15th International Conference on Arabidopsis Research - TAIR

T03-057
A Proteomic Analysis of Leaf Peroxisomes
Lavanya Babujee(1), Franziska Lüder(1), Virginie Wurtz(2), Hartmut Kratzin(3),
Sigrun Reumann(1)
1-University of Göttingen, Dept. Plant Biochemistry, Justus-von-Liebig-Weg 11, D-37077 Göttingen,
email: sreuman@gwdg.de
2-LSMBO, Batiment R5, 25, rue becquerel, F-67087 Strasbourg cedex 2
3-Max-Planck-Institut für experimentelle Medizin, Hermann-Rein-Str. 3, 37075 Göttingen
The soluble matrix proteome of purified leaf peroxisomes from spinach and
Arabidopsis was partially characterized by two-dimensional electrophoresis
followed by mass spectrometry (MS). Apart from well-known peroxisomal
proteins, such as those involved in photorespiration, fatty acid beta oxidation
and metabolism of reactive oxygen species, some proteins were identified,
including a short chain reductase, two enoyl-CoA hydratases/isomerases
and a small heat shock protein (sHSP) as novel protein components of leaf
peroxisomes. Because fractions highly enriched in leaf peroxisomes still tend
to be contaminated by proplastids and mitochondria, the subcellular localization
of the novel proteins was verified by differential 2D gels. Further support
for peroxisomal localization is provided by cloning of the corresponding
Arabidopsis genes and analyzing subcellular targeting of fusion proteins with
spectral variants of green fluorescent protein in onion epidermal cells. Posttranslational
modifications (PTMs) including phosphorylation appeared to a
salient feature of several peroxisomal proteins, even though not reported yet
for any peroxisomal protein. Some novel proteins and/or isoforms appeared
to be induced during conditions of stress. The MS identification of the novel
proteins and the characterization of the PTMs are currently underway and are
likely to contibute to a better understanding of the metabolic capacity of leaf
peroxisomes.
15 th <strong>International</strong> <strong>Conference</strong> on Arabidopsis Research 2004 · Berlin
T03-058
Genetic analysis of the AtRabGDI family
Hana Soukupova(1), Michal Hala(1), Lukas Synek(1, 2), Viktor Zarsky(1, 2)
1-Laboratory of Cell Biology, IEB ASCR, Rozvojova 135, Praha 6, 165 02 Czech Republic
2-Department of Plant Physiology, Charles University, Vinicna 5, Praha 2, 128 44 Czech Republic
Rab GDP dissociation inhibitors (RabGDIs) are required for proper recycling
of small RabGTPases by their retrieval from target membrane compartments
and for cytosolic binding of RabGTPases in the inactive form. Therefore,
RabGDIs activity is essential for the control of vesicular transport.
Sequencing of Arabidopsis thaliana and Oryza sativa genomes suggested
that angiosperm possess three isotypes of RabGDI. We have started our
analysis of the AtRabGDI family using L59 mutant containing T-DNA insertion
in the first intron of AtRabGDI1 (provided by dr. Klaus Palme (Köln, Germany)).
Detailed analysis of this insertion showed that homozygous plants have no
observable phenotype. The same hold true about the AtRabGDI2 insertional
mutant (from the Salk Institute database) with T-DNA insertion in the fifth
exon. There are no convenient candidates for any insertion in AtRabGDI3 in
public collections. We have obtained no double homozygous mutant from
genetical analysis of reciprocal crosses between AtRabGDI1 and AtRabGDI2
mutants suggesting that disruption of both AtRabGDIs function is lethal.
Female and male gamets harboring both AtRabGDI1 and AtRabGDI2 mutated
alleles are functional. Specific expression of AtRabGDI3 (according to Affymetrix
data) may complement this defficiency in male gamets.
This work was supported by GACR 206/99/1138 and MSMT CR-
LN00A081”SIDROS” grants.
T03 Cell Biology