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<h3><font>Research Interests in Maeda Lab</font></h3>
 
<h3><font>Research Interests in Maeda Lab</font></h3>
  
−
Plants produce a diverse array of secondary (specialized) metabolites in order to survive in various ecological niches. In contrast to the well-documented diversification of plant secondary metabolism, primary metabolism, which provides precursors of secondary metabolites, is generally thought to be conserved across different plant species. However, '''it is not understood to what extent primary metabolic pathways of plants have evolved in different lineages to support the downstream synthesis of diverse secondary metabolites'''. Such basic knowledge of the interface between plant primary and secondary metabolism will be also crucial for future metabolic engineering and breeding to improve production of plant natural products. To address this question, we are investigating in different plant species <font face=arial color=orangered> '''the biosynthetic pathways of aromatic amino acids'''</font>.
+
'''Our goal is to understand remarkable chemical and metabolic diversity that evolved in the plant kingdom, in order to redesign plants for efficient and sustainable production of natural and bio-products'''. Plants produce a diverse array of secondary (specialized) metabolites, which play critical roles in plant adaption to various ecological niches. In contrast to well-documented diversification of plant secondary metabolism, primary metabolism, which provides precursors of secondary metabolites, is generally thought to be conserved across different plant species. However, our lab has recently uncovered <font face=arial color=blue>'''lineage-specific diversifications of primary metabolic pathways'''</font> in different plant groups, likely to support downstream synthesis of diverse secondary metabolites (Schenck et al. 2015; 2017a; Lopez-Nieves et al. 2018; Maeda 2019a). Such basic knowledge of the interface between plant primary and secondary metabolism will be crucial for future breeding, metabolic engineering, and synthetic biology to improve production of natural and bio-products in plant hosts (Maeda 2019b).  
  
 +
We are currently conducting various projects that focus on <font face=arial color=blue>'''aromatic amino acid biosynthesis'''</font>, which connects central carbon metabolism and diverse plant natural product pathways. Currently, we are further '''i)''' exploring other examples of primary metabolic diversity in the aromatic amino acid pathways, '''ii)''' investigating underlying genetic and biochemical mechanisms, '''iii)''' utilizing the knowledge to engineer aromatic amino acid biosynthesis and improve precursor metabolite availability, and '''iv)''' analyzing physiological impacts of metabolic engineering on plant growth and development.
  
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'''Tyrosine Biosynthetic Pathways in Plants''' [[Image:Tyrpathway.png|right|400px|]] 
 
−
*Tyrosine (Tyr) is an aromatic amino acid required for protein biosynthesis in all living cells and, due to the absence of Tyr biosynthesis in animals, is an essential nutrient in human diets. In plants, Tyr also serves as a precursor of numerous natural products, which include tocopherols (vitamin E), cyanogenic glycosides, and isoquinoline alkaloids (e.g., analgesic morphine and codeine). These Tyr-derived plant metabolites have a remarkable structural complexity and a variety of pharmacological and biological activities, making them effective nutritional compounds and pharmaceutical drugs. However, often the low yields of these compounds in plant tissues hamper their commercial production in plants, and there is a growing need to rationally engineer the plant Tyr pathway.
 
  
−
*Although the Tyr biosynthetic pathway has been extensively studied in microbes (e.g., ''E. coli'', yeast), '''we still do not know how and where Tyr is synthesized in plants'''. To investigate Tyr biosynthetic pathways, their localization and regulation in different plant species, we use two contrasting model plants, ''Arabidopsis thaliana'' and ''Medicago truncatula'', which appear to use different pathways for Tyr synthesis (see our paper, [[Maeda_Lab:Publications |'''Schenck ''et al''., 2014''']]). We particularly focus on prephenate dehydrogenase (TyrAp/'''PDH''') and arogenate dehydrogenase (TyrAa/'''ADH''') enzymes, which compete prephenate and arogenate substrates, respectively, against phenylalanine biosynthesis. Accurate understanding of the Tyr pathways may provide an opportunity to dramatically increase Tyr availability in plants by redirecting carbon flow from phenylalanine biosynthesis, which consumes major carbon flow for lignin biosynthesis in vascular plants.
+
'''Evolutionary Diversification of Tyrosine Biosynthesis and Downstream Natural Product Pathways'''
 +
*Primary metabolites are indispensable cellular components that directly impact plant growth and development. Consequently, '''''primary metabolic pathways are evolutionarily constrained, typically conserved among the plant kingdom, and difficult to manipulate'''''. Our lab, however, uncovered lineage-specific diversification of a primary amino acid pathway, tyrosine biosynthesis, in legumes (Schenck et al. 2015; 2017a; 2017b; Maeda 2019a). We also revealed that de-regulation of tyrosine biosynthesis occurred at the base of core Caryophyllales and facilitated later evolution of a novel pigment pathway, betalain biosynthesis (Lopez-Nieves et al. 2018).
 +
*Combining evolutionary biochemistry, metabolomics, and phylogenomic analyses, we are further exploring evolutionary alterations of tyrosine biosynthesis and their impacts on associated metabolic network in various plant lineages. The study can advance our understanding of the step-wise evolution of complex metabolic traits at macroevolutionary scale and explore how the evolution of such traits influence subsequent adaptation and diversification in plants. As a part of broader impact activity, we will be also generating '''chemotaxonomic databases''' together with students to study evolutionary history, mechanism, and functions of tremendous chemical diversity that exist in the plant kingdom.
  
