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Plant Signaling & Behavior 7:7, 1–3; July 2012;
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2012 Landes Bioscience
Additional cause for reduced JA-Ile
in the root of a Lotus japonicus phyB mutant
Tamaki Shigeyama,1 Akiyoshi Tominaga,1,2 Susumu Arima,1,2 Tatsuya Sakai,3,4 Sayaka Inada,3 Yusuke Jikumaru,3
Yuji Kamiya,3 Toshiki Uchiumi,5 Mikiko Abe,5 Masatsugu Hashiguchi,6 Ryo Akashi,6 Ann M. Hirsch7 and Akihiro Suzuki1,2,*
1
Department of Environmental Sciences; Faculty of Agriculture; Saga University; Honjyo-machi, Saga, Japan; 2United Graduate School of Agricultural Sciences; Kagoshima University;
Korimoto, Kagoshima, Japan; 3RIKEN Plant Science Center; Yokohama; Kanagawa, Japan; 4Graduate School of Science and Technology; Niigata University; Nishiku, Niigata, Japan;
5
Department of Chemistry and Bioscience; Faculty of Science; Kagoshima University; Korimoto, Kagoshima, Japan; 6Frontier Science Research Center; University of Miyazaki;
Miyazaki; Miyazaki, Japan; 7Department of Molecular; Cell and Developmental Biology and Molecular Biology Institute; University of California-Los Angeles; Los Angeles, CA USA
Keywords: symbiotic nitrogen fixation, shade avoidance syndrome, phytochrome, R/FR ratio, root nodule, nodulation, jasmonic acid
Light is critical for supplying carbon for use in the energetically expensive process of nitrogen-fixing symbiosis between
legumes and rhizobia. We recently showed that root nodule formation in phyB mutants [which have a constitutive shade
avoidance syndrome (SAS) phenotype] was suppressed in white light, and that nodulation in wild-type is controlled by
sensing the R/FR ratio through jasmonic acid (JA) signaling. We concluded that the cause of reduced root nodule formation
in phyB mutants was the inhibition of JA-Ile production in root. Here we show that the shoot JA-Ile level of phyB mutants is
higher than that of the wild-type strain MG20, suggesting that translocation of JA-Ile from shoot to root is impeded in the
mutant. These results indicate that root nodule formation in phyB mutants is suppressed both by decreased JA-Ile
production, caused by reduced JAR1 activity in root, and by reduced JA-Ile translocation from shoot to root.
© 2012 Landes Bioscience.
Light is an important environmental factor controlling plant
growth. It is well known that plants require light for photosynthesis and are able to monitor both light quality and quantity
for optimal survival. Plants have photoreceptors that sense the
presence of their neighbors by monitoring the ratio of red light
(R), which is absorbed by chlorophyll, and far red light (FR),
which is not. A low R/FR ratio indicates the presence of neighbors
that may compete for photosynthetically active radiation (PAR)
and initiates the shade avoidance syndrome (SAS), causing plants
to grow taller or bend to the light to avoid shade.1-4
Many leguminous plants establish a symbiosis with nitrogenfixing soil bacteria called rhizobia. Inside the root nodules,
the rhizobia differentiate into nitrogen-fixing bacteroids. The
bacteroids convert atmospheric nitrogen into ammonia, a source
of fixed nitrogen for the host plant. Because assimilates from
photosynthesis provide energy to fuel the symbiosis between
legumes and rhizobia, the light conditions under which host
plants grow are very important.5-8
Previously we reported that root nodule formation was
suppressed in a Lotus japonicus phytochrome B (phyB) mutant
having a constitutive SAS phenotype.9 In that paper, we
concluded that the cause of reduced root nodule formation in
low-R/FR-grown MG20 (wild-type) plants and white-light-grown
phyB mutants is inhibition of JA-Ile (an active JA derivative)
production in root. By using grafted plants prepared from MG20
and phyB mutant plants, we also showed that shoot genotype
controls root nodule formation.9 Here we report additional data
confirms that root nodulation is controlled by shoot genotype.
