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Plant Physiol, March 2000, Vol. 122, pp. 925-932
Auxin Metabolism in the Root Apical Meristem1
Nancy M.
Kerk,2
Keni
Jiang, and
Lewis J.
Feldman*
Department of Plant and Microbial Biology, 111 Koshland Hall,
University of California, Berkeley, California 94720-3102
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ABSTRACT |
Within
the root meristem of flowering plants is a group of mitotically
inactive cells designated the quiescent center (QC). Recent work links
the quiescent state to high levels of the growth regulator auxin that
accumulates in the QC via polar transport. This in turn results in
elevated levels of the enzyme ascorbic acid oxidase (AAO), resulting in
a reduction of ascorbic acid (AA) within the QC and mitotic quiescence.
We present evidence for additional interactions between auxin, AAO, and
AA, and report that, in vitro, AAO oxidatively decarboxylates auxin,
suggesting a mechanism for regulating auxin levels within the QC. We
also report that oxidative decarboxylation occurs at the root tip and that an intact root cap must be present for this metabolic event to
occur. Finally, we consider how interaction between auxin and AAO may
influence root development by regulating the formation of the QC.
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INTRODUCTION |
It has long been known that auxin is transported from the shoot to
the root in a polar manner (Scott and Wilkins, 1968 ; Morris et al.,
1969). However, little attention has been given to the fate of the
auxin accumulating in the root tip. Since root cells are significantly
more sensitive to auxin concentrations than are cells of the shoot
(Thimann, 1937), it seems unlikely that high concentrations of auxin
could be tolerated in a growing root tip.
Our previous work has provided evidence for polar transport of
indole-3-acetic acid (IAA) and its accumulation in maize (Zea mays) root tips (Kerk and Feldman, 1994, 1995). We showed that this resulted in an increased level of ascorbic acid oxidase (AAO) mRNA, protein, and enzyme activity, as well as the localized depletion of ascorbic acid (AA) within the quiescent center (QC), a group of
mitotically inactive cells within the root meristem. Since AA is
necessary for the G1 to S transition in the cell cycle in root tips
(Liso et al., 1988), and regulation of AA is thought to be dependent on
AAO, we proposed that its depletion in root tips may be responsible for
the formation and maintenance of the QC (Kerk and Feldman, 1995).
In the course of these experiments we discovered additional
interactions between auxin and AAO. Using the radish (Raphanus sativus) root auxin bioassay system (Blakely et al., 1982), we found that increased levels of AAO in root segments in sterile culture
decreased the response of radish roots to auxin. Moreover, if auxin is
first reacted in vitro with AAO and then bioassayed, root cultures fail
to exhibit a normal response.
Finally, we report on a possible mechanism underlying these auxin
responses. We report here that AAO oxidatively decarboxylates auxin in
vitro, suggesting a mechanism for regulating auxin levels within the QC
and other root tissues. Oxidative decarboxylation occurs at the root
tip, and an intact root cap must be present for this metabolic event to
occur. These observations provide a robust model for the organization
and functioning of the maize root tip, in which auxin transported from
the shoot both regulates and is regulated by the level of AAO, which
then regulates entry into the S phase by controlling levels of AA.
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MATERIALS AND METHODS |
Plant Growth Conditions
Maize (Zea mays var Merit, Asgrow Seed Co., Kalamazoo,
MI) caryopses were imbibed and germinated in the dark at 25°C for
2 d. Bioassays were carried out on 3-d-old radish (Raphanus
sativus Scarlet Globe, Asgrow Seed Co., Kalamazoo, MI) roots.
Radish seeds were surface-sterilized and germinated aseptically in the
dark at 25°C for 3 d. The terminal 2- to 5-mm tips were removed
and 1-cm segments were placed into liquid culture. Medium was complete Murashige and Skoog (MS) medium (Murashige and Skoog, 1962) with added CuSO4, auxin, or auxin previously reacted
with AAO, as reported in "Results." AAO activity was assayed as
described in Kerk and Feldman (1995).
