Targets of the WRKY53 transcription factor and its role during leaf
senescence in Arabidopsis
Y. Miao, T. Laun, P. Zimmermann and U. Zentgraf *
ZMBP (Centre of Molecular Biology of Plants), Department of General Genetics, University Tu
¨
bingen, Auf
der Morgenstelle 28, 72076 Tu
¨
bingen, Germany (*author for correspondence; email ulrike.zentgraf@uni-
tuebingen.de)
Received 23 April 2004; accepted in revised form 13 August 2004
Key words: catalases, hydrogen peroxide, leaf senescence, target genes, WRKY transcription factors
Abstract
Arabidopsis WRKY proteins comprise a family of plant specific zinc-finger-type transcription factors
involved in the regulation of gene expression during pathogen defense, wounding, trichome development,
and senescence. To understand the regulatory role of the senescence-related WRKY53 factor, we identified
target genes of this transcription factor by a pull down assay using genomic DNA and recombinant
WRKY53 protein. We isolated a number of candidate target genes including other transcription factors,
also of the WRKY family, stress- and defence related genes, and senescence-associated genes (SAGs).
WRKY53 protein could bind to these different promoters in vitro and in vivo and it could act either as
transcriptional activator or transcriptional repressor depending on the sequences surrounding the W-boxes.
Overexpression, RNAi and knock-out lines showed accelerated and delayed senescence phenotypes,
respectively, and exhibited altered expression levels of the target genes. WRKY53 can be induced by H
2
O
2
and can regulate its own expression in a negative feed back loop. Our results suggest that WRKY53 acts in a
complex transcription factor signalling network regulating senescence specific gene expression and that
hydrogen peroxide might be involved in signal transduction.
Introduction
Senescence is not a chaotic breakdown but an
orderly loss of normal cell functions which is
under the control of the nucleus. Bleaching of
leaves, loss of chlorophyll and changes in total
protein and RNA content are some of the first
obvious signs and measurable parameters of leaf
senescence. These parameters have been examined
throughout the life cycle of Arabidopsis to char-
acterize the molecular mechanisms involved in this
process (Hensel et al., 1993; Lohman et al., 1994).
Specific sets of genes are down-regulated during
senescence (designated senescence-down-regulated
genes; SDGs), including photosynthesis related
genes. Different sets of genes, designated senes-
cence-associated genes (SAGs) are up-regulated
during senescence and encode proteins involved in
the breakdown of cellular components, including
nucleases, proteases, and cell wall hydrolases.
Other gene products are associated with the
mobilization of nutrients and minerals like fatty
acids, nitrogen compounds or phosphate out of
the senescing tissue in developing parts of the
plants (reviewed in Bleecker and Patterson, 1997;
Buchanan-Wollaston, 1997; Hadfield and Bennett,
1997; Nam, 1997; Noode
´
n et al., 1997; Biswal and
Biswal, 1999; Quirino et al., 2000, Buchanan-
Wollaston et al., 2003). The ability of SAG pro-
moters to direct senescence specific expression of
the GUS reporter gene revealed that SAGs appear
to be regulated mainly on the transcriptional level
(Hanfrey et al., 1996; Oh et al., 1996; Butt et al.,
1998; Noh and Amasino, 1999).
Plant Molecular Biology 55: 853–867, 2004.
Ó 2004 Kluwer Academic Publishers. Printed in the Netherlands.
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