , wherein the plant is homozygous for a mutation in a polynucleotide encoding a polypeptide at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ ID NO:1. 
     
     
         20 . The method of  claim 10 , wherein the plant is homozygous for a mutation in a polynucleotide encoding a polypeptide at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ ID NO:2. 
     
     
         21 . The method of  claim 10 , wherein the modification of glutamate receptors may also include knock-outs, activation tags, RNA interference or ribozyme mediated inhibition or down regulation of the gene(s), promoter(s) or regulatory elements including transcription factors and regulatory small RNA molecules. 
     
     
         22 . The method of  claim 1 , wherein identification of natural allelic variations of SEQ. ID 1 and SEQ ID 2, including SNPs 
     
     
         23 . The method  claim 22  wherein the natural allelic variation selection and transfer by conventional breeding methods using markers such as marker assisted selection. 
     
     
         24 . The method of  claim 22 , wherein identification of natural allelic variations of polynucleotide encoding a polypeptide at least 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% or 99% identical to SEQ ID NO:1 or SEQ ID NO:2, including SNPs 
     
     
         25 . The method of  claim 24 , wherein identification of natural allelic variation, selection and transfer by conventional breeding methods using markers such as marker assisted selection. 
     
     
         26 . The method of  claim 22 , wherein the markers may be morphological, physical, biochemical or DNA based marker such as but not limited to PCR or RFLP or AFLP or SSR such as microsatellite based markers which locate glutamate receptors. 
     
     
         27 . The method of  claim 1  and/or  4  wherein, the activity is modified using chemicals such as agonist/antagonists or modulators of ion channel activities. 
     
     
         28 . The method of  claim 1  and/or  4  wherein, the activity is modified using proteins that interact with glutamate receptors. 
     
     
         29 . The method of  claim 4 , wherein the plant exhibits modified levels of N assimilation/metabolism enzymes such as GS, AAT, and Fd-GOGAT. 
     
     
         30 . The method of  claim 4 , wherein the plant exhibits one or more traits selected from: i) faster rate of growth, ii) greater fresh or dry weight at maturation, iii) greater fruit or seed yield, iv) greater total plant N content, v) greater fruit or seed N content, vi) greater free amino acid content in the whole plant, vii) greater free amino acid content in the fruit or seed, viii) greater protein content in seed or fruit, ix) greater protein content in a vegetative tissue, x) greater amount of products including one or more of oils, polymers, enzymes, antibodies, or metabolites, or xi) greater ability to resist disease or pest damage, than a progenitor plant which does not contain the gene construct. 
     
     
         31 . The method of  claim 4 , wherein the plant is modified in specific C and N metabolic pathways to a particular product. 
     
     
         32 . The method of  claim 4 , wherein the plant is efficient in nutrient uptake wherein nutrient uptake is improved compared to the progenitor plant. 
     
     
         33 . The method of  claim 4 , wherein the plant is efficient in water use, 
     
     
         34 . The method of  claim 4 , wherein water efficiency comprises altered stomatal conductance and reduced transpirational loss. 
     
     
         35 . A transgenic plant having a gene construct comprising a gene encoding a glutamate receptor operably linked to a plant promoter so that the glutamate receptor is overexpressed in the transgenic plant, and the transgenic plant exhibits:
 i) faster rate of growth;   ii) greater fresh or dry weight at maturation;   ii) greater fresh or dry weight at maturation;   iii) greater fruit or seed yield;   iv) greater total plant nitrogen content;   v) greater fruit or seed nitrogen content;   vi) greater free amino acid content in the whole plant;   vii) greater free amino acid content in the fruit or seed;   viii) greater protein content in seed or fruit; or   ix) greater protein content in a vegetative tissue;   than a progenitor plant which does not contain the gene construct, when the transgenic plant and the progenitor plant are cultivated under identical growth conditions, wherein the glutamate receptor gene product is substantially identical to AtGLR3.2 polypeptide (SEQ. ID: 2).   
     
     
         36 . The transgenic plant of  claim 35 , wherein the plant promoter is a strong, constitutively expressed plant promoter. 
     
     
         37 . The transgenic plant of  claim 35 , wherein the glutamate receptor gene product is at least 99% identical to AtGLR3.2 polypeptide (SEQ. ID: 2). 
     
     
         38 . The transgenic plant of  claim 35 , wherein the transgenic plant exhibits at least about 60% increase in total lateral root elongation. 
     
     
         39 . The transgenic plant of  claim 35 , wherein the plant exhibits at least about 21% increase in primary root growth. 
     
     
         40 . A seed of the transgenic plant of  claim 35 , wherein the seed has the gene construct. 
     
     
         41 . A progeny, clone, cell line or cell of the transgenic plant of  claim 35  wherein said progeny, clone, cell line or cell has the gene construct. 
     
     
         42 . An isolated nucleic acid comprising the mRNA of SEQ ID 2. 
     
     
         43 . An isolated polypeptide comprising the polypeptide of SEQ ID 2.