In Vitro Auxin Metabolism
In a total volume of 0.5 mL, 10 7 to
108 M IAA (Sigma-Aldrich,
St. Louis) ± 5-3H-IAA and/or
1-14C-IAA were incubated with 50 µM
p-coumaric acid, 100 µM
Mn2+, and 25 units of AAO (Biozyme Labs, San
Diego) in a 7 mM Na-phosphate buffer, pH 5.3, at
27°C in the dark on a shaker for 1 to 4 h. For assessing
decarboxylation activity, aliquots were periodically removed from the
incubation mixture and counted on a scintillation counter set (model
60001C, Beckman Instruments, Fullerton, CA) to separately window
3H and 14C. To determine
the degree to which this reaction is mediated by a copper-containing
oxidase (such as AAO), we also performed the incubation in the presence
of 1 to 5 mM bathocuproinedisulfonic acid
(Sigma-Aldrich), a compound with high specificity for inhibiting copper-containing enzymes (Li et al., 1996), and in 1 mM diethyldithiocarbamate (DDC) (Sigma-Aldrich),
another copper-chelating agent and inhibitor of AAO (Wang and Fuast,
1992).
For HPLC analysis, the incubation mixture was acidified to pH 3.0, extracted three times against ethyl acetate, dried, solubilized in a
small amount of methanol, and subjected to HPLC analysis on a HPLC
(Kratos, Chestnut Ridge, NY) using a reversed phase small pore
silica C18 10-µm column (4.6 mm × 25 cm; Vydac, Hesperia, CA) and a 40-min linear gradient of 5%
to 30% (v/v) acetonitrile in 0.08% (v/v)
trifluoroacetic acid with a flow rate of 0.7 mL/min; fractions were
collected every 30 s. Elution profiles were obtained by monitoring
at 254 nm, and fractions representing products of IAA catabolism were
collected, characterized, and counted as described above. For each run
typically 90% of the injected radioactivity was recovered. After
identifying the radioactive elution profiles of the IAA metabolites, we
scaled up the reaction 10× and repeated the previously described
incubation but instead used non-radioactive auxin. The products were
then separated by HPLC and the two fractions corresponding to the
radiolabeled IAA metabolites collected and subjected to
diffuse-reflectance Fourier transform infrared (IR) spectroscopy. In
preparation for IR analysis, each HPLC fraction was dried down under a
stream of N2 and the residue was dissolved in 20 µL of CH2Cl2. The
solution was applied to powdered IR-grade potassium bromide in a 3-mm
microsampling cup and the IR spectrum (32 scans) obtained after
evaporation of the CH2Cl2.
The same HPLC fractions were also examined using UV spectroscopic analysis.
In Vivo Auxin Metabolism
For monitoring in vivo auxin metabolism the terminal 2 cm of roots
from aseptically grown maize seedlings were excised from 48-h-old
seedlings and placed tip down into microfuge tubes containing 0.5 mL of
one-half-strength MS medium, pH 6.8, supplemented with 1% Suc
and 0.9% agar. For some experiments roots were decapped (Kerk
and Feldman, 1995) prior to excising the terminal 2 cm, which, as
before, were placed tip down into the microfuge tubes. On the basal cut
surface (the surface protruding from the tube) was placed a 1%
agar block (approximately 1 mm2)
containing 1 × 109 M
non-radioactive IAA, plus 108 M
5-3H-IAA (specific activity, 16.7 Ci/mM; Amersham) and/or 1-14C-IAA
(specific activity, 9.4 mCi/mM; Sigma). The roots with the attached agar blocks were returned to a moistened chamber and incubated
for 12 h in the dark. For most experiments 40 roots were used.
Following incubation the root was divided into three sections: the
distal terminal millimeter, the subtending 1 cm (root minus terminal
millimeter), and the 1- to 2-cm section contacting the agar block. The
terminal millimeter and subtending 1-cm sections of roots were
harvested separately, pooled, homogenized, extracted in 80%
(v/v) methanol (McDougall and Hillman, 1978), and subjected to
HPLC analysis as described above. Fractions were collected every
30 s. To determine whether any decarboxylation occurred, 14C- and 3H-IAA in a known
ratio were together incubated with root tissues. At the end of the 12-h
incubation period, the ratios of 14C- and
3H-IAA were compared for each fraction using a
scintillation counter set to separately window 3H
and 14C, and the counts were normalized for any
change in the ratio (indicating a loss of 14C).
14C-3H ratios were
calculated after setting the background to 50 dpm.
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RESULTS |
Bioassay Results
Response of Cultured Roots to Exogenous IAA
To establish a baseline for experiments involving the interaction
between IAA and AAO, we examined the effect of supplying IAA in vitro
to radish root segments. Roots exposed to a range of concentrations of
IAA initiated increasing numbers of lateral roots, but the outgrowth of
these roots was progressively inhibited (Fig.
