15th International Conference on Arabidopsis Research - TAIR

T07-087
Flavonol glycosyltransferases in Arabidopsis thaliana
Burkhard Messner(1), Patrik Jones(2), Birgit Geist(1), Susanna Holzinger(1), Kazuki
Saito(2), Tony R. Schaeffner(1)
1-Institute of Biochemical Plant Pathology, GSF ¯ National Research Center for Environment and
Health, D-85764 Neuherberg, Germany
2-Department of Molecular Biology and Biotechnology, Chiba University, CREST of Japan Science
and Technology Corporation, Yayoi-cho 1-33, Inage-ku, Chiba 263-8522, Japan
Flavonol glycosides constitute a prominent class of secondary metabolites
in A. thaliana as well as several crop plants. In Arabidopsis, mainly kaempferol
and quercetin occur in glycosylated forms with carbohydrate residues
attached at different positions. However, the conjugating enzymes have been
elusive so far. Arabidopsis thaliana harbors more than 100 UDP-sugar dependent
glycosyltransferase genes (UGT). Based on sequence homologies to
known flavonoid glycosyltransferases from other plant species, the UGT78D
and UGT73B/C subgroups were targeted as candidate flavonoid-UGTs from
Arabidopsis. Indeed, metabolic profiles of T-DNA knockout lines showed
the reduction of quercetin-3-O-rhamnoside-7-O-glucoside (ugt73C6 and
ugt78D1) as well as flavonol-3-O-rhamnoside-7-O-rhamnosides (ugt78D1)
and two flavonol-3-O-glucoside derivatives (ugt78D2). The latter two mutants
indicated that UGT78D1 and UGT78D2 were responsible for the initial 3-Oglycosylation
of flavonols but discriminated between rhamnosylation and glucosylation.
Interestingly, UGT78D2 was recently also found to be upregulated
in an anthocyanin-overaccumulating mutant and shown to be responsible for
anthocyanin-3-O-glucosylation as well (Nisiyama et al, 2004). In accordance
with these mutant data, recombinantly expressed UGT78D1 and UGT78D2
catalyzed the rhamnosylation of the 3-OH group of quercetin and kaempferol,
whilst UGT73C6 catalysed the glucosylation of the 7-OH group of kaempferol-
and quercetin-3-O-rhamnosides. Thus, these analyses identified the first
A. thaliana flavonol glycosyltransferases. The respective mutants and double
mutants created will be used for analyzing the function of differential and
complex glycosylation of flavonols in Arabidopsis.
Nisiyama Y et al. (2004) Plant Cell Physiol., Supplement 45: s157.
T07 Metabolism (Primary, Secondary, Cross-Talk and Short Distance Metabolite Transport)
T07-088
MOLECULAR AND GENOMIC ANALYSIS OF
NITROGEN REGULATION OF AMINO ACID PERMEASE
I (AAP1) IN ARABIDOPSIS
Mengjuan Guo(1), Daniel R. Bush(2, 1)
1-Plant Biology, UIUC Urbana, IL
2-ARS Photosynthesis Research, IL. and Biology Department, Colorado State University, Fort
Collins, CO 8053
In higher plants, amino acids are the currency of nitrogen exchange
between source and sink tissues at every stage of plant growth and development.
We are investigating AAP1 as a prototypical example of a plant
proton-amino acid symporter that plays an important role in amino acid
partitioning. We’ve previously shown that AAP1 expression is regulated by
changes in nitrate, ammonia, and amino acid abundance, as well as changes
in carbon metabolism.
Semi-quantitative RT-PCR showed nitrate and glutamate are metabolite
signals that induce AAP1 expression after nitrogen starvation. Microarray
analysis showed that application of exogenous nitrate, glutamate and glutamine
differentially regulate the expression of AAP1 and several other genes
involved in key aspects of plant metabolism and assimilate partitioning.
Significantly, there was little congruence between the genes regulated by
each of these metabolic signals. This observation was supported in a T-DNA
insertion knockout of AAP1 in which a short fragment of the AAP1 promoter
was sufficiently long to retain nitrate regulatory elements, but the same
region was too short to preserve sensitivity to glutamate regulation.
AAP1:GUS plants demonstrated that AAP1 expression is developmentally
regulated in both source and sink tissues. In situ hybridization identified the
cell-specific AAP1 expression in the phloem cells of minor veins, consistent
with a role in phloem loading of amino acids. Overexpression of AAP1 results
in shorter roots and altered leaf shape in seedlings, while RNAi knockouts
flowered later than wild type plants. These data support the hypothesis that
AAP1 plays an important role in nitrogen partitioning.
Taken together, the evidence presented here shows that there are multiple
nitrogen-metabolite mediated regulatory pathways that differentially modulate
gene expression in response to dynamic changes in nitrogen status.
15 th <strong>International</strong> <strong>Conference</strong> on Arabidopsis Research 2004 · Berlin