Nitric oxide and hydrogen peroxide mediated defence responses in Arabidopsis thaliana.
Incompatible plant/pathogen interactions are often manifested as the
hypersensitive response (HR), characterised by host cell death and rapid tissue
collapse at the site of attempted infection. A key early response during the HR is the
generation of reactive oxygen species (ROS), such as the superoxide anion ( 0; -) and
hydrogen peroxide (H20 2), in an oxidative burst. The ROS produced during the
oxidative burst have been implicated as cellular signalling molecules for the induction
of defences responses including hypersensitive cell death. Increasing evidence exist
that the free radical, nitric oxide (NO) also acts as a signalling molecule in plants
during plant/pathogen interactions.
The generation of NO in response to bacterial challenge, and the potential
signalling pathways involved in H20 2- and NO-induced defence responses in
Arabidopsis were therefore investigated
Arabidopsis suspension cultures were found to generate elevated levels of NO
and undergo cell death analogous to HR seen in planta, in response to challenge by
avirulent bacteria. Using NO donors, elevated levels of NO were found to be
sufficient to induce cell death independently of ROS, but not the expression of the
defence-related genes PAL or GST. The NO-induced cell death was sensitive to
inhibitors of RNA processing and protein synthesis, suggesting that NO-induced cell
death is a form of programmed cell death (PCD), requiring the expression of at least
one gene. However, the source of NO production by Arabidopsis remains to be
elucidated, but appears to be independent of nitric oxide synthase-like activity.
Pharmacological studies using specific inhibitors of mammalian mitogen
activated protein kinase (MAPK) signalling cascades, and guanylate cyclase, the
enzyme responsible for the production of second messenger cyclic guanosine
monophosphate (cGMP), suggest that a MAPK signalling cascade acts downstream or
independently of the oxidative burst to initiate H20 2-induced defence responses, while
NO-induced cell death requires the production of cGMP in Arabidopsis.
A number of studies have attempted to establish whether PCD induced during
the HR in plants is similar to apoptotic cell death of anin1al cells. The key
executioners of apoptosis in animal cells are caspases. NO was found to induce
caspase-like activity in Arabidopsis cells, while a specific inhibitor of caspase-l
blocked harpin-, H20 2- and NO-induced cell death. A characteristic of apoptosis is
chromatin condensation and DNA fragmentation into nucleosomal fragments.
Chromatin condensation was observed in Arabidopsis cells treated with the NO donor
Roussin's black salt, but no DNA fragmentation was found in DNA extracted from
cells treated with harpin, H20 2 or NO. In addition, random DNA degradation
indicative of necrosis was found in DNA extracted from cells following avirulent