 +
''Collaborators: Michael McKain (Univ. Arabama), James Leebens-Mack (Univ. Georgia), Ya Yang (Univ Minnesota), Samuel Brockington (Cambridge), and Stephen Smith (Univ. Michigan)''
  
−
'''Role of Tyrosine Pathway Regulation in the Formation of Tyr-Derived Natural Products'''
+
''Read more at '''our NSF PGRP project website''' (under construction)'':
−
*Towards improving the production of Tyr-derived plant natural products, we also address the question, "'''how does the regulation of the Tyr pathway contribute to the formation of downstream Tyr-derived compounds?'''" In this project, we use table beet (''Beta vulgaris'') as a novel model system, which produces high levels of Tyr-derived pigments, betalains. Using functional genomics approach, we aim to isolate and characterize of ''B. vulgaris'' ADH (or PDH) enzymes involved in Tyr biosynthesis. In collaboration with [http://goldman.horticulture.wisc.edu Dr. Irwin Goldman] at UW Madison, we will investigate biochemical and genetic changes that has been introduced in the Tyr and betalain pathways during breeding of ''B. vulgaris'' germplasm. Finally, we will further engineer the pathway in order to improve the production of Tyr and betalains in table beets as a proof of concept, which can be applied to other plant species for enhanced production of Tyr-derived compounds with nutritional and medicinal values.
 
  
  
−
'''Regulation of the Shikimate Pathway in Plants'''
+
'''Regulation of the Shikimate and Aromatic Amino Acid Pathways in Plants'''
−
*The shikimate pathway provides chorismate, a common precursor of all three aromatic amino acids (phenylalanine, tyrosine, and tryptophan). In microbes, the first enzyme of the shikimate pathway, 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase (DAHPS), is tightly regulated by aromatic amino acids and controls carbon flux through the shikimate pathway. In plants, previous biochemical studies showed that the plant DAHPS enzymes are not sensitive to aromatic amino acids, suggesting that plants have a different mechanism regulating the shikimate pathway. However, the underlying regulatory mechanism of the plant shikimate pathway is poorly understood, due to limited knowledge of the plant enzymes involved in the early steps of the shikimate pathway. We apply integrated approaches of bioinformatics, enzymology, forward/reverse genetics, cell biology, and analytical chemistry, including stable isotope-assisted metabolic flux analyses, to investigate the enzyme organization and regulatory mechanisms of the plant shikimate pathway.
+
*The shikimate pathway directs carbon flux from central carbon metabolism (e.g. Calvin-Benson cycle, glycolysis) to biosynthesis of aromatic amino acids (AAAs) and numerous AAA-derived plant natural products (Maeda and Dudareva, 2012). However, '''''it is unknown how plants regulate the shikimate pathway'''''. We are investigating molecular mechanisms underlying de-regulation of plant AAA biosynthesis and analyzing their quantitative impacts on overall plant metabolic network. Here we are utilizing enzyme biochemistry, forward and reverse genetics, 13CO2 labeling, and structure-function enzyme analyses, as well as metabolic pathway modeling. The obtained enzyme variants and knowledge can be used to boost the production of AAAs and their derived natural products in plants.  
  
