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Identification and characterization of the maize arogenate dehydrogenase gene family.

Holding DR, Meeley RB, Hazebroek J, Selinger D, Gruis F, Jung R, Larkins BA - J. Exp. Bot. (2010)

Bottom Line: In plants, the amino acids tyrosine and phenylalanine are synthesized from arogenate by arogenate dehydrogenase and arogenate dehydratase, respectively, with the relative flux to each being tightly controlled.A Mutator insertion at an equivalent position in AroDH-3, the most closely related family member to AroDH-1, is also associated with opaque endosperm and stunted vegetative growth phenotypes.Overlapping but differential expression patterns as well as subtle mutant effects on the accumulation of tyrosine and phenylalanine in endosperm, embryo, and leaf tissues suggest that the functional redundancy of this gene family provides metabolic plasticity for the synthesis of these important amino acids. mto140/arodh-1 seeds shows a general reduction in zein storage protein accumulation and an elevated lysine phenotype typical of other opaque endosperm mutants, but it is distinct because it does not result from quantitative or qualitative defects in the accumulation of specific zeins but rather from a disruption in amino acid biosynthesis.

View Article: PubMed Central - PubMed

Affiliation: Center for Plant Science Innovation, University of Nebraska, 1901 Vine St., Lincoln, NE 68588, USA. dholding2@unl.edu

ABSTRACT
In plants, the amino acids tyrosine and phenylalanine are synthesized from arogenate by arogenate dehydrogenase and arogenate dehydratase, respectively, with the relative flux to each being tightly controlled. Here the characterization of a maize opaque endosperm mutant (mto140), which also shows retarded vegetative growth, is described The opaque phenotype co-segregates with a Mutator transposon insertion in an arogenate dehydrogenase gene (zmAroDH-1) and this led to the characterization of the four-member family of maize arogenate dehydrogenase genes (zmAroDH-1-zmAroDH-4) which share highly similar sequences. A Mutator insertion at an equivalent position in AroDH-3, the most closely related family member to AroDH-1, is also associated with opaque endosperm and stunted vegetative growth phenotypes. Overlapping but differential expression patterns as well as subtle mutant effects on the accumulation of tyrosine and phenylalanine in endosperm, embryo, and leaf tissues suggest that the functional redundancy of this gene family provides metabolic plasticity for the synthesis of these important amino acids. mto140/arodh-1 seeds shows a general reduction in zein storage protein accumulation and an elevated lysine phenotype typical of other opaque endosperm mutants, but it is distinct because it does not result from quantitative or qualitative defects in the accumulation of specific zeins but rather from a disruption in amino acid biosynthesis.

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Phylogenic analysis of maize and other plant AroDH genes. Genus-species codes were prepended to the accessions and names: At, Arabidopsis thaliana; Cr, Chlamydomonas reinhardtii; Os, Oryza sativa; Ol, Ostreococcus lucimarinus; Pp, Physcomitrella patens; Ps, Picea sitchensis; Sb, Sorghum bicolor; Vv, Vitis vinifera; and Zm, Zea mays. Coloured backgrounds denote taxonomic groups as follows: light green designates non-vascular plants (moss and algae); light yellow designates vascular plants (gymnosperms and angiosperms); blue-green designates algae (Chlamydomonas and Ostreococcus); peach designates gymnosperms (Picea); and gold designates monocots (Orzya, Sorghum, and Zea).
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fig4: Phylogenic analysis of maize and other plant AroDH genes. Genus-species codes were prepended to the accessions and names: At, Arabidopsis thaliana; Cr, Chlamydomonas reinhardtii; Os, Oryza sativa; Ol, Ostreococcus lucimarinus; Pp, Physcomitrella patens; Ps, Picea sitchensis; Sb, Sorghum bicolor; Vv, Vitis vinifera; and Zm, Zea mays. Coloured backgrounds denote taxonomic groups as follows: light green designates non-vascular plants (moss and algae); light yellow designates vascular plants (gymnosperms and angiosperms); blue-green designates algae (Chlamydomonas and Ostreococcus); peach designates gymnosperms (Picea); and gold designates monocots (Orzya, Sorghum, and Zea).

Mentions: A My iQ Real-time PCR thermocycler (Bio-Rad) was used with the following program: 95 °C for 5 min, followed by 45 cycles of 95 °C (10 s), and 60 °C (10 s) with 20 C s−1 ramp rates. Melting curves were obtained by heating from 65 °C to 95 °C with a 0.1 C s−1 ramp rate. Expression levels of AroDH-2 and -3 genes were calculated relative to the reference (AroDH-1) for each tissue type (Fig. 4B). Tissue-specific expression levels for each gene were calculated relative to the reference (endosperm expression level) for each gene (Fig. 4C). The internal control gene was glyceraldehyde phosphate dehydrogenase (GAPDH) which showed minimal variation between samples. For each expression calculation, the average crossing temperature (Ct) value was determined for three biological replicate samples of arodh-1 and the wild type for each tissue type. Relative expression was calculated using the following equation, where X=subject gene, C=control gene (GAPDH), W=average Ct of three reference samples, and G=average Ct of three subject samples: 2[(WX–WC)–(GX–GC)]. The final fold change values shown are the mean of three independent experiments (±SD).