1A). This result confirms the work of
previous investigators (Blakely et al., 1982). Furthermore, we found
that when roots that had produced laterals in response to a particular
concentration of exogenous auxin were subsequently exposed to a higher
concentration of IAA, numerous supernumerary lateral roots spaced
between existing ones were formed (Fig. 1B).
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Figure 1.
Radish root segments cultured with IAA. A, Root
segments cultured for 48 h in MS medium containing from left to
right, 0, 1, 3, 10, and 30 µM IAA. Scale bar increments
indicate 1 mm. B, Root segment cultured for 24 h in MS plus 3 µM IAA and then shifted to MS plus 30 µM
IAA for 48 h. Scale bar increments indicate 0.5 mm.
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Response of Cultured Roots to Increased AAO
Esaka et al. (1992) have demonstrated a marked increase in
ascorbate oxidase protein in pumpkin cells by adding copper to the
culture medium. We used this method to cause a 1.7-fold increase in AAO
in seedling radish roots grown in culture supplemented with 10 µM CuSO4. We also measured the
response of these roots to auxin by determining the lateral root
frequency, which increases with auxin concentration (Blakely et al.,
1982; Kerk, 1990) (Fig. 1A).
When auxin was added to the culture medium simultaneously with copper,
there was no diminution in the number of lateral roots (Fig.
2, A and B). However, when roots were
preincubated in a copper-containing culture medium for 24 h before
auxin was added, there was a significant decrease in the number of
lateral roots produced, yet these roots were healthy and showed no
toxic effect of the copper (Fig. 2C). This suggests that levels of AAO
in root tissues affect the auxin response.
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Figure 2.
Radish root segments cultured in MS medium with 3 µM IAA only for 72 h (A); with 3 µM
IAA and 10 µM CuSO4 added simultaneously for
72 h (B); roots incubated first for 24 h with 10 µM CuSO4 and then 3 µM IAA was
added for a further 48 h (C).
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In Vitro Reactions of IAA and AAO as Visualized by the Bioassay
Experiments were carried out to determine if AAO could directly
react with auxin as a substrate. In vitro reactions were set up
containing auxin and AAO in buffers and at concentrations comparable to
those found at physiological conditions. The reaction mixtures were
filter-sterilized and used as the auxin source for bioassays. Reaction
mixtures with increasing units of AAO resulted in a marked decrease in
the number of lateral roots and failed to inhibit tip growth, as would
be expected in a standard auxin bioassay in roots (Fig.
3, A-C). When the AAO was denatured by
boiling before being reacted with IAA in the buffer, lateral root
formation was similar to that in roots treated with IAA alone (Fig.
3D).
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Figure 3.
Radish root segments cultured in MS and IAA, or
IAA prereacted with AAO and then filter-sterilized and added to
cultures. A, Effect of increasing concentrations of IAA. As the
concentration is increased, tip growth becomes inhibited and lateral
root frequency increases. B, Effects of IAA prereacted with increasing
amounts of AAO. As AAO units are increased, inhibition of tip growth
and lateral root frequency are decreased. C, Representative repetition
of the effect on roots in culture with IAA pretreated with 5 units of
AAO. The original tip region of the root before elongation growth in
culture is marked by the swollen cortical region located proximally
about one-third the distance to the present tip. D, Radish root
segments cultured with 3 µM IAA reacted with 5 units of
AAO previously boiled to denature the protein.
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Tips of Lateral Root Primordia Lower the Effective Auxin Levels in
Roots in Culture
Roots were cultured in 3 µM IAA. The tips of the
lateral roots that developed in response to IAA treatment were excised
and the roots were returned to the same culture medium. The roots with
decapitated tips formed a large number of supernumerary laterals spaced
between the stumps of the pre-existing laterals. In control roots in
which needle punctures were made in the cortical region but tips
were left intact (to simulate the wounding effect of decapitation), no
supernumerary laterals were formed (Fig.
4).
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Figure 4.
Effect of removing lateral root tips on new
lateral root initiation. A, Root cultured for 5 d in MS plus 3 µM IAA. B, Root grown as in A, but after 5 d lateral
root tips were clipped off, leaving stumps, and placed back in 3 µM IAA medium. New laterals were initiated at high
frequency on the parent root.
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Biochemical Results
The bioassay experiments described in the preceding sections have
provided evidence that AAO and auxin interact to decrease the effective
auxin concentration in roots. To investigate this reaction, its
products, and the likely reaction mechanism, we examined HPLC profiles
of reactions carried out with AAO and radioactively labeled auxin.
