15th International Conference on Arabidopsis Research - TAIR

T07-071
Mutants in medium long and long chain acyl-
CoA oxidase activity demonstrate that long chain
acyl-CoA activity is important in seed viability and
essential for seedling establishment.
Elizabeth L. Rylott(1), Helen Pinfield-Wells(1), Alison D. Gilday(1), Ian A. Graham(1)
1-CNAP Dept. of Biology, University of York
The Arabidopsis acyl-CoA oxidase (ACX) family comprises isozymes with distinct
fatty acid chain-length specificities that together catalyse the first step
of peroxisomal fatty acid ß-oxidation. We have isolated and characterised T-
DNA insertion mutants in the medium long-chain (ACX1) and long chain acyl-
CoA oxidases (ACX2). In acx1 seedlings, medium-long (C10:0-C20:0)-chain
acyl-CoA oxidase activity was greatly reduced by greater than 95 percent for
C16:0. In acx2 mutant seedlings, specifically long-chain activity was reduced,
by greater than 90 percent for C18:0. Whilst lipid catabolism during germination
and early post-germinative growth was unaltered in the acx1 mutant
and only slightly delayed in the acx2 mutant, both mutants accumulated acyl-
CoAs. Three-day-old acx1 seedlings accumulated mainly medium/long chain
acyl-CoAs, whilst acx2, accumulated only long chain acyl-CoAs. In acx1 and
acx2, seedling growth and establishment in the absence of an exogenous
supply of sucrose, was unaffected. Furthermore, no alterations in vegetative
or reproductive phenotype were obvious throughout the remaining life cycle
of the acx1 or acx2 plants.
Seedlings of the double mutant acx1acx2 exhibited a 98% reduction in
C16:0 activity, were unable to catabolise seed storage lipid and accumulated
long chain acyl-CoAs. In the absence of an exogenous sucrose supply,
acx1acx2 seedlings were unable to establish photosynthetic competency.
However when rescued on sucrose at the seedling stage, no alterations in
vegetative or reproductive phenotype were seen throughout the remaining
lifecycle. Germination frequency was less than 30 percent of wild type seeds,
and was unaffected by the addition of exogenous sucrose. This phenotype
resembles the comatose (cts2) mutant.
We propose that the acx1 and acx2 mutants are partially complemented by
ACX2 and ACX1 respectively, and that acx1acx2 seedlings require sucrose
for seedling establishment because they have a more severe restriction
in long-chain acyl-CoA oxidase activity. Furthermore, long chain acyl-CoA
oxidase activity plays an important role in seed viability.
T07 Metabolism (Primary, Secondary, Cross-Talk and Short Distance Metabolite Transport)
T07-072
Reserve Mobilisation in the Arabidopsis Endosperm
Fuels Hypocotyl Elongation in the Dark, is
Independent of Abscisic Acid and Requires the
PHOSPHOENOLPYRUVATE CARBOXYKINASE1 Gene
Steven Penfield(1), Elizabeth R. Rylott(1), Alison D. Gilday(1), Stuart Graham(1), Tony
R. Larson(1), Ian A. Graham(1)
1-CNAP, Department of Biology, University of York, PO BOX 373, York ,YO10 5YW, United Kingdom
Arabidopsis is used as a model system to study triacylglycerol (TAG) accumulation
and seed germination in oilseeds. Here we consider the partitioning of
these lipid reserves between embryo and endosperm tissues in the mature
seed. The Arabidopsis endosperm accumulates significant quantities of
storage lipid, and this is effectively catabolised upon germination. This lipid
differs in composition from that in the embryo and has a specific function
during germination. Removing the endosperm from wild-type seeds resulted
in a reduction in hypocotyl elongation in the dark, demonstrating a role
for endospermic TAG reserves in fuelling skotomorphogenesis. Seedlings
of two allelic gluconeogenically compromised PHOSPHOENOLPYRUVATE
CARBOXYKINASE (PCK1) mutants show a reduction in hypocotyl length in
the dark compared to wild-type, but this is not further reduced by removing
the endosperm. The short hypocotyl phenotypes were completely reversed
by the provision of an exogenous supply of sucrose. The PCK1 gene is
expressed in both embryo and endosperm, and the induction of PCK1:GUS at
radical emergence occurs in a robust wave-like manner around the embryo
suggestive of the action of a diffusing signal. Strikingly, the induction of PCK1
promoter reporter constructs and measurements of lipid breakdown demonstrate
that while lipid mobilisation in the embryo is inhibited by abscisic acid
(ABA), no effect is seen in the endosperm. This insensitivity of endosperm
tissues is not specific to lipid breakdown as hydrolysis of the seed coat cell
walls also proceeded in the presence of concentrations of ABA that effectively
inhibit radical emergence. Both processes still required gibberellin however.
These results suggest a model whereby the breakdown of seed carbon
reserves is regulated in a tissue specific manner and shed new light on
phytohormonal regulation of the germination process.
15 th <strong>International</strong> <strong>Conference</strong> on Arabidopsis Research 2004 · Berlin