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Plant Physiol. (1998) 116: 495-502
Light Promotion of Hypocotyl Gravitropism of a
Starch-Deficient Tobacco Mutant Correlates with
Plastid Enlargement and Sedimentation1
Stanislav Vitha,
Ming Yang2,
John Z. Kiss3, and
Fred D. Sack*
The Ohio State University, Department of Plant Biology, 1735 Neil
Avenue, Columbus, Ohio 43210
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ABSTRACT |
Dark-grown
hypocotyls of a starch-deficient mutant (NS458) of tobacco
(Nicotiana sylvestris) lack amyloplasts and plastid sedimentation, and have severely reduced gravitropism. However, gravitropism improved dramatically when NS458 seedlings were grown in
the light. To determine the extent of this improvement and whether
mutant hypocotyls contain sedimented amyloplasts, gravitropic sensitivity (induction time and intermittent stimulation) and plastid
size and position in the endodermis were measured in seedlings grown
for 8 d in the light. Light-grown NS458 hypocotyls were gravitropic but were less sensitive than the wild type (WT). Starch occupied 10% of the volume of NS458 plastids grown in both the light
and the dark, whereas WT plastids were essentially filled with starch
in both treatments. Light increased plastid size twice as much in the
mutant as in the WT. Plastids in light-grown NS458 were sedimented,
presumably because of their larger size and greater total starch
content. The induction by light of plastid sedimentation in NS458
provides new evidence for the role of plastid mass and sedimentation in
stem gravitropic sensing. Because the mutant is not as sensitive as the
WT, NS458 plastids may not have sufficient mass to provide full
gravitropic sensitivity.
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INTRODUCTION |
The availability of starch-deficient mutants has provided new
opportunities to test the starch-statolith hypothesis, i.e. the idea
that gravity sensing relies on the mass of amyloplasts that sediment.
In WT plants amyloplast sedimentation occurs in highly specific tissues
such as the central rootcap (columella) and in the starch sheath
(endodermis) in stems (Sack, 1987 , 1991). In mutants with little or no
starch, plastid sedimentation appears to be absent from these locations
(Caspar and Pickard, 1989; Kiss and Sack, 1990). Roots and hypocotyls
of starchless or starch-deficient mutants of Arabidopsis
thaliana are gravitropic, but sensing is reduced, as measured by
an increased threshold of stimulation and by a greater variability in
organ orientation in response to prolonged stimulation (Caspar and
Pickard, 1989; Kiss et al., 1989, 1996). Light-grown roots of a
starch-deficient mutant of tobacco (Nicotiana
sylvestris), NS458, are also gravitropic and have reduced
sensitivity (Kiss and Sack, 1989).
The findings that five different mutants at four loci have no or
decreased starch and reduced gravitropism support the conclusions that
starch plays a role in sensing when present, and that at least moderate
levels of starch are necessary for full gravitropic sensitivity (Poff
et al., 1994; Kiss et al., 1996; Sack, 1997). These hypotheses are also
supported by many other correlative data, including older experiments
with maize plastid mutants and with the depletion of starch by
experimental manipulation (Hertel et al., 1969; Sack, 1991, 1997).
Data from NS458 hypocotyls provide striking support for the
starch-statolith hypothesis, since the endodermis contains small, unsedimented plastids and these hypocotyls are severely deficient in
gravitropism when grown in the dark (Kiss and Sack, 1990). However,
preliminary experiments indicated that light-grown NS458 hypocotyls
were strongly gravitropic without obvious increases in starch content,
data that raised the possibility that light restored gravitropism
independent of plastid mass or position. To analyze further the effects
of light, we determined whether light-grown NS458 hypocotyls are as
sensitive to gravity as WT hypocotyls, and whether light affects the
amount of starch or plastid position compared with dark-grown mutant
hypocotyls. This analysis shows that light induces plastid
sedimentation in the NS458 endodermis, probably through an increase in
plastid volume and in total starch content, and that light-grown mutant
hypocotyls exhibit significant gravitropism but are less sensitive than
the WT.
