15th International Conference on Arabidopsis Research - TAIR

T05-069
Cell-specific Gene Activation by Salicylic Acid
Kate Wilson(1), John Carr(1)
1-Dept. of Plant Sciences, Cambridge University, UK
Cell-specific Gene Activation by Salicylic Acid.
Induction of systemic acquired resistance (SAR) to plant pathogens (fungi,
bacteria and viruses) requires activation of a signal transduction pathway
that includes salicylic acid (SA) as a key component. The signal transduction
pathway branches downstream of SA. One branch leads to induction of
pathogenesis-related (PR) protein genes and resistance to bacteria and fungi.
The other branch activates resistance to virus replication and movement
(Sing et al., 2004, MPP 5: 57-64). SA induced changes in host gene expression
may affect viruses by: a) induction of host genes encoding inhibitors of
virus replication or movement, or b) repression of genes encoding proteins
required to support virus replication or movement. These changes are likely
to occur in a cell/tissue-specific manner because findings in our laboratory
showed that in the leaf mesophyll cells of SA-treated tobacco plants the
replication of TMV is greatly decreased (Murphy & Carr, 2002, Plant Phys. 28:
552-563). In contrast, SA did not decrease TMV replication in the initially inoculated
epidermal cells. Instead, it induced resistance to movement between
the epidermal cells. To identify cell-specific SA-induced changes in gene
expression, more than 2000 transgenic lines of Arabidopsis which carry random
insertions of the enhancer trap T-DNA (Haseloff, 1999, Methods in Cell
Biology, Vol. 58) have been generated. If the enhancer trap T-DNA integrates
close to a SA-activated promoter/enhancer, the synthesis of GFP will be activated.
Approximately 700 of these lines have been screened for SA-induced
cell-specific GFP fluorescence using epifluorescent microscopy. Eight lines,
showing SA-induced alterations in tissue-specific GFP fluorescence in the
root, have been rescued to date. These lines are being further characterized
with the aim of isolating new genes regulated by the viral defense pathway.
15 th <strong>International</strong> <strong>Conference</strong> on Arabidopsis Research 2004 · Berlin
T05-070
dsRNAi ¯ A reverse genetic tool to discover gene
function in plant nonhost resistance
Christina Neu(1), Bekir Ülker(1), Paul Schulze-Lefert(1)
1-Max-Planck Institut für Züchtungsforschung Carl-von-Linné-Weg 10. D-50829 Köln, Germany
Post-transcriptional gene silencing triggered by self-complementary hairpin
RNA is widely used to knockdown transcripts of genes to elucidate their
function. Compared to conventional mutant screens, dsRNAi technology has
the potential to overcome the problem of genetic redundancy by targeting
highly sequence related members of a gene family. When used in combination
with an inducible expression system, dsRNAi might facilitate the analysis
of genes whose heritable inactivation lead to severe developmental defects
or lethality. To identify new components of nonhost disease resistance,
hundreds of dsRNAi lines will be generated to target genes whose expression
is shown to be up-regulated upon pathogen infection. The dsRNAi
lines will be screened for altered infection phenotypes upon challenge with
inappropriate foliar pathogens. To streamline the cloning procedure we
developed GATEWAY compatible dsRNAi plant transformation vectors for
constitutive and ethanol(EtOH)-inducible gene silencing. In order to evaluate
the efficiency and effectiveness of these vectors we generated dsRNAi lines
for GLABROUS1 (GL1) and the phytoene desaturase gene (PDS) as there
knock-out phenotypes are readily discernable. In addition, dsRNAi constructs
for silencing of EDS1, PAD4 and PEN1 were generated to optimize the timing
of inducer (EtOH) and pathogen challenge. All tested dsRNAi lines were found
to phenocopy heritable mutants of the respective genes. Because phenotypic
stability of gene silencing over several generations is of crucial importance
for our long-term goal, we are currently analyzing T3 and T4 generations of
the dsRNAi lines mentioned above.
T05 Interaction with the Environment 2 (Biotic)