 +
''Collaborators: Joseph Jez (WUSTL), Alisdair Fernie (MPI-Golm), Mark Stitt (MPI-Golm), Zoran Nikoloski (Univ. Potsdam)''
  
−
'''Evolution of Plant Phenylalanine Biosynthesis'''
 
−
[[Image:Camilla_image_2.png|right|320px|]] 
 
−
*In plants, phenylalanine (Phe) is a precursor of abundant and diverse phenylpropanoid compounds, such as flavonoids, tannins, and lignin. Plants synthesize Phe predominantly via the arogenate pathway ([[Maeda_Lab:Publications |'''Maeda ''et al''., 2010; 2011''']]) in contrast to model microbes (e.g. ''E. coli'') that use exclusively the phenylpyruvate pathway for Phe biosynthesis. Together with [https://www.ppws.vt.edu/people/faculty/jelesko-john.html Dr. John Jelesko] at Virginia Tech, we conducted phylogenetically-informed biochemical characterization of prephenate aminotransferase (PPA-AT), which catalyzes the first committed step of the arogenate pathway, and uncovered a unique evolutionary history of the arogenate Phe pathway. Our results revealed that prephenate-specific aminotransferase containing Thr84 and Lys169 was transferred from a Chloribi/Bacteroidetes ancestor to a eukaryotic ancestor of the Plantae. This ancient lateral gene transfer likely led to efficient Phe production via the arogenate pathway, which currently supports the production of abundant phenylpropanoid natural products such as lignin. ([[Maeda_Lab:Publications |'''Dornfeld ''et al''., 2014''']])
 
  
 +
'''Crop-Based Production of Natural Products through Synthetic Biology'''
 +
*Plant natural products provide valuable nutraceuticals, pharmaceuticals, and bio-products. Global crop production systems provide sustainable and potentially efficient chemical production platforms, as plants can naturally produce and store abundant chemicals. However, '''''little effort has been made to identify and build plant hosts or chassis for sizable production of natural products'''''.
 +
*We are addressing two outstanding questions thwarting this issue: i) Which plants serve as efficient chemical production platforms and ii) what is the contribution of primary metabolite precursor supply for natural product synthesis? Utilizing our de-regulated tyrosine biosynthetic enzymes (Schenck et al. 2015; 2017; Lopez-Nieves et al. 2018) we are conducting proof-of-concept experiments by expressing tyrosine-utilizing enzymes in transgenic Arabidopsis and other crops accumulating different levels of tyrosine. The study will generate prototype crops for nutrient-enriched functional food, feed, and natural dye, and also lay essential groundwork towards crop-based production of natural and bio-products.
 +
 +
''Collaborator: Wisconsin Crop Innovation Center (WCIC)''
 +
 +
 +
'''Amino Acid Sensing and Response Mechanisms in Plants'''
 +
*The long-term goal of the Maeda laboratory is to improve the production of amino acids and amino acid-derived plant natural products. Perturbation of amino acid metabolism, however, often leads to pleiotropic growth and developmental responses and '''''very little is known how amino acid homeostasis is regulated and linked to plant growth and development'''''. We previously isolated partially tyrosine-deficient ''tyra2'' mutants as well as tyrosine hyper-accumulating lines of Arabidopsis, both of which exhibit compromised growth and unique reticulate leaf phenotypes (de Oliveira et al. 2019). These findings suggest that imbalance of aromatic amino acids impacts plant growth and development.  Here we are conducting physiological, genetic, and biochemical analyses to elucidate mechanisms underlying these pleiotropic plant responses to amino acid imbalance.  Fundamental understanding of amino acid homeostasis mechanisms will enhance plants’ resilience to amino acid perturbation and hence have broad impacts on improving yield and nutritional quality of crops.
 +
 +
 +
'''Defining the Nitrogen Flux Maps (NFMs) of Plants'''
 +
*Nitrogen (N) is a critical element of organic molecules, including aromatic amino acids, but is highly limited in plants. Thus, N use efficiency directly impacts overall yield and performance of plants. Unlike extensively-studied carbon (C) flux map of plant metabolism,'''''little is known how assimilated N flows through the metabolic network, namely “N flux map (NFM)”'''''. The core of NFM is different branches of amino acid metabolism interconnected by aminotransferases (ATs), which play pivotal roles in distributing reduced N for synthesis of various organonitrogen compounds (Dornfeld et al., 2014; Wang et al., 2016; Wang and Maeda, 2017; Wang et al., 2019). Utilizing the wealth of sequenced plant genome information, high-throughput gene and protein syntheses, and enzyme assay platforms, we aim to determine multi-substrate specificities of all ATs from different plant species and construct NFMs from Arabidopsis and Sorghum. The resulting NFMs will provide foundation to further optimize N metabolic network and to generate plants that can produce high levels of N-containing compounds (e.g. alkaloids) while maintaining robust growth even in reduced N fertilizer. 
 +
 +
''Collaborators: Trent Northen (Lawrence Berkeley National Laboratory, LBNL), Yasuo Yoshikuni (LBNL, JGI), Zoran Nikoloski (Max Planck Institute Golm/Univ Potsdam), Taichi Takasuka (Hokkaido Univ)''
  
