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Plant Physiol, July 2002, Vol. 129, pp. 954-956
SCIENTIFIC CORRESPONDENCE
Nitric Oxide Is Required for Root
Organogenesis1
Gabriela Carolina
Pagnussat,
Marcela
Simontacchi,
Susana
Puntarulo, and
Lorenzo
Lamattina*
Instituto de Investigaciones Biológicas, Facultad de Ciencias
Exactas y Naturales, Universidad Nacional de Mar del Plata, 7600 Mar
del Plata, Argentina (G.C.P., L.L.); and Cátedra de
Físico-Química, Facultad de Farmacia y
Bioquímica, Universidad de Buenos Aires, Junín 956, Buenos Aires, Argentina (M.S., S.P.)
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ARTICLE |
In this report, we
demonstrate that nitric oxide (NO) mediates the auxin response leading
the adventitious root formation. A transient increase in NO
concentration was shown to be required and to be part of the
molecular events involved in adventitious root development induced by
indole acetic acid (IAA).
The discovery of signal molecules involved in the intricate network
that triggers root formation remains a major goal for a large number of
biotechnological procedures. Adventitious rooting involves the
development of a meristematic tissue after removal of the primary root
system. The plant hormone auxins promote this process through the
dedifferentiation of cells to reestablish the new apical meristem.
Although a variety of components of auxin transport and signal
transduction were identified, the molecular mechanism underlying the
initiation of new root meristems is poorly understood (Doerner, 2000 ;
Berleth and Sachs, 2001).
NO is a diffusible multifunctional second messenger first described in
mammals, where it plays variable functions ranging from dilation of
blood vessels to neurotransmission and defense during immune response
(Gow and Ischiropoulos, 2001). Several researches have shown the
presence of NO in plants and have attributed novel roles to this gas in
the plant kingdom (Beligni and Lamattina, 2001a and refs. therein).
Of late, and contemporary to genomics and proteomics, it is
interesting to note the revival of pharmacological and surgical techniques in the field of plant developmental biology (Nemhauser et
al., 2000; Reinhardt et al., 2000). In this communication, we
demonstrate through pharmacological and surgical approaches that NO is
required for root organogenesis.
Two NO donors, sodium-nitroprussiate (SNP) and S-nitroso,
N-acetyl penicillamine (SNAP), applied to hypocotyl cuttings
(primary roots removed) of cucumber (Cucumis sativus) were
able to mimic the effect of the auxin IAA in inducing de novo root
organogenesis (Fig. 1). In addition, NO-
and IAA-induced roots presented similar anatomic structure when
they were analyzed by optic microscopy (not shown). This NO-mediated
effect was prevented when the specific NO-scavenger carboxy-PTIO
(cPTIO) was added with SNP or SNAP (Fig. 1). Result of treatments
performed with different SNP concentrations confirmed that the effect
was dose dependent, with a maximal biological response at 10 µM SNP (Fig. 2).
Within 3 d after removal of the primary root system, adventitious
root development was detected in the explants treated with IAA, SNP, or
SNAP. Two parameters of root growth were considered, and length and
number of adventitious roots exhibit similar behavior among these
treatments (Fig. 3, t test,
P < 0.05). In control experiments, when hypocotyl
cuttings were kept in water or in
NO2 /NO3
(normal products of NO decomposition, not shown), adventitious roots
emerged 4 d after primary root removal, and they reached only 22%
of the length obtained from NO- or IAA-treated explants (Fig. 3). The
treatment of hypocotyls with SNP or SNAP plus IAA resulted in an
increased response, displaying roots longer than those from hypocotyls
treated with the NO donors or IAA alone in a factor of 1.6 (Figs. 1 and
3). The specific NO scavenger, cPTIO, delayed adventitious root
emergency and significantly reduced the root length and number of the
IAA-treated explants (Figs. 1 and 3, t test,
P < 0.05). It is interesting that this inhibitory effect of cPTIO was reversible because adventitious root emergency was
triggered by the addition of 200 µM SNP after
cPTIO treatment (not shown). In addition, a direct inactivation of IAA
by cPTIO per se may be discarded because a mixture of IAA plus cPTIO
was able to induce epinasty, an IAA-mediated effect, at the same extent as IAA alone (not shown).
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Figure 1.
NO donors induce adventitious root development in
cucumber explants. The primary root system was removed from hypocotyls
of 10-d-old germinated cucumbers. Explants were incubated with water,
10 µM each of NO donors (SNP or SNAP), NO donors plus 200 µM cPTIO, or 10 µM IAA. The treatments were
also done with a combination of IAA with SNP, SNAP, or the NO scavenger
cPTIO. Photographs were taken after 5 d of treatments. Bar = 5 mm.
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Figure 2.
The effect of the NO donor on the induction of
adventitious root formation is dose dependent. The Primary root system
was removed from hypocotyls of 10-d-old germinated cucumbers. Explants
were incubated with water or different concentrations of SNP as
indicated. Adventitious root numbers were quantified after 5 d of
treatment and are expressed as mean ± SE
(n = 15 explants from at least three independent
experiments).
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Figure 3.
NO effect on adventitious root number and length
in cucumber explants. Ten-day-old germinated cucumber seedlings were
treated as described in Figure 1. Root number (A) and length (B) values
are expressed as mean ± SE (n = 20 explants from at least three independent experiments). Bars with
different letters are significantly different with P < 0.05 (t test).
