Plant Molecular Biology 36: 673–680, 1998.
1998 Kluwer Academic Publishers. Printed in Belgium.
High-level expression of a viscotoxin in Arabidopsis thaliana gives enhanced
resistance against Plasmodiophora brassicae
, Jutta Ludwig-M
and Holger Bohlmann
Swiss Federal Institute of Technology (ETH), Institute of Plant Sciences, ETH-Zentrum, Universit
LFW D.58, 8092 Z
urich, Switzerland (
author for correspondence);
Johann Wolfgang Goethe-Universit
Botanisches Institut, Postfach 111932, 60054 Frankfurt/Main, Germany;
Present address: European Patent
Ofﬁce. Directorate 1.2.12, Chemistry, Postbus 5818, 2280 HV Rijswijk (ZH), Netherlands
Received 18 March 1977; accepted in revised form 13 October 1997
Key words: plant defence, thionins, cysteine-rich, antimicrobial
Viscotoxins are a group of toxic thionins found in several mistletoe species. The constitutive CaMV-
wasused to drive the expression of the viscotoxin A3 cDNA from Viscum album in transgenic Arabidopsis thaliana
C24. Lines with high viscotoxin A3 levels in all parts of the plant were selected and tested for resistance against
the clubroot pathogen Plasmodiophora brassicae. The transgenic lines were more resistant to infection by this
pathogen than the parental line.
Several groups of basic and cysteine-rich polypeptides
in the defence against phytopathogens. These include
thionins [for review see 8, 10, 29], lipid transfer pro-
teins , and plant defensins . The toxic activit-
ies of thionins have been studied for several decades.
Soonaftertheisolation ofpurothioninfromwheat ﬂour
, it was discovered that it has antimicrobial activ-
ity . Such antimicrobial activity has since been
demonstrated for several other thionins [11, 25, 26,
44, 65, 67], supportingthe suggestion that thionins are
defenceproteins . Further evidence for such a pos-
sionofseveralthioningenes is induciblebyphytopath-
ogenic fungi [11, 12, 20, 68, 69]. Furthermore, it has
been shown that the expression of
transgenic tobacco plants can lead to enhanced res-
istance against a phytopathogenic bacterium . In
similar experiments, however, Florack et al.  did
expressing a synthetic
We have recently shown that an Arabidopsis thali-
ana thionin gene is inducible by pathogenic fungi via
a signal transduction pathway different from that for
PR proteins . The induction of this Thi2.1 gene by
Fusariumoxysporumf.sp. matthiolae ismuchstronger
in resistant than in susceptible ecotypes (Epple, Vign-
utelli, Apel and Bohlmann, manuscript submitted).
Finally, overexpression of the THI2.1 thionin in the
susceptible ecotype Col-2 leads to higher resistance
against F. oxysporum f. sp. matthiolae .
Viscotoxins are a group of thionins which have
been isolated and sequenced from different mistletoe
species [52, 72]. Mistletoe extracts are being used in
the treatment of cancer  and as antihypertensive.
[e.g. 2, 17, 39, 50]. The toxicity of viscotoxins and
other thionins has been attributed to a damaging effect
on membranes [e.g. 15, 17, 39]. Like other thionins,
viscotoxins are also produced as preproproteinswith a
molecular mass of about 15 kDa [8, 10, 29].
Four viscotoxin isoforms with six cysteine residues
53, 54, 55]. Additional new viscotoxin variants were
identiﬁed by a PCR-based approach . The thionin
domains encoded by two of these cDNAs have eight
cysteine residues, in contrast to six in the classical vis-