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<font face=arial color=black size=3> '''If you are interested in joining our lab, please send email to  [mailto:maeda2@wisc.edu maeda2@wisc.edu].''' </font>
+
''Read more at '''our DOE BER project website'''''
  
  
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::::::::'''''Our studies are supported by UW-Madison, NSF IOS, and USDA NIFA AFRI.'''''
+
::::::::'''''Our studies are supported by UW-Madison, NSF, USDA NIFA, and DOE BER.'''''
 
[[Image:UWlogo.png|center|150px|]]
 
[[Image:UWlogo.png|center|150px|]]
 
[[Image:nsf2.png|center|300px|]]
 
[[Image:nsf2.png|center|300px|]]
 
[[Image:USDAlogo4.jpg|center|280px|]]
 
[[Image:USDAlogo4.jpg|center|280px|]]
 +
 +
 +
'''COMPLETED PROJECTS'''
 +
 +
'''Defining the Tyrosine Biosynthetic Pathways in Plants'''  [[Image:Tyrpathway.png|right|400px|]] 
 +
''Funded by NSF IOS grant-1354971, 9/01/2014 – 8/31/2018''
 +
*This project uncovered unique diversification of the primary amino acid pathway—tyrosine biosynthesis—in different plants, likely to support tremendous chemical diversity of tyrosine-derived natural products (Schenck and Maeda, 2018). We found that Arabidopsis makes tyrosine mainly via plastidic arogenate dehydrogenase (TyrAa) enzymes (Wang et al., 2016; de Oliveira et al., 2019; Wang 2019), whereas legumes have an additional alternative tyrosine pathway catalyzed via a novel feedback-insensitive prephenate dehydrogenase (TyrAp) enzymes (Scheck et al., 2015, 2017a). Phylogeny-guided structure-function analyses of plant and microbial TyrA enzymes further identified a single aspartate residue that confers arogenate dehydrogenase activity and simultaneously affects tyrosine inhibition (Schenck et al., 2017a; 2017b). The project also provided rigorous independent research opportunities for ten undergraduate students and conducted “Pigment-Art” outreach activity to inform general public about the nutritional and pharmacological values of plant natural products (see our outreach page).
 +
 +
 +
'''Elucidation of the Tyrosine Pathway in Table Beets'''
 +
''Funded by USDA NIFA grant-2015-67013-11631139, 3/01/2015 – 2/28/2019)''
 +
*Table beets uniquely accumulate high levels of tyrosine-derived red/yellow pigments, betalains (Wang et al., 2017). Similar to Arabidopsis, beets (''Beta vulgaris'') had two genes (''BvTyrAα'' and ''BvTyrAβ'') encoding plastidic arogenate dehydrogenase (TyrA) enzymes. Interestingly, however, BvTyrAα but not BvTyrAβ exhibited reduced sensitivity to tyrosine inhibition (Lopez-Nieves et al., 2018). Detailed phylogenetic analysis of ''BvTyrAα'' and ''BvTyrAβ'' orthologs identified from transcriptome data of nearly one hundred Caryophyllales plants showed that ''BvTyrAα'' orthologs emerged prior to the appearance of betalain pigmentation. The results suggest that a lineage-specific de-regulation of primary metabolism (i.e. tyrosine biosynthesis) facilitated the evolution of downstream betalain biosynthesis (Lopez-Nieves et al., 2018) and various natural product pathways derived from tyrosine (Lopez-Nieves et al., In preparation). The identified de-regulated BvTyrAα enzymes can be further utilized to enhance the availability of the tyrosine precursor and the production of tyrosine-derived natural products (Timoneda et al., 2018; Maeda 2019b).
 +
 +
 +
'''Evolution of Plant Phenylalanine Biosynthesis'''
 +
[[Image:Camilla_image_2.png|right|320px|]] 
 +
*In plants, phenylalanine (Phe) is a precursor of abundant and diverse phenylpropanoid compounds, such as flavonoids, tannins, and lignin. Plants synthesize Phe predominantly via the arogenate pathway ([[Maeda_Lab:Publications |'''Maeda ''et al''., 2010; 2011''']]) in contrast to model microbes (e.g. ''E. coli'') that use exclusively the phenylpyruvate pathway for Phe biosynthesis. Together with [https://www.ppws.vt.edu/people/faculty/jelesko-john.html Dr. John Jelesko] at Virginia Tech, we conducted phylogenetically-informed biochemical characterization of prephenate aminotransferase (PPA-AT), which catalyzes the first committed step of the arogenate pathway, and uncovered a unique evolutionary history of the arogenate Phe pathway. Our results revealed that prephenate-specific aminotransferase containing Thr84 and Lys169 was transferred from a Chloribi/Bacteroidetes ancestor to a eukaryotic ancestor of the Plantae. This ancient lateral gene transfer likely led to efficient Phe production via the arogenate pathway, which currently supports the production of abundant phenylpropanoid natural products such as lignin. ([[Maeda_Lab:Publications |'''Dornfeld ''et al''., 2014''']])

