Following pathogen recognition, nitric oxide (NO) is rapidly produced in plants, this
small molecule has emerged as a key signal in plant defence responses. S-nitrosylation
is the major route of NO signal transduction in plants, a redox-based
modification by addition of an NO moiety on cysteine thiol to form an S-nitrosothiol
(SNO). S-nitrosoglutathione reductase (GSNOR) regulates cellular levels of S-nitrosylation
and displays a key role in regulating the plant defence response. In this
context, NO is important to orchestrate both defence gene expression and the
hypersensitive response (HR) during attempted microbial infection. However, how
the plant immune system recognizes NO and how NO level could elicit plant defence
responses are poorly understood.
The Arabidopsis thaliana (Arabidopsis) mutant NO overproducing 1 (nox1) was
employed to characterize how NO level elicits defence dynamics. In response to
microbial infection, resistance (R) gene-mediated defence and basal resistance were
found to be compromised in the nox1 mutant relative to wild type Col-0 plants.
Interestingly, nox1 mutant exhibit similar levels of HR and pathogen susceptibility to
the GSNOR loss-of-function mutant atgsnor1-3. This phenomenon suggests that NO
might regulate defence responses via GSNOR-mediated S-nitrosylation. Therefore,
the nox1 atgsnor1-3 double mutant was generated and characterized to clarify this
hypothesis. Accelerated HR and increased pathogen susceptibility are shown in the
double mutant, which implies that increased NO mediated by nox1 and elevated
SNOs resulting from atgsnor1-3, are additive with respect to the plant defence
response.
To identify genes responsible for NO perception, forward genetic screens were
developed to identify Arabidopsis mutants with abnormal NO recognition. NO
marker genes for genetic screens were identified from both lab and open source
microarray data. Two genes, At3g28740 and At1g76600 were selected and
experimentally confirmed to be strongly induced by NO. Transgenic Arabidopsis
plants were generated carrying a NO reporter cassette, which consist of a luciferase
reporter gene (LUC) driven by the promoter of NO marker gene. This forward
genetic approach might be a powerful tool to identify genes integral to NO signal
transduction.
Three C2H2 zinc finger transcription factors (ZnTFs) ZAT7, ZAT8 and ZAT12 were
identified as being rapidly and strongly induced by NO donors, which could be
modulators of redox/NO-dependent signalling pathway. T-DNA insertion mutants
within these ZnTFs have been identified. Basal resistance against Pseudomonas
syringae pv tomato (Pst) DC3000 is compromised in all single knockout lines.
Therefore, the full characterisation of defence phenotype of these mutants would be
necessary to explore the role of these TFs in the plant defence. Furthermore, zat8
mutant is more sensitive to nitrosative stress when compared to wild type Col-0. This
suggests that ZAT8 may be involved in protecting plants against nitrosative stress.
However, the molecular mechanisms that underpin this function remain to be
determined.
In conclusion, NO and SNOs might regulate plant disease resistance via distinct
pathways. Our work has also established NO-reporter lines to identify genes
responsible for NO perception. In addition, three NO-induced ZnTFs have been
identified that participate in regulation of basal resistance, which might unveil
aspects of NO signalling related to the regulation of transcription