We identified two major metabolic products of the reaction that had a
pH optimum of 5.3 (Fig. 5). The reaction
was completely inhibited when DDC was added to the reaction mix (Fig.
5). DDC is an inhibitor of AAO that chelates copper away from this blue copper protein, permitting the subunits to dissociate (Wang and Fuast,
1992). The addition of ascorbic acid, dehydroascorbate, and ascorbate
free radical to reaction mixes with or without AAO had no effect on
auxin metabolism (data not shown).
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Figure 5.
HPLC profiles of the reaction products of AAO and
3H-IAA. The pH optimum was pH 5.3 and the inclusion of DDC,
an inhibitor of AAO, completely blocked the reaction. Peak 2, IAA.
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HPLC Elution Profiles of in Vitro 14C-IAA and/or
3H-IAA Catabolism
IAA is oxidatively decarboxylated in vitro by AAO, forming as one
of the primary products oxindole-3-methanol (Figs.
6 and 7).
Identification of this compound was based on its IR spectrum (diffuse
reflectance on powdered KBr) (3,397 cm1 [OH],
3,207 cm1 [NH], 1,704 cm1, and 1,621 cm1
[C = O]) and on its UV spectrum (in methanol,
 max = 250 nm with a shoulder at 280 nm)
(Hinman and Lang, 1965; Kobayashi et al., 1984). Additional support for
this identification was obtained from the non-biological conversion of
oxindole-3-methanol to a compound identified as 3-methyleneoxindole,
based on its twin UV absorption peaks at max
248 and 252 nm (Hinman and Lang, 1965). This in vitro activity is
dependent on the addition of Mn2+ and
p-coumaric acid (Fig. 6). The addition of
bathocuproinedisulfonic acid, a compound with high specificity for
inhibiting copper-containing enzymes, resulted in a decrease in the
decarboxylation of IAA, suggesting that the decarboxylating activity
was due to a copper-containing enzyme such as AAO (Li et al., 1996)
(Table I).
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Figure 6.
AAO facilitates IAA decarboxylation. Oxidative
decarboxylation of 1-14C-IAA, measured as a loss of
14CO2. Data reflect the amount of radioactivity
in a 75-µL aliquot after various periods of incubation with and
without AAO and with and without co-factors (Mn2+ and
p-coumaric acid).
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Figure 7.
HPLC elution profiles of in vitro
14C-IAA and/or 3H-IAA catabolism. A,
5-3H-IAA plus Mn2+ plus
p-coumaric acid. B, 5-3H-IAA plus
Mn2+ plus p-coumaric acid plus AAO. C,
1-14C-IAA plus Mn2+ plus
p-coumaric acid. D, 1-14C-IAA plus AAO plus
Mn2+ plus p-coumaric acid. Peak 1, Oxindole-3-methanol, the primary in vitro catabolic product; peak 2, IAA; peak 3, 3-methyleneoxindole, a non-biological breakdown product of
peak 1.
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Table I.
Effects of an inhibitor of copper-containing
oxidases on IAA decarboxylation
Effect of bathocuproinedisulfonic acid (BA), an inhibitor of
copper-containing enzymes, on the oxidative decarboxylation of
1-14C-IAA in the presence of AAO, Mn+2, and
p-coumaric acid. Data reflect the amount of radioactivity in
a 75 µL-aliquot after various periods of incubation with and without
BA.
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HPLC Elution Profiles of in Vivo 14C-IAA and/or
3H-IAA Catabolism
At least 30% of polarly transported auxin is oxidatively
decarboxylated in vivo by intact root tips (terminal millimeter) (Fig.
8). Oxidative decarboxylation in the root
occurs almost exclusively at the tip (Fig. 8). Excision of the cap
prevents decarboxylation (Fig. 8).
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Figure 8.
HPLC elution profiles of 1-14C-IAA
metabolites in various maize root tissues. Black portions for each
curve represent the actual recoverable 14C counts in each
fraction after a 12-h incubation. The white portions under the curve
represent the 14C counts that would have been recovered had
there not been any decarboxylation. To determine which fractions show
decarboxylation, 1-14C- and 5-3H-IAA in a known
ratio were together incubated with root tissues. At the end of the 12-h
incubation period the ratios of 14C- and 3H-IAA
were compared for each fraction and the curve normalized for any change
in the ratio (indicating a loss of 14C) (white portion of
the curve). The only significant changes in the ratio are in IAA
metabolites in intact terminal millimeter root tissues. For all other
treatments the ratios are more or less unchanged and the white and
black curves overlap. Note that all root tissues completely catabolize
the radiolabeled IAA.