Revision as of 18:23, 27 January 2020

Maeda lab banner 2020.jpg

Home        Research        Members        Publications        Protocols        Outreach       


Research Interests in Maeda Lab

Our goal is to understand remarkable chemical and metabolic diversity that evolved in the plant kingdom, in order to redesign plants for efficient and sustainable production of natural and bio-products. Plants produce a diverse array of secondary (specialized) metabolites, which play critical roles in plant adaption to various ecological niches. In contrast to well-documented diversification of plant secondary metabolism, primary metabolism, which provides precursors of secondary metabolites, is generally thought to be conserved across different plant species. However, our lab has recently uncovered lineage-specific diversifications of primary metabolic pathways in different plant groups, likely to support downstream synthesis of diverse secondary metabolites (Schenck et al. 2015; 2017a; Lopez-Nieves et al. 2018; Maeda 2019a). Such basic knowledge of the interface between plant primary and secondary metabolism will be crucial for future breeding, metabolic engineering, and synthetic biology to improve production of natural and bio-products in plant hosts (Maeda 2019b).

We are currently conducting various projects that focus on aromatic amino acid biosynthesis, which connects central carbon metabolism and diverse plant natural product pathways. Currently, we are further i) exploring other examples of primary metabolic diversity in the aromatic amino acid pathways, ii) investigating underlying genetic and biochemical mechanisms, iii) utilizing the knowledge to engineer aromatic amino acid biosynthesis and improve precursor metabolite availability, and iv) analyzing physiological impacts of metabolic engineering on plant growth and development.


Evolutionary Diversification of Tyrosine Biosynthesis and Downstream Natural Product Pathways

  • Primary metabolites are indispensable cellular components that directly impact plant growth and development. Consequently, primary metabolic pathways are evolutionarily constrained, typically conserved among the plant kingdom, and difficult to manipulate. Our lab, however, uncovered lineage-specific diversification of a primary amino acid pathway, tyrosine biosynthesis, in legumes (Schenck et al. 2015; 2017a; 2017b; Maeda 2019a). We also revealed that de-regulation of tyrosine biosynthesis occurred at the base of core Caryophyllales and facilitated later evolution of a novel pigment pathway, betalain biosynthesis (Lopez-Nieves et al. 2018).
  • Combining evolutionary biochemistry, metabolomics, and phylogenomic analyses, we are further exploring evolutionary alterations of tyrosine biosynthesis and their impacts on associated metabolic network in various plant lineages. The study can advance our understanding of the step-wise evolution of complex metabolic traits at macroevolutionary scale and explore how the evolution of such traits influence subsequent adaptation and diversification in plants. As a part of broader impact activity, we will be also generating chemotaxonomic databases together with students to study evolutionary history, mechanism, and functions of tremendous chemical diversity that exist in the plant kingdom.