The expression level of marker gene NIN,10 which is required for
infection thread formation and nodule primordium initiation, was
analyzed in the root of grafted plants by using real time RT-PCR
by the methods described in Tominaga et al.11 The roots and
shoots of five-day-old MG20 and phyB mutant plants were grafted
in various combinations, as described by Magori et al.12 After 7 d,
the grafted plants were inoculated with M. loti, and expression was
analyzed 7 d after inoculation. Figure 1 shows the relative
expression levels of NIN in roots of grafted plants. The mean
value of expression in grafted MG20(scion)/MG20(root stock)
plants was set as one. When phyB was the scion, both root nodule
number and NIN expression levels were significantly lower than
in the MG20/MG20 control regardless of the rootstock genotype.
When MG20 was used as the scion on a phyB rootstock, no
significant change in N1N expression was observed relative to the
MG20/MG20 control. These results support the hypothesis that
shoot genotype controls root nodule formation.
Furthermore, we previously showed that levels of JAR1 gene
expression and JA-Ile concentration are lower in roots of phyB
than in roots of MG20.9 Because JAR1 codes for an enzyme that
conjugates JA with amino acids to produce the active JA derivative
[most likely jasmonoyl-isoleucine (JA-Ile)],13 we suggested that
inhibition of root nodule formation in phyB mutants is caused by
suppression of the conversion of JA to JA-Ile. To investigate
whether JA-Ile levels decreased throughout the whole plant or
only in root, we measured the endogenous concentrations of JA
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*Correspondence to: Akihiro Suzuki; Email: [email protected]
Submitted: 03/23/12; Revised: 04/13/12; Accepted: 04/16/12
http://dx.doi.org/10.4161/psb.20407
www.landesbioscience.com
Plant Signaling & Behavior
1
observed in JA concentration between MG20 and
phyB mutant plants (Fig. 2A); however, the
concentration of JA-Ile was significantly higher
in the shoot of phyB mutants than in MG20
(Fig. 2B). This result suggested that the translocation of JA-Ile from shoot to root is blocked
and that JA-Ile accumulates in the shoot of phyB
mutants.
Taken together, these results indicate that
decreased JA-Ile concentration caused reduced
root nodule formation in the root of low-R/FRgrown MG20 plants, and that the phenotype of
white-light-grown phyB mutants was produced
both by decreased JAR1 activity in root and by
decreased translocation of JA-Ile from shoots
to roots.
Thus, in wild-type plants exposed to low R/FR
Figure 1. Relative expression of NIN gene in root of grafted plants. The mean value of
light, SAS is triggered by the inactivation of
expression in MG20(scion) / MG20(root stock) was set as 1.0. Transcript amounts were
normalized against ATP synthase (internal control) transcripts. The data represent
PHYB, and root nodule formation is suppressed
the averages ± SE of three independent experiments using roots derived from 3–4
through regulation of the JA-Ile concentration. We
different plants. Statistical significance in comparison to grafted MG20/MG20 is indicated
conclude that this shade avoidance syndrome for
by asterisks (**p , 0.01).
root nodule formation is required for L. japonicus
nodule development and is essential for establishing
and JA-Ile in shoots of white light-grown MG20 and phyB. and maintaining a successful nitrogen-fixing symbiosis.
Fifteen-day-old plants were inoculated with M. loti; after 7 d,
Disclosure of Potential Conflicts of Interest
endogenous JA and JA-Ile concentrations were measured by a
14
previously reported method. No significant difference was No potential conflicts of interest were disclosed.
© 2012 Landes Bioscience.
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Figure 2. Endogenous concentration of JA and JA-Ile in shoots of white light-grown MG20 and phyB. The data represent the averages ± SE of three
independent experiments using shoots derived from six different plants. Statistical significance is indicated by asterisks (*p , 0.05).
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Plant Signaling & Behavior
Volume 7 Issue 7
Acknowledgments
L. japonicus Miyakojima MG20 seeds were provided by the
National BioResource Project of the Ministry of Education,
Culture, Sports, Science and Technology, Japan. This work was
7.
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supported by a Grant-in-Aid for Challenging Exploratory
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© 2012 Landes Bioscience.
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