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MATERIALS AND METHODS |
Plant Material and Cultivation
Seeds used were WT tobacco (Nicotiana sylvestris Speg.
et Comes) and the starch-deficient mutant NS458 (fourth generation with
reselection for the phenotype after the first backcross). NS458 is
deficient in the activity of plastidic phosphoglucomutase (Hanson and
McHale, 1988).
Seedlings were grown in square polystyrene Petri dishes (100 × 15 mm) on 1% (w/v) agar containing nutrients supplemented with 1%
(w/v) Suc, as described by Kiss and Sack (1990). The dishes were sealed
with Parafilm (American National Can, Greenwich, CT) and placed on the
edge so that the surface of the agar was vertical, and were placed
under continuous illumination (60-80 µmol m 2
s1 from 40-W cool-white fluorescent lamps,
General Electric). After 8 d of cultivation, the hypocotyls were
about 1 to 1.5 mm long. For assessment of plastid sedimentation,
dark-grown seedlings were also used and cultivated as described by Kiss
and Sack (1990).
Measurement of Gravitropic Curvature and Growth
To measure curvature and growth, light-grown seedlings were
photographed and then transferred to the dark and either turned to the
horizontal (for measurement of curvature) or kept upright (for
measurement of growth). Dark-grown seedlings were intermittently photographed using Kodak T-Max 400 ASA film and illumination with dim-green light (intensity of approximately 0.9 µmol
m2 s1 at the level of
the hypocotyl) provided by an incandescent lamp filtered through two
layers of a Roscolux filter (no. 1090, Rosco Laboratories, Port
Chester, NY) with a peak transmission of 526 nm and one-half bandwidth
of 58 nm. Growth rates were measured during the first 6 h in
darkness using digitally scanned photographic negatives and NIH
Image program (National Institutes of Health, Bethesda, MD).
Gravitropic curvature was measured from photographic prints as the
increment over the initial angle of each individual hypocotyl. At the
start of the gravitropism experiments hypocotyl angles of both
genotypes were within 10° from the vertical, since seedlings were
grown for 8 d with the light source directly above, inducing
phototropism.
For the measurement of the presentation time, hypocotyls were turned to
the horizontal and immediately placed in the dark for 15 to 150 min and
then rotated on a clinostat for 2 h. Seedlings were rotated at 1 rpm on a horizontal clinostat, with the axis of rotation parallel to
the root-hypocotyl axis. A plot of curvature versus the stimulation
time was constructed and regression lines for the WT and NS458 were
calculated. The intercept of the extrapolated regression line and the
x axis was taken as an estimate of the presentation time.
To measure the perception time by intermittent stimulation, Petri
dishes were turned to the horizontal and immediately mounted on a
clinostat in the dark. The seedlings were exposed to a total of 12 cycles (10 min each) consisting of gravistimulation (for 0.5-8 min)
when the clinostat was stationary and the seedlings were horizontal,
followed by rotation for the rest of the cycle. For 0.5-min
stimulations, seedlings were rotated for only 9 min in each cycle.
After the 12th cycle, the dishes were rotated for an additional 60 min.
Curvatures were tested for significance using a paired Student's
t test.
Plastid Sedimentation, Size, and Starch Content
Petri dishes containing light-grown seedlings were inverted
(placed upside down) and kept in darkness for 1 h. The dishes were then filled with fixative consisting of 1.8% (w/v) formaldehyde, 5% (v/v) acetic acid, and 45% (v/v) ethanol, through a small hole in
the top edge of the dish. Seedlings were then briefly vacuum infiltrated and fixed overnight, and were kept in an inverted orientation throughout. The hypocotyls were then washed in 50% ethanol, dehydrated, and embedded in paraffin or Steedman's wax (Vitha
et al., 1997). Longitudinal 10-µm sections were dewaxed and stained
for starch with an IKI solution (2% [w/v] KI, 1% [w/v] I).