Identification and characterization of the maize arogenate dehydrogenase gene family.

Holding DR, Meeley RB, Hazebroek J, Selinger D, Gruis F, Jung R, Larkins BA - J. Exp. Bot. (2010)

Phylogenic analysis of maize and other plant AroDH genes. Genus-species codes were prepended to the accessions and names: At, Arabidopsis thaliana; Cr, Chlamydomonas reinhardtii; Os, Oryza sativa; Ol, Ostreococcus lucimarinus; Pp, Physcomitrella patens; Ps, Picea sitchensis; Sb, Sorghum bicolor; Vv, Vitis vinifera; and Zm, Zea mays. Coloured backgrounds denote taxonomic groups as follows: light green designates non-vascular plants (moss and algae); light yellow designates vascular plants (gymnosperms and angiosperms); blue-green designates algae (Chlamydomonas and Ostreococcus); peach designates gymnosperms (Picea); and gold designates monocots (Orzya, Sorghum, and Zea).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2921203&req=5

fig4: Phylogenic analysis of maize and other plant AroDH genes. Genus-species codes were prepended to the accessions and names: At, Arabidopsis thaliana; Cr, Chlamydomonas reinhardtii; Os, Oryza sativa; Ol, Ostreococcus lucimarinus; Pp, Physcomitrella patens; Ps, Picea sitchensis; Sb, Sorghum bicolor; Vv, Vitis vinifera; and Zm, Zea mays. Coloured backgrounds denote taxonomic groups as follows: light green designates non-vascular plants (moss and algae); light yellow designates vascular plants (gymnosperms and angiosperms); blue-green designates algae (Chlamydomonas and Ostreococcus); peach designates gymnosperms (Picea); and gold designates monocots (Orzya, Sorghum, and Zea).
Mentions: A My iQ Real-time PCR thermocycler (Bio-Rad) was used with the following program: 95 °C for 5 min, followed by 45 cycles of 95 °C (10 s), and 60 °C (10 s) with 20 C s−1 ramp rates. Melting curves were obtained by heating from 65 °C to 95 °C with a 0.1 C s−1 ramp rate. Expression levels of AroDH-2 and -3 genes were calculated relative to the reference (AroDH-1) for each tissue type (Fig. 4B). Tissue-specific expression levels for each gene were calculated relative to the reference (endosperm expression level) for each gene (Fig. 4C). The internal control gene was glyceraldehyde phosphate dehydrogenase (GAPDH) which showed minimal variation between samples. For each expression calculation, the average crossing temperature (Ct) value was determined for three biological replicate samples of arodh-1 and the wild type for each tissue type. Relative expression was calculated using the following equation, where X=subject gene, C=control gene (GAPDH), W=average Ct of three reference samples, and G=average Ct of three subject samples: 2[(WX–WC)–(GX–GC)]. The final fold change values shown are the mean of three independent experiments (±SD).

Bottom Line: In plants, the amino acids tyrosine and phenylalanine are synthesized from arogenate by arogenate dehydrogenase and arogenate dehydratase, respectively, with the relative flux to each being tightly controlled.A Mutator insertion at an equivalent position in AroDH-3, the most closely related family member to AroDH-1, is also associated with opaque endosperm and stunted vegetative growth phenotypes.Overlapping but differential expression patterns as well as subtle mutant effects on the accumulation of tyrosine and phenylalanine in endosperm, embryo, and leaf tissues suggest that the functional redundancy of this gene family provides metabolic plasticity for the synthesis of these important amino acids. mto140/arodh-1 seeds shows a general reduction in zein storage protein accumulation and an elevated lysine phenotype typical of other opaque endosperm mutants, but it is distinct because it does not result from quantitative or qualitative defects in the accumulation of specific zeins but rather from a disruption in amino acid biosynthesis.

View Article: PubMed Central - PubMed

Affiliation: Center for Plant Science Innovation, University of Nebraska, 1901 Vine St., Lincoln, NE 68588, USA. dholding2@unl.edu

ABSTRACT
In plants, the amino acids tyrosine and phenylalanine are synthesized from arogenate by arogenate dehydrogenase and arogenate dehydratase, respectively, with the relative flux to each being tightly controlled. Here the characterization of a maize opaque endosperm mutant (mto140), which also shows retarded vegetative growth, is described The opaque phenotype co-segregates with a Mutator transposon insertion in an arogenate dehydrogenase gene (zmAroDH-1) and this led to the characterization of the four-member family of maize arogenate dehydrogenase genes (zmAroDH-1-zmAroDH-4) which share highly similar sequences. A Mutator insertion at an equivalent position in AroDH-3, the most closely related family member to AroDH-1, is also associated with opaque endosperm and stunted vegetative growth phenotypes. Overlapping but differential expression patterns as well as subtle mutant effects on the accumulation of tyrosine and phenylalanine in endosperm, embryo, and leaf tissues suggest that the functional redundancy of this gene family provides metabolic plasticity for the synthesis of these important amino acids. mto140/arodh-1 seeds shows a general reduction in zein storage protein accumulation and an elevated lysine phenotype typical of other opaque endosperm mutants, but it is distinct because it does not result from quantitative or qualitative defects in the accumulation of specific zeins but rather from a disruption in amino acid biosynthesis.

Show MeSH
Related in: MedlinePlus