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Projects
Enzymes of
Steroidal Glycoalkaloid Biosynthesis
Despite
the world-wide consumption of potatoes and tomatoes, little is known about
the formation of potentially lethal toxins called steroidal gycoalkaloids (SGAs) that are present in these
plants. At the concentrations they are found in potato tubers and tomato fruit, they give a slightly bitter flavor that is characteristic of these vegetables. At higher concentrations, they are part of the plant chemical defense against herbivores such as Colorado potato beetle. Fortunately, both plants have been bred to limit the accumulation of
steroidal glycoalkaloids in the edible parts of the plant. We are
using wild potato to understand the biosynthesis of these compounds and
their natural biological role so that we can improve the cultivated
varieties for both chemical defense in the nonedible parts and food safety
in tomato fruits or potato tubers. We have developed tools to rapidly screen the function of targeted genes in root cultures of wild potato. .
The primary metabolic pathway leading to SGA formation is shared with biosynthesis of phytosterol, triterpene and brassinosteroid classes of compounds. We are investigating the enzymes found in common with the formation of all four classes to identify mechanisms that allow for the specialization or channeling of substrates for one or more class of compounds. We have determined that wild potato has at least three genes coding for enzymes that are identical or very similar to squalene synthase. The genes have an unusual structure whose function is unknown but may be related to the fidelity of protein synthesis.
Antiherbivore
Properties of Different Glucosinolate Hydrolysis Products
The glucosinolate-myrosinase system is a substrate-enzyme-product reaction
associated with plant defense against herbivores and pathogens and is found
in the crucifer family. The substrate and enzyme are normally
separate, but upon tissue damage, the two components interact resulting in the formation of cytotoxic breakdown products. These toxins
interfere with insects feeding on the plant and are thought to function
similarly with microbial pathogens. The chemical defenses of plants usually
involve such so-called secondary metabolites. A central dilemma in our
understanding of secondary metabolites--their biosynthesis and biological
function, is the diversity of compounds that a plant makes.
Leaves of Arabidopsis thaliana have 15 easily-identified
glucosinolates. The entire species produces over 35 different
glucosinolates. To understand why the plant makes so many different
glucosinolates, the Tokuhisa lab is interested in the levels of antifeeding activity created by the various breakdown products that arise
from the hydrolysis of glucosinolates by myrosinase. Different herbivores
and microbes will be assessed for their growth response to various
glucosinolate breakdown products.
The formation of specific cytotoxic breakdown products is dependent not only
on the glucosinolate and the myrosinase reaction but also by the specific
glucosinolate structure, accessory proteins that guide the enzymatic
reaction, and the pH and metal ion environment of the enzyme reaction.
These conditions are defined by the plant and can be altered by transgenic
technologies. The Tokuhisa lab has altered the glucosinolate profile in
Arabidopsis thaliana and will change other components of the
glucosinolate-myrosinase system using gene knockout, overexpression and gene
silencing strategies to modify the profile of glucosinolate breakdown
products in transgenic Arabidopsis. These plants will be tested for defense responses to various herbivores and pathogens.
The Physiology of Glucosinolate
Accumulation
We are mining Arabidopsis transcriptome, proteome and metabolome data to
formulate models of the dynamic physiology of the glucosinolate-myrosinase
system during development and environmental stress. We are using
mutant and transgenic plant lines coupled with analytical biochemistry to
test our models.
Grafted Tomato for Sustainable Field Production
We are analyzing the potential of grafted tomato to expand the phenotypic plasticity of tomato for field production. |