In WT stems the endodermis is obvious because of the presence of large,
sedimented amyloplasts. In NS458 the endodermis can still be recognized
by its location as the innermost layer of the cortex. Only endodermal
cells within the most apical 0.5 mm (light-grown hypocotyls) or 3 mm
(dark-grown hypocotyls) were analyzed for sedimentation and plastid
size. Sedimentation was evaluated by counting the number of plastids in
the top, middle, and bottom thirds of individual endodermal cells
visualized in the microscope. Sections from five hypocotyls were
assessed for each genotype and light treatment (WT versus NS458; light
versus dark).
For measurement of plastid size, the sections were photographed using a
Plan 100× oil-immersion lens (model NA 1.3, Zeiss), and negatives were
digitally scanned. NIH Image program software was used to trace plastid
outlines at a total magnification of 20,000 to determine the minimum
and maximum diameters of each plastid. For NS458, individual starch
grains within plastids were also measured. Optical sectioning of
endodermal plastids in hypocotyls indicated that plastid shape could
best be approximated as an ellipsoid in both genotypes. Thus, plastid
volume, V, was calculated for an ellipsoid, where
V = [4/3 ×  × (length/2)] × (width/2)2. The same calculation was used for the
volume of starch grains within the NS458 plastids.
Stokes' law was used to calculate the theoretical velocity,
v, of plastid sedimentation, where v = 2/9 × (d1  d2) × g × r2/ , and
(d1 d2) is the difference between the densities of the plastid and the cytoplasm, g is the
gravitational constant, r is the plastid radius (calculated
from plastid volume assuming a sphere), and is the viscosity of the
cytoplasm (here assumed to be 0.3 Pa; Björkman, 1988). The
density values used were 1.015 g cm3 for
cytoplasm, 1.42 for amyloplasts, and 1.45 for starch. For NS458
plastids, starch occupied 10% of plastid volume (see ``Results''), and the density of the remaining volume was assumed to be 1.113 g
cm3 (approximated from the density of 30%
Glc). The potential energy of the sedimenting particle was calculated
as (d1 d2) × V × g × s, where
V is the plastid volume and s is the distance
displaced (here set at 10 µm).
Endodermal cell length was measured using an ocular micrometer from
4-µm sections of hypocotyls that were fixed in glutaraldehyde and
embedded in Spurr's resin as described by Kiss and Sack (1990). Cell-length measurements were made only in regions also used to measure
plastid size and sedimentation.
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RESULTS |
Light Significantly Improves NS458 Hypocotyl Gravitropism
NS458 hypocotyls grown in the dark for an extended period appeared
almost agravitropic (Fig. 1E), whereas WT
hypocotyls under similar conditions were upright (Fig. 1A). Light
substantially improved the gravitropism of NS458 hypocotyls compared
with those grown in the dark. (Note that gravitropism was allowed to
develop in the dark, regardless of whether the plants had previously
been grown in the light or the dark.) Whereas gravitropic curvature was
about 10° (after 70 h horizontal) for dark-grown NS458
hypocotyls, it was almost 50° (after 50 h) for light-grown NS458
hypocotyls, a value close to that of the light-grown WT hypocotyls
(Fig. 2). In addition, the onset of
gravitropic curvature was much earlier and the rate of curvature was
much greater in light- versus dark-grown NS458 hypocotyls (Fig. 2).
This difference was not the result of growth rate, because dark-grown
hypocotyls elongated twice as fast as light-grown hypocotyls (Table
I). Also, within the same light
treatment, WT and NS458 hypocotyls had the same growth rates (Table I).
Thus, the 50% slower rate of gravitropic curvature of light-grown
NS458 compared with WT hypocotyls (Fig. 2B) also cannot be explained by
a difference in growth rate. Cultivation in the light had much less of
an effect on the gravitropism of the WT hypocotyls (Fig. 2).
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| Figure 1.
Gravitropism and endodermal plastid sedimentation.