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DISCUSSION |
The major findings reported here suggest that: (a) elevated levels
of AAO in roots are correlated with a decreased auxin response, as
visualized by bioassays in radish root culture; and (b) the decreased
auxin response may be due to the fact that AAO oxidatively decarboxylates auxin. This reaction renders the reacted auxin ineffective in stimulating an auxin response in root cultures. Support
for the likely occurrence of this reaction was demonstrated by in vitro
experiments and in vivo through bioassays for auxin response and by
recovering metabolites of radiolabeled auxin generated through
oxidative decarboxylation in intact maize root tips. We suggest that
AAO, previously reported to be highly expressed in the root meristem,
may function in vivo in auxin catabolism. This was demonstrated
visually in the bioassays.
In the experiment shown in Figure 4, the tips, or putative
auxin-catabolizing regions were removed from laterals on a cultured parent root and the result was a dramatic production of supernumerary lateral root primordia on the parent root. If roots with an existing, stable pattern of lateral roots are shifted to medium with higher auxin
concentration, as shown in Figure 1B, this same effect is demonstrated.
Our interpretation of this tip removal experiment is that we have
removed the main sites of auxin metabolism in root tissue and, once
removed, the parent root is now in an environment of high auxin and
reacts by organizing new lateral root meristems, thus generating new
root tips to replace these sites of auxin catabolism.
Until recently, it was believed that there were two major mechanisms
for auxin turnover in plants: (a) a peroxidase- and/or oxidase-mediated
oxidative decarboxylation, resulting in a loss of the number 1 carbon
(the carboxyl group) from the indole side chain; and (b) a
non-decarboxylative pathway (Normanly, 1997; Östin et al., 1998).
While the non-decarboxylative pathway is today accepted as the likely
process mediating free auxin levels, the significance of the
decarboxylation pathway has recently been challenged (Normanly, 1997;
Östin et al., 1998). Because the products of in vitro
decarboxylation could not be shown to occur in vivo (Reinecke and
Bandurski, 1983; Ernstsen et al., 1987; Bandurski et al., 1995),
oxidative decarboxylation is now considered an unimportant, perhaps
even an artifactual, route for auxin turnover. The reason for this
conclusion is evident when one considers the plant materials (primarily
shoot tissues) recently used to study auxin catabolism (Normanly, 1997;
Östin et al., 1998). Root tissues have rarely been employed, and,
when occasionally used, either the terminal 1 to 4 mm was excised prior
to initiating IAA turnover experiments or the entire plant was
extracted, which could lead to a masking of any decarboxylation
occurring in the small amounts of tissue comprising the root tips
(Nonhebel et al., 1983; Sztein et al., 1995). Additionally, when
decarboxylation products have been detected in extracts, these products
have often been viewed as resulting from experimentally induced
(artifactual) exposure of auxin to peroxidases (Bandurski et al., 1995;
Normanly, 1997; Östin et al., 1998). However, our results support
a specific (non-artifactual) capacity of root tips to decarboxylate
IAA, as suggested by several previous investigators (Morris et
al., 1969; Bourbouloux and Bonnemain, 1974; Pernet and Pilet,
1979).
Here we have shown that at least 30% of polarly transported auxin is
oxidatively decarboxylated in vivo by intact root tips (terminal
millimeter) (Fig. 8). Oxidative decarboxylation in the root occurs
almost exclusively at the tip (Fig. 8). Excision of the cap prevents
decarboxylation (Fig. 8). We also show that IAA is oxidatively
decarboxylated in vitro by AAO, forming as one of the primary products
oxindole-3-methanol (Fig. 7).
Although we have not yet identified the in vivo-generated,
auxin-decarboxylated products, it is clear that decarboxylation is a
significant route for free auxin turnover in root tips (as much as 30%
of the auxin is oxidatively decarboxylated) (Fig. 8). Moreover, our
results do not support the contention that these decarboxylated
products arise from an artifactual mixing of IAA and a
peroxidase/oxidase, because if this were so, decarboxylation of IAA in
both intact and decapped root tips would have been seen, and instead
decarboxylation only occurred in intact tips (Fig. 8).
Furthermore, recent data with transgenic plants show that significant
alterations in endogenous peroxidases (increases or decreases) have no
effect on endogenous auxin levels (Lagrimini et al., 1992). This was
shown for either a 10-fold increase or a 90% decrease in peroxidase
levels (Lagrimini et al., 1992). These data thus argue against the
likelihood that the observed in vivo IAA decarboxylation was due to
peroxidase activity.