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Previous results reported by Gouvêa et al. (1997) showed that
NO-releasing substances induced root tip elongation. Although a nitric
scavenger prevented the growth induced by NO-releasing substances, it
had no effect on IAA-induced cell expansion (Gouvêa et al.,
1997). In contrast, our results using the specific NO scavenger, cPTIO,
strongly indicated a role of endogenous NO in IAA-mediated root
organogenesis. Consequently, the endogenous NO level was determined by
electron paramagnetic resonance (EPR) in explants treated with IAA or
water (control). High levels of NO were detected at
T0 in both treatments, probably reflecting a
response to wounding after removing the primary root. However, a
transient increase of NO (60 nmol g1 fresh
weight) could be measured in explants after 24 h of treatment with
IAA. NO concentration remained at detectable levels (20 nmol g1 fresh weight) until 96 h of treatment.
On the contrary, in control explants, the level of endogenous NO
dramatically shut down at 24 h (Fig.
4A). We also detected the presence of
endogenous NO by using a fluorescent probe. Explants were exposed to
the permeable and specific NO-sensitive fluorophore
4,5-diamino-fluorescein diacetate, which allows the detection of NO
presence in animal and plant cells (Kojima et al., 1998; Foissner et
al., 2000). The IAA-treated explants displayed 4-fold more fluorescence
than control explants (medium pixel intensity: 138 versus 39; Fig. 4, B
and C, respectively).
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Figure 4.
IAA induces a transient increase of endogenous NO
in cucumber explants. Endogenous NO level was determined by EPR (A; MGD
kit M-0010; OMRF SPIN TRAP SOURCE, Oklahoma City, OK) in
explants treated with 10 µM IAA (red circles) or water
(blue circles). Inset, EPR profile obtained from IAA- and water-treated
explants after 24 h of incubation. B and C,
4,5-Diamino-fluorescein diacetate fluorescence detected in a
longitudinal section from the tip of the hypocotyls, where new
meristematic tissue and adventitious roots are formed. Pictures were
taken after 24 h of treatment from an IAA-treated explant (B) and
an untreated one (C). Bars = 0.5 mm.
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It still remains to be elucidated if NO alone has the capability to
trigger root formation in the complete absence of IAA. Preliminary
results in IAA-depleted cucumber hypocotyls showed that they were able
to induce the adventitious root formation after treatment with NO
donors (G. Pagnussat, L. Lanteri, and L. Lamattina, unpublished
data). However, further investigations are needed to conclusively
confirm these observations.
Although a direct effect of NO cannot be discarded, our
findings indicate that NO could mediate the auxin response during the
adventitious rooting process in cucumber. Nevertheless, a serial
linkage IAA  NO rooting cannot be distinguished from a scenario
in which IAA and NO could be acting in parallel on a third
intermediate. The involvement of NO in this auxin-signaling pathway
opens a wide field of research for every reported IAA effect in plant
biology. Considering that NO has been postulated as a signal molecule
during development and adaptive plant responses (Klessig et al., 2000;
Beligni and Lamattina, 2001b; Garcia Mata and Lamattina, 2001), our
results strongly support the idea of NO as a versatile molecule with
variable functions in plants, as occurs in animal systems.
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FOOTNOTES |
Received February 12, 2002; returned for revision March 11, 2002; accepted April 5, 2002.
1
This work was supported by Agencia Nacional de
Promoción Científica y Tecnológica (grant no.
PICT 1-6496-99 to L.L.), by Consejo Nacional de Investigaciones
Científicas y Técnicas (grant no. PIP 0898/98 to L.L.),
and by institutional grants from Universidad Nacional de Mar del
Plata. G.C.P. is a postdoctoral fellow, and L.L. is member of
the research career of Consejo Nacional de Investigaciones Científicas y Técnicas.
*
Corresponding author; e-mail lolama{at}mdp.edu.ar; fax
54-223-475-3150.
www.plantphysiol.org/cgi/doi/10.1104/pp.004036.
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LITERATURE CITED |
-
Beligni MV, Lamattina L
(2001a)
Plant Cell Environ
24: 267-278[CrossRef]
-
Beligni MV, Lamattina L
(2001b)
Trends Plant Sci
6: 508-509[CrossRef][ISI][Medline]
-
Berleth T, Sachs T
(2001)
Curr Opin Plant Biol
4: 57-62[CrossRef][ISI][Medline]
-
Doerner P
(2000)
Curr Biol
10: 201-303
-
Foissner I, Wendehenne D, Langebartels C, Durner J
(2000)
Plant J
23: 817-824[CrossRef][ISI][Medline]
-
Garcia Mata C, Lamattina L
(2001)
Plant Physiol
126: 1196-1204[Abstract/Free Full Text]
-
Gouvêa CMCP, Souza JF, Magalhães, Martins IS
(1997)
Plant Growth Regul
21: 183-187
-
Gow AJ, Ischiropoulos HJ
(2001)
Cell Physiol
187: 277-282
-
Klessig DF, Durner J, Noad R, Navarre DA, Wendehenne D, Kumar D, Zhou JM, Shah J, Zhang S, Kachroo P, et al
(2000)
Proc Natl Acad Sci USA
97: 8849-8855[Abstract/Free Full Text]
-
Kojima H, Nakatsubo N, Kikuchi K, Urano Y, Higuchi T, Tanaka J, Kudo Y, Nagano T
(1998)
Neuroreport
9: 3345-3348[ISI][Medline]
-
Nemhauser JL, Feldman LJ, Zambryski PC
(2000)
Development
127: 3877-3888[Abstract]
-
Reinhardt D, Mandel T, Kuhlemeier C
(2000)
Plant Cell
12: 507-518[Abstract/Free Full Text]
© 2002 American Society of Plant Physiologists
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LITERATURE CITED