Dark-grown WT plants (WT-D) are gravitropic (A) and contain large,
sedimented amyloplasts (B) located approximately at the arrow in A. Dark-grown NS458 hypocotyls (M-D) are severely disoriented (E) and the
endodermis (at the arrow in E) contains small plastids with small
starch grains (F). Light-grown WT seedlings (WT-L, C) contain large
amyloplasts that are sedimented in the endodermis (D, arrows in J and
K). Light-grown NS458 seedlings (M-L, G) contain plastids that are larger and contain more starch (H) (compared with dark-grown mutants, F) that are sedimented in the endodermis (arrows in L and M). The
position of the lower cell wall is indicated by an arrowhead in M. The
endodermis in light-grown NS458 hypocotyls (arrows in I, L, M, and O)
can be identified based on its position as the innermost layer of the
cortex adjacent to the stele, even though it has less starch than the
WT (N). All sections come from tissues that were fixed after being
inverted for 1 h (see ``Materials and Methods'') except for the
section in I, which was kept horizontal for 1 h before fixation. All sections are longitudinal except for those in N and O, which are
cross-sections. The gravity vector is toward the bottom of all figures,
except for the cross-sections. All sections are of Steedman's wax
stained with IKI to localize starch (blue-black) except for the section
in I, which is of Spurr's resin stained with toluidine blue. Scale
bars = 5 µm (B, D, F, and H), 50 µm (J-M), 100 µm (I, N,
and O), or 10 mm (A, C, E, and G).
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| Figure 2.
Time course of gravitropic curvature after
rotating dishes 90°. A, Dark-grown plants (data redrawn from Kiss and
Sack, 1990). B, Seedlings grown for 8 d in the light were placed
in the dark and immediately reoriented to the horizontal. Compared with
dark-grown NS458 hypocotyls that exhibit almost no gravitropism,
light-grown NS458 hypocotyls eventually curve almost as much as the WT.
se bars are shown when greater than the size of the symbol.
The experiment was repeated three times with similar results.
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Table I.
Growth rates (mean ± se) of
vertically grown hypocotyls
Growth was measured during a 6-h period in the dark, regardless of
whether the plants were previously grown in the light or in the dark
for 8 d; n = 37 to 83 plants.
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Light-Grown WT Hypocotyls Are More Sensitive
One measure of gravitropic sensitivity is the presentation time
(also known as the induction time), which is the shortest single dose
of gravistimulation that is a threshold to curvature. Regression lines
were calculated from plots of stimulation time versus curvature (Fig.
3). The regression lines shown in Figure 3 were calculated from all stimulation times, which included at least
one value (from different experiments) that was statistically different
(P < 0.05) from zero curvature. This criterion excluded the 0.25- and 0.5-h values (Fig. 3,  ) for NS458 from the calculations. For the
regression lines shown in Figure 3, the presentation times were 4.8 and
30 min for the WT and for NS458, respectively. Varying combinations of
inclusion of the zero-curvature values and exclusion of values for
longer stimulation times (2 or 2.5 h) produced a range of values
for the presentation times from 14.8 to 16.3 min for the WT and 15.8 to 33.1 min for NS458 (regression lines not shown). Whereas these
variations indicate the difficulty of obtaining a precise value for
this threshold, they do show that WT hypocotyls are more sensitive to
single, short periods of reorientation than are hypocotyls of NS458.
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| Figure 3.
Presentation time of light-grown hypocotyls.
Gravitropic curvature in response to single horizontal doses from 0.25 to 2.5 h. The intercept of regression lines (r = 0.95 for the WT [ ] and 0.61 for NS458 [ , ]) with the
x axis yields estimated presentation times of 0.08 and
0.50 h for the WT and NS458, respectively. Only time points with
at least one nonzero value for curvature were included (open symbols)
in the calculation of the regression. Thus, the values indicated by the
filled triangles for NS458 were not used for the regression shown in
this figure. se bars are shown when greater than the size
of the symbol. Each symbol represents one experiment with 30 to 44 seedlings.
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Another measure of sensitivity was derived from intermittent
stimulation experiments, which indicate the shortest repeated stimulation times that can be integrated to produce statistically significant curvature. WT hypocotyls showed significant curvature if
they were repeatedly horizontal (stationary) for 1 min or longer, whereas at least 2-min periods were necessary to induce significant curvature in NS458 hypocotyls (Table II).
