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The role of amino acid metabolism in response to biotic and abiotic stress factors in Arabidopsis thaliana

Friday, 24 June, 2011 - 17:00
Campus: Brussels Humanities, Sciences & Engineering campus
Faculty: Science and Bio-engineering Sciences
Sigrid Brauč
phd defence

Due to the increasing population and as a result of the increasing demand for plant derived
products, plant production has received extremely important economical interest. However,
modern agriculture is facing important challenges. Plants must continuously protect themselves
against different kinds of stresses. Biotic stresses, caused by attack from bacteria, viruses, fungi,
insects, etc. and abiotic stresses, caused by physical and chemical challenges such as drought,
salinity, high or low temperatures, etc. (ir)reversibly affect plants’ growth, development and
production. Therefore, plants have developed complex defence mechanisms to cope with different
environmental stresses. These defence mechanisms are known to interact, most probably evolving
from the simultaneous multiple stress confrontation in their natural environment. Nevertheless, the
sophisticated defence mechanisms present in plants are mostly not sufficient to cope with the
deleterious effects of stresses and result in reduced growth and productivity. Therefore, substantial
research effort goes to unravelling plant defence mechanisms against abiotic as well as biotic
stresses in order to find novel sustainable agronomic applications.

The result of defence mechanisms of plants during biotic and abiotic stress exposure can lead to
up-regulation of genes, leading to the production or accumulation of defence molecules, which
could eventually be amino acids, amino acid derivates or proteins (built up of amino acids). The
amino acid proline and ã-aminobutyric acid (GABA) and polyamines are well-studied molecules in
this topic. The first one is known to accumulate to high concentrations upon abiotic stress, whereas
the last two accumulate upon biotic as well as abiotic stress imposition.

Most defence molecules have an amino acid as precursor. Therefore our question arose: Are other
amino acids also involved in abiotic stress response?

Gene-expression analysis of Arabidopsis, tomato and a tomato mutant showing increased
resistance to the necrotrophic pathogen Botrytis cinerea revealed the up-regulation of a wide range
of genes upon infection with B. cinerea. Genes encoding enzymes of the amino acid metabolic
pathways were found to be induced, beyond the expected up-regulation of genes known to be
involved in defence mechanisms. Five genes directly involved in amino acid metabolism were found
to be up-regulated in both (Arabidopsis and tomato) or at least one organism: lysine-ketoglutarate
reductase/saccharopine dehydrogenase (LKR/SDH), genes belonging to the aspartate
aminotransferase (AspAT) and alanine aminotransferase (AlaAT) family, arginase (ARG) and
glutamate decarboxylase (GAD). These genes are not only differentially expressed after infection
with B. cinerea, but also abiotic stress appeared to influence the expression of those genes in
Arabidopsis. Using quantitative RT-PCR we demonstrated that those genes are induced early after
stress treatment albeit with different amplitudes.

The aim of this study was to investigate the contribution of the listed genes, involved in amino acid
metabolism, to defence mechanisms in Arabidopsis. Therefore, over-expression, knockout and/or
silencing lines have been created in Arabidopsis thaliana. These lines were characterised and
subsequently submitted to salt and osmotic stress treatments and further infected with the
necrotrophic pathogen B. cinerea. Results of this study demonstrated that all those genes
influenced plant defence mechanisms in one way or another.

Arginase over-expression lines exhibited about 10% reduced susceptibility to B. cinerea compared
to the wild-type. This increased resistance probably arises from increased stress-related
metabolites in those transgenic lines. Further, transgenic lines with disturbed nitrogen metabolism,
AspAT transgenic lines, showed up to 20% increased lesion formation upon infection with B.
cinerea. This is supposed to be due to decreased stress-related metabolites or due to perturbed
nitrogen assimilation, which would be in favour of the pathogen. Finally, LKR/SDH over-expression
lines, predicted to produce increased stress-related metabolites via glutamate, showed to possess
increased resistance against the pathogen. LKR/SDH knockout mutants similarly displayed
decreased susceptibility towards the pathogen resulting from another, yet unknown, defence

The photorespiratory AlaAT was shown to play an important role in sustaining growth on standard
conditions and upon abiotic stress. Similarly, transgenic lines possessing increased and decreased
lysine contents were affected in their growth in most of the tested conditions. However, the way
these transgenic lines are affected in their response to stress is different, demonstrating the
diversity of the role of amino acids in plants.

The results presented in this manuscript confirmed the role of already described stress-related
molecules such as proline and GABA. Furthermore, through over-expression and silencing lines,
novel roles of genes which were not expected to be directly related to stress response were
discovered. Amino acid metabolism, thus demonstrated to play an important role in stress
responses. The proposed results open new insights in the role of amino acids in stress responses
and create new horizons for orienting research of these important molecules.

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