Previously we advanced a model in which polarly transported auxin
accumulated at the root apex, resulting in an enhancement in AAO
activity, a localized depletion of AA, and as a consequence, the
formation of the QC (Kerk and Feldman, 1995). Here we add an additional
step to that model by presenting evidence that AAO can also metabolize
auxin. We suggest that in the intact root tip, AAO functions as an
auxin oxidase regulating endogenous auxin levels at the root tip. Our
previous study localized high levels of AAO specifically in the QC and
cap of intact root tips (Kerk and Feldman, 1995).
One way of integrating the proposed auxin/AAO interactions in regard to
the maintenance of the QC is to suggest that a classical feedback loop
involving AAO and auxin exists: high auxin enhances AAO activity, which
leads to a decline in auxin, promoting a decline in AAO, allowing auxin
levels to again increase (Fig. 9).
Layered over this hypothesized feedback loop we suggest a parallel
regulation in the levels of AA (Fig. 9). While it is widely known that
AAO is present in most if not all higher plants, its regulation and biological function are not clearly defined (Arrigoni, 1994;
Córdoba and González-Reyes, 1994; Smirnoff, 1996). That
ascorbate is a substrate in vivo for AAO is highly probable (Avigliano
and Finazzi-Agro, 1997). Recent data reported by Kato and Esaka (1996, 1999) showed correlations between levels of AAO mRNA and ascorbate metabolism in synchronous, non-synchronous, and elongating cultured tobacco cells. Kato and Esaka (1999) proposed that AAO expression and
metabolic reaction are under control of the cell cycle and may be
involved in the process of cell elongation.
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Figure 9.
Hypothetical scheme of possible interactions
between polarly transported auxin (IAA), AA, and AAO.
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Our earlier data showing low levels of AA in regions with high AAO
support this suggestion of cell cycle regulation of AAO, and as
suggested earlier, could result in the formation of the QC. Therefore,
the maintenance and functioning of the QC may be a consequence not only
of the accumulation of auxin (Kerk and Feldman, 1995), but may also be
a result of the regulation by auxin of an auxin-catabolizing enzyme.
Whether AAO could regulate endogenous auxin levels within the QC has
not yet been proven. However, its in vitro ability to oxidize auxin,
its high concentration within the QC, and the property of auxin to
regulate AAO mRNA and protein levels and activity
(Esaka et al., 1992; Kerk and Feldman, 1995)
argue in favor of in vivo catabolic interactions between AAO and auxin.
Many interrelated factors need to be considered when proposing a
complex interaction between a hormone, an abundant metabolic enzyme,
and the cell cycle. Kato and Esaka (1999) suggested a role for
AAO-mediated metabolism in the apoplast during the process of cell
elongation. Our findings showing a pH optimum of 5.3 for the oxidative
decarboxylation of auxin are consistent with pH measurements in cell
walls. In this regard, and considering the acid-growth hypothesis
(Goodwin and Mercer, 1983), it is interesting to speculate on the
possible metabolism of auxin in the walls by AAO. One could speculate
that the subcellular pH and other forms of compartmentalization
play important roles in favoring different phases of the feedback
loop at different times of the cell cycle, in different regions of the
cell, and even in different regions of the root.
Thus, while oxidative decarboxylation may be a minor pathway for
regulating auxin levels in the whole plant, the occurrence of this
pathway within the root may have consequences for root development.
Determining whether this pathway operates in roots should provide an
understanding of the establishment, maintenance, and function of the
QC, and compartmentalization of the cap and other classic zones of the
root such as the zone of elongation.
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ACKNOWLEDGMENTS |
We thank Ian Sussex for discussions and review of the
manuscript, Steve Ruzin for his gracious and skilled assistance in
figure preparation, Larry Cool of the University of California Forest Products Laboratory for the IR analysis, Sarah Reisinger for assistance with AAO assays, Richard Malkin for biochemical advice, and Robert Bandurski for sharing his enthusiasm and thoughts on auxin metabolism.
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FOOTNOTES |
Received September 13, 1999; accepted November 8, 1999.
1
This work was supported by the National Science
Foundation (grant no. IBN-9404842).
2
Present address: Department of Molecular,
Cellular and Developmental Biology, Yale University, New Haven, CT
06520-8104.
*
Corresponding author; e-mail feldman{at}nature.berkeley.edu; fax
510-642-4995.
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(1982)
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