Thus, by the measure of intermittent stimulation, light-grown NS458
hypocotyls are only half as sensitive as WT hypocotyls.
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Table II.
Gravitropic curvature after intermittent
stimulation
The seedlings were intermittently stimulated for 2 h (12 10-min
cycles with 0.5-8 min of stationary, horizontal stimulation for each
cycle) and then rotated continuously on a clinostat for 1 additional
hour. The curvatures (degrees) were tested by paired Student's
t tests to determine whether the values were significantly different from 0. Each number represents the mean from one experiment with 33 to 50 seedlings.
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Light Increases Mutant Plastid Size but Not the Volume Fraction of
Starch
Light increased the plastid size in the endodermis of both
genotypes (Table III). Regardless of
light treatment, WT plastids were larger than NS458 plastids (Fig. 1,
B, D, F, and H). The volume of WT plastids consisted almost entirely of
starch, whereas only about 10% of plastid volume was occupied by
starch in both light- and dark-grown NS458 plastids (Table III). Thus,
although plastids of the light-grown NS458 have more starch on an
absolute basis than in the dark-grown mutant, the volume fraction of
starch is the same in both the light and in the dark.
Light Induces Mutant Plastid Sedimentation
In both dark- and light-grown WT hypocotyls, the plastids in the
endodermis (starch sheath) were amyloplasts that were filled with
starch and consistently sedimented (Fig. 1, B, D, J, and K). Previous
qualitative observations have shown that dark-grown NS458 hypocotyls
lack obvious plastid sedimentation (Kiss and Sack, 1990).
Quantification of plastid position confirms this conclusion because
plastids were found with equal frequency in the apical, middle, and
basal thirds of endodermal cells (Fig. 4). However, plastid sedimentation did
occur in the endodermis of light-grown NS458 hypocotyls (Fig. 1, L, and
M). Endodermal cells can be identified based on their position outside
of the stele and inside of the other cortical layers, even in the
absence of an obvious starch sheath (Fig. 1, I, L, and O).
Quantification of plastid position in light-grown NS458 hypocotyls
confirmed the presence of plastid sedimentation, even though it was not as complete as in the WT (Fig. 4). NS458 endodermal cells were shorter
than those of the WT in light-grown hypocotyls (Table IV), a difference that might overestimate
the extent of sedimentation in NS458 relative to the wild type because
sedimentation was measured by dividing the cell into thirds rather than
into cell segments of fixed lengths. Endodermal cell length was
essentially unaffected by light treatment in the wild type, whereas
these cells were much longer in dark- versus light-grown NS458
hypocotyls (Table IV).
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| Figure 4.
Plastid sedimentation in light- and dark-grown
hypocotyls. Shown is the percentage of plastids present in each third
of endodermal cells of seedlings that were inverted for 1 h, fixed
in this orientation, embedded in wax, sectioned, and stained. Plastids
are sedimented, except in dark-grown NS458 hypocotyls. Bars = +se. A total of 39 to 61 endodermal cells from five
different hypocotyls were scored for plastid sedimentation for each
genotype and condition. Open bars, Light-grown WT; black bars,
dark-grown WT; white bars with horizontal lines, light-grown NS458; and
black bars with horizontal white lines, dark-grown NS458.
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Table IV.
Length of endodermal cells of WT and NS458
hypocotyls
Cell length (mean ± se) from the apical part of the
hypocotyl (see text). n = 40 to 85 cells. All values
are different ( < 0.01; analysis of variance, Newman-Keuls multiple
comparisons).
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DISCUSSION |
The light promotion of hypocotyl gravitropism in NS458 correlates
with the onset of plastid sedimentation, a finding consistent with the
hypothesis that gravitropic sensing is plastid based. Whereas
gravitropism in dark-grown NS458 hypocotyls is severely impaired (Fig.
2) (Kiss and Sack, 1990), mutant hypocotyls from seedlings grown in the
light are strongly gravitropic. This light promotion cannot be
attributed to effects on growth, because the growth rates of WT and
NS458 hypocotyls are equal and because the growth of both genotypes is
twice as fast in the dark as in the light.
