Discussion
GSTs played the great role in biotic and abioic stress, plant growth and
development, secondary metabolism (Vaish et al. 2020). A soybeanGST gene have been found to increase the resistance
toS. sclerotiorum using GWAS
and linkage mapping (Jianan et al. 2021). To our knowledge, there was no
research about the function study of GSTs in B. napus . In
the study, we found that BnGSTU12 could enhance S.
sclerotiorum resistance inB . napus through
scavenging ROS, and inducing
by
JA signaling pathway.
In our study, we found that BnGSTU12 was located in nuclear and
cytoplasm. GSTs were universe in eukaryotes and prokaryotes and showed
diverse function. GST families have been found to locate in the cytosol,
nucleus, chloroplasts, mitochondria and
ER (Dixon et al. 2009). GSTU12in A. thaliana were found to locate in the nucleus (Dixon et al.
2009). And AtGSTU12 was highly induced by the bacterial elicitor,
flagellin, but is not responsive to H2O2and SA, the key signaling molecules in the hypersensitive response
(Sappl et al. 2009). AtGSTU12 would be unlikely to be involved in
reduce the H2O2, and it might function
to regulate the activity of transcriptional regulators (Sappl et al.
2009).
BnGSTU12
was located in nuclear and cytoplasm. BnGSTU12 was highly induced
by H2O2, JA and SA. The expression
pattern of BnGSTU12 and AtGSTU12 was different. A.
thaliana and B. napus belong to Brassicaceae family, andBrassica and Arabidopsis lineages diverged about 20 MYA.Brassica species have undergone whole genome triplication after
divergence from Arabidopsis (Cheng et al. 2013). Brassica genome
have evolved through extensive genome replication followed by chromosome
fusion and arrangement (Lagercrantz 1998).
The
primary fate of duplicate genes was subfunctionalization,
neofunctionalization and pseudogenization (Birchler and Yang 2022).
Tic40 (for translocon at the inner membrane of chloroplasts with
40 kDa ) genes in Brassica retain the ancestral function similar
with AtTic40, but BnaC9.Tic40 evolved a novel function than can
rescue the fertility due to the amino acid replacements (Dun et al.
2014). In the study, we found that the expression divergence existed
between BnGSTU12 and AtGSTU12 .
Reactive
oxygen species (ROS) including O2-, -OH,
H2O2, the general term for
containing-oxygen molecules, are reactive in the nature. Oxidative burst
is an important defense response for the host during the early stages of
infection through the production of large amounts of ROS (Gonzaleztorres
and Lacruz 2010, del, Rio et al. 2015). ROS, as the signaling molecular,
induced the expression of defense-related genes and cell programmed
death. However, excessive ROS could cause the plant DNA damage and cell
death. In order to
reduce
oxidative damage, antioxidant compounds and enzymes will work to
maintain the steady-state level in plant cells, and then GST could be
used as one of the antioxidase to keep intracellular redox levels by
scavenge reactive oxygen species in plants, which will prevent the
plants from damage (Apel and Hirt 2004; Sonia et al. 2009; GratãO et al.
2005). S. sclerotiorum was the necrotrophic pathogen, which kill
the host cells and then uptake the nutrients. In the study, we found
that BnGSTU12 enhanced the resistance to S. sclerotiorumin B. napus . The GST, SOD, POD and CAT enzyme activities
were
higher in BnGSTU12 -OE lines than WT, and
H2O2 content was less in OE plants.
GSTU12 could be the scavengers of ROS during the hypersensitive
response, which might result in less dead cells and inhibit the growth
ofS.
sclerotiorum .
In the study, JA content increased more in OE-plants than WT after
inoculation with S. sclerotiorum . And the expression of genes
involving in JA signaling and JA-responsive pathway were up-regulated,
showing that these genes contribute to the increasedS.
sclerotiorum resistance of the BnGSTU12 overexpression plants.
JA and SA played the great role in defense response. JA activates
defenses against necrotrophic pathogens, while SA against (hemi-)
biotrophic pathogens (Glazebrook 2005; Pieterse et al. 2012).
Bio-active
JA-isoleucine
(JA-Ile)
was conjugated JA to isoleucine by jasmonoyl-isoleucine synthetase
(JAR). JA-Ile promote the formation of COI1-JAZ complex, and this
molecular interaction leads to proteasomal degradation of JAZ proteins.
The degradation of JAZ induced activity of the defense regulatory
transcription factors and pathogen-response gene plant
defensin1.2 (PDF1.2 ) (Wasternack and Song 2017). In our study,
the expression levels of JAR and PDF1.2 genes in
inoculated OE plants were up-regualted, and JAZ gene was
down-regulated. Improved S. sclerotiorum resistance and GST
enzyme activities were observed in the early stage of inoculated OE
plants, which might feedback activate the JA signaling and JA-responsive
genes. GaGSTF9 in cotton affects endogenous SA levels, not only
SA induces GSTs, but also GSTs can affect SA through the complex reverse
process (Gong et al. 2018). In addition, GSTF2, GSTF8, GSTF10 and GSTF11
were the SA-binding proteins identified by photoaffinity labeling
technique and surface plasmon resonance-based technology (Tian et al.
2012). BnGSTU12 might be the JA-binding protein, which need to be
verified further.
BnGSTU12 played the great role inS.
sclerotiorumresistance
inB. napus through changing the equilibrium of ROS and mediating by
JA signaling pathway. This study not only is important for understanding
the molecular mechanism of S. sclerotiorum resistance, but also
provide the valuable
resistance
breeding strategy in B. napus .