Collaborators: Michael McKain (Univ. Arabama), James Leebens-Mack (Univ. Georgia), Ya Yang (Univ Minnesota), Samuel Brockington (Cambridge), and Stephen Smith (Univ. Michigan)

Read more at our NSF PGRP project website (under construction):


Regulation of the Shikimate and Aromatic Amino Acid Pathways in Plants

  • The shikimate pathway directs carbon flux from central carbon metabolism (e.g. Calvin-Benson cycle, glycolysis) to biosynthesis of aromatic amino acids (AAAs) and numerous AAA-derived plant natural products (Maeda and Dudareva, 2012). However, it is unknown how plants regulate the shikimate pathway. We are investigating molecular mechanisms underlying de-regulation of plant AAA biosynthesis and analyzing their quantitative impacts on overall plant metabolic network. Here we are utilizing enzyme biochemistry, forward and reverse genetics, 13CO2 labeling, and structure-function enzyme analyses, as well as metabolic pathway modeling. The obtained enzyme variants and knowledge can be used to boost the production of AAAs and their derived natural products in plants.

Collaborators: Joseph Jez (WUSTL), Alisdair Fernie (MPI-Golm), Mark Stitt (MPI-Golm), Zoran Nikoloski (Univ. Potsdam)


Crop-Based Production of Natural Products through Synthetic Biology

  • Plant natural products provide valuable nutraceuticals, pharmaceuticals, and bio-products. Global crop production systems provide sustainable and potentially efficient chemical production platforms, as plants can naturally produce and store abundant chemicals. However, little effort has been made to identify and build plant hosts or chassis for sizable production of natural products.
  • We are addressing two outstanding questions thwarting this issue: i) Which plants serve as efficient chemical production platforms and ii) what is the contribution of primary metabolite precursor supply for natural product synthesis? Utilizing our de-regulated tyrosine biosynthetic enzymes (Schenck et al. 2015; 2017; Lopez-Nieves et al. 2018) we are conducting proof-of-concept experiments by expressing tyrosine-utilizing enzymes in transgenic Arabidopsis and other crops accumulating different levels of tyrosine. The study will generate prototype crops for nutrient-enriched functional food, feed, and natural dye, and also lay essential groundwork towards crop-based production of natural and bio-products.

Collaborator: Wisconsin Crop Innovation Center (WCIC)


Amino Acid Sensing and Response Mechanisms in Plants

  • The long-term goal of the Maeda laboratory is to improve the production of amino acids and amino acid-derived plant natural products. Perturbation of amino acid metabolism, however, often leads to pleiotropic growth and developmental responses and very little is known how amino acid homeostasis is regulated and linked to plant growth and development. We previously isolated partially tyrosine-deficient tyra2 mutants as well as tyrosine hyper-accumulating lines of Arabidopsis, both of which exhibit compromised growth and unique reticulate leaf phenotypes (de Oliveira et al. 2019). These findings suggest that imbalance of aromatic amino acids impacts plant growth and development. Here we are conducting physiological, genetic, and biochemical analyses to elucidate mechanisms underlying these pleiotropic plant responses to amino acid imbalance. Fundamental understanding of amino acid homeostasis mechanisms will enhance plants’ resilience to amino acid perturbation and hence have broad impacts on improving yield and nutritional quality of crops.


Defining the Nitrogen Flux Maps (NFMs) of Plants

  • Nitrogen (N) is a critical element of organic molecules, including aromatic amino acids, but is highly limited in plants. Thus, N use efficiency directly impacts overall yield and performance of plants. Unlike extensively-studied carbon (C) flux map of plant metabolism,little is known how assimilated N flows through the metabolic network, namely “N flux map (NFM)”. The core of NFM is different branches of amino acid metabolism interconnected by aminotransferases (ATs), which play pivotal roles in distributing reduced N for synthesis of various organonitrogen compounds (Dornfeld et al., 2014; Wang et al., 2016; Wang and Maeda, 2017; Wang et al., 2019). Utilizing the wealth of sequenced plant genome information, high-throughput gene and protein syntheses, and enzyme assay platforms, we aim to determine multi-substrate specificities of all ATs from different plant species and construct NFMs from Arabidopsis and Sorghum. The resulting NFMs will provide foundation to further optimize N metabolic network and to generate plants that can produce high levels of N-containing compounds (e.g. alkaloids) while maintaining robust growth even in reduced N fertilizer.