Light, especially red light, has been shown to promote gravitropism in
stems in other genera (Britz and Galston, 1982; Woitzik and Mohr,
1988), and light and phytochrome modulate stem gravitropism (Poppe et
al., 1996; Robson and Smith, 1996). We did not investigate either the
wavelength(s) or the duration of irradiation necessary for the light
promotion of gravitropism in NS458 hypocotyls. But unlike in the
studies cited above, we examined the effects of light on gravitropic
sensitivity and on the size and sedimentation of endodermal plastids.
The finding that endodermal plastid sedimentation is absent in
dark-grown NS458 hypocotyls but is largely present in light-grown mutant hypocotyls suggests that this sedimentation is responsible for
the significant promotion by light of gravitropism. This is supported
by data for the WT in which the extent of both amyloplast sedimentation
and gravitropism are comparable in the light and in the dark. However,
we cannot rule out the possibility that light promotes gravitropism
independently of an effect on plastid size, e.g. via signal
transduction.
Plastid sedimentation in light-grown NS458 hypocotyls was seen only in
the endodermis close to the shoot apex. This is the same location for
amyloplast sedimentation in the stems of WT plants of most genera
(Perbal and Rivière, 1980; Sack, 1987, 1991; Caspar and Pickard,
1989; Volkmann et al., 1993). Of course, the concept that plastid
sedimentation functions in gravitropic sensing is a century old and is
supported by numerous correlational data (Sack, 1991, 1997). This
hypothesis is also strongly supported by the recent finding that
hypocotyls of the scarecrow mutant of Arabidopsis are
agravitropic and lack an endodermis and amyloplast sedimentation
(Fukaki et al., 1997). Our data for NS458 hypocotyls provide one more
correlation that plastid mass and sedimentation in the endodermis are
necessary for substantial gravitropism in stems.
Plastid sedimentation in light-grown NS458 probably results from an
increase in plastid mass. Although the NS458 mutant has been described
as starchless (Hanson and McHale, 1988; Sicher and Kremer, 1996), its
plastids do have small amounts (approximately 10% by volume) of starch
(Kiss and Sack, 1990; this study). In endodermal cells the percent of
plastid volume occupied by starch is no different in light- or in
dark-grown NS458 hypocotyls (Table III). This may indicate that light
does not alter the already defective enzyme activity of plastidic
phosphoglucomutase or the possible metabolic feedback mechanisms that
affect the volume fraction of starch. Nevertheless, light increases
plastid volume overall, both in the WT and in NS458, and this results
in a greater absolute amount of starch, which may be sufficient to
induce endodermal plastid sedimentation in the mutant. However, it is
possible that sedimentation also results from additional effects of
light, such as a decrease in cytoplasmic viscosity or from an increase
in the density of the plastid through the accumulation of soluble sugars or of other compounds.
Although light-grown NS458 plastids have sufficient mass to sediment,
they do so with much less consistency than heavier, WT amyloplasts. We
calculate from Stokes' law that the theoretical velocity of
sedimentation of light-grown NS458 plastids (approximately 2 µm
min1) is about twice that of dark-grown mutant
plastids, but much less than that of WT amyloplasts (7.8 and 10.3 µm
min1 from dark- and light-grown hypocotyls,
respectively). According to Björkman (1988), the potential energy
of the sedimenting particle must be much higher than thermal noise
(Brownian motion; 2 × 1021 J) to trigger
gravitropic sensing. It is estimated that dark-grown NS458 plastids
possess a potential energy (approximately 1 × 1019 J) that is only 1 to 2 orders of magnitude
higher than thermal noise, and that is only 25% of the potential
energy of light-grown mutant plastids (4 × 1019 J). The potential energies of WT
amyloplasts from dark- and light-grown plants are estimated to be
1.5 × 1018 J and 2.3 × 1018 J, respectively.