Collaborators: Trent Northen (Lawrence Berkeley National Laboratory, LBNL), Yasuo Yoshikuni (LBNL, JGI), Zoran Nikoloski (Max Planck Institute Golm/Univ Potsdam), Taichi Takasuka (Hokkaido Univ)

Read more at our DOE BER project website


Our studies are supported by UW-Madison, NSF, USDA NIFA, and DOE BER.
UWlogo.png
nsf2.png
USDAlogo4.jpg


COMPLETED PROJECTS

Defining the Tyrosine Biosynthetic Pathways in Plants
Tyrpathway.png

Funded by NSF IOS grant-1354971, 9/01/2014 – 8/31/2018

  • This project uncovered unique diversification of the primary amino acid pathway—tyrosine biosynthesis—in different plants, likely to support tremendous chemical diversity of tyrosine-derived natural products (Schenck and Maeda, 2018). We found that Arabidopsis makes tyrosine mainly via plastidic arogenate dehydrogenase (TyrAa) enzymes (Wang et al., 2016; de Oliveira et al., 2019; Wang 2019), whereas legumes have an additional alternative tyrosine pathway catalyzed via a novel feedback-insensitive prephenate dehydrogenase (TyrAp) enzymes (Scheck et al., 2015, 2017a). Phylogeny-guided structure-function analyses of plant and microbial TyrA enzymes further identified a single aspartate residue that confers arogenate dehydrogenase activity and simultaneously affects tyrosine inhibition (Schenck et al., 2017a; 2017b). The project also provided rigorous independent research opportunities for ten undergraduate students and conducted “Pigment-Art” outreach activity to inform general public about the nutritional and pharmacological values of plant natural products (see our outreach page).


Elucidation of the Tyrosine Pathway in Table Beets Funded by USDA NIFA grant-2015-67013-11631139, 3/01/2015 – 2/28/2019)

  • Table beets uniquely accumulate high levels of tyrosine-derived red/yellow pigments, betalains (Wang et al., 2017). Similar to Arabidopsis, beets (Beta vulgaris) had two genes (BvTyrAα and BvTyrAβ) encoding plastidic arogenate dehydrogenase (TyrA) enzymes. Interestingly, however, BvTyrAα but not BvTyrAβ exhibited reduced sensitivity to tyrosine inhibition (Lopez-Nieves et al., 2018). Detailed phylogenetic analysis of BvTyrAα and BvTyrAβ orthologs identified from transcriptome data of nearly one hundred Caryophyllales plants showed that BvTyrAα orthologs emerged prior to the appearance of betalain pigmentation. The results suggest that a lineage-specific de-regulation of primary metabolism (i.e. tyrosine biosynthesis) facilitated the evolution of downstream betalain biosynthesis (Lopez-Nieves et al., 2018) and various natural product pathways derived from tyrosine (Lopez-Nieves et al., In preparation). The identified de-regulated BvTyrAα enzymes can be further utilized to enhance the availability of the tyrosine precursor and the production of tyrosine-derived natural products (Timoneda et al., 2018; Maeda 2019b).


Evolution of Plant Phenylalanine Biosynthesis

Camilla image 2.png
  • In plants, phenylalanine (Phe) is a precursor of abundant and diverse phenylpropanoid compounds, such as flavonoids, tannins, and lignin. Plants synthesize Phe predominantly via the arogenate pathway (Maeda et al., 2010; 2011) in contrast to model microbes (e.g. E. coli) that use exclusively the phenylpyruvate pathway for Phe biosynthesis. Together with Dr. John Jelesko at Virginia Tech, we conducted phylogenetically-informed biochemical characterization of prephenate aminotransferase (PPA-AT), which catalyzes the first committed step of the arogenate pathway, and uncovered a unique evolutionary history of the arogenate Phe pathway. Our results revealed that prephenate-specific aminotransferase containing Thr84 and Lys169 was transferred from a Chloribi/Bacteroidetes ancestor to a eukaryotic ancestor of the Plantae. This ancient lateral gene transfer likely led to efficient Phe production via the arogenate pathway, which currently supports the production of abundant phenylpropanoid natural products such as lignin. (Dornfeld et al., 2014)
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