How then does a relatively small increase in mutant plastid mass or
potential energy provide an orientational signal sufficient to restore
significant gravitropism to light-grown NS458 hypocotyls? Spaceflight
experiments have demonstrated that gravitropic sensing can occur at
thresholds as low as 0.1g (Brown et al., 1995). This suggests that the gravitropic sensing apparatus is "overbuilt" and
may be capable of discriminating a signal from noise at some threshold
greater than thermal noise (Sack, 1997). If all gravitropic sensing is
plastid based, then the potential energy in the plastids of dark-grown
NS458 may be sufficient to barely trigger enough gravitropic sensing to
produce the residual levels of gravitropism observed in these
hypocotyls (Kiss and Sack, 1990). The estimated 4-fold greater
potential energy of light-grown (compared with dark-grown) mutant
plastids may be sufficiently higher than threshold levels to
dramatically increase the orientational signal and thus gravitropism.
The increased mass may also be important in inducing sedimentation,
which places these plastids in a part of the endodermal cell that may
contain or be enriched in mechanosensitive receptors that transduce
plastid mass.
This reasoning might also explain why light-grown WT hypocotyls are at
least twice as sensitive as NS458 hypocotyls, as estimated by the
presentation time and by intermittent stimulation. The reduced
sensitivity of the light-grown mutant compared with the WT might result
both from the lower plastid mass and from the less-consistent
sedimentation of mutant plastids. Based on data from roots of
starch-deficient mutants of Arabidopsis, Kiss et al. (1996, 1997)
estimated that mutants, the plastids of which contain  60% of the
starch of the WT, exhibit full WT levels of gravitropic sensitivity.
The starch content of NS458 plastids is significantly below this level.
The relative importance of plastid sedimentation to gravitropic
sensitivity may differ in stems and roots. As in hypocotyls, light-grown roots of NS458 are strongly gravitropic but less sensitive than the WT, but mutant roots apparently lack sedimentation in columella cells (Kiss and Sack, 1989). These data show that plastid sedimentation is not necessary for significant gravitropism in roots,
whereas there is a strong correlation for hypocotyls of the same
mutant. This difference might be related to the much smaller size of
columella versus endodermal cells.
Although light promotes gravitropism in NS458 hypocotyls, comparable
data are not available for an Arabidopsis mutant that is also deficient
in plastidic phosphoglucomutase, the pgm1 locus represented
by two alleles, TC7 and ACG21 (Caspar and Pickard, 1989; Kiss et al.,
1996). pgm1 appears to be completely devoid of starch
(Caspar, 1994). Dark-grown hypocotyls are severely disoriented, although they are capable of weak gravitropism (Caspar and Pickard, 1989; Kiss et al., 1997). Depending on the report, light-grown pgm1 hypocotyls either show WT levels of curvature (fig. 7 in Caspar and Pickard, 1989) or less gravitropic curvature than in mutant hypocotyls grown in the dark (figs. 5 and 6 in Kiss et al.,
1997). Neither sensitivity nor plastid size and position were measured
in these studies. Based on our results with NS458, we predict that
light-grown pgm1 hypocotyls should be slightly more
sensitive than dark-grown mutant hypocotyls; whereas light might
enlarge pgm1 plastids, the increase in plastid mass should be much less than in NS458 because, unlike NS458 plastids,
pgm1 plastids entirely lack starch.
In summary, our data are consistent with the conclusions that the light
promotion of gravitropism in NS458 results from an increase in plastid
mass that induces plastid sedimentation and greatly increases
gravitropic sensitivity compared with the dark-grown mutant. These data
support the importance of both plastid mass and plastid sedimentation
in gravitropic sensing.
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FOOTNOTES |
1
This research was supported by the National
Aeronautics and Space Administration (grant no. NAGW-4472).
2
Present address: Cold Spring Harbor Laboratory,
Cold Spring Harbor, NY 11724.
3
Present address: Botany Department, Miami
University, Oxford, OH 45056.
*
Corresponding author; e-mail sack.1{at}osu.edu; fax
1-614-292-6345.
Received July 24, 1997;
accepted November 10, 1997.
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ABBREVIATIONS |
Abbreviation:
WT, wild type.
| |
ACKNOWLEDGMENTS |
We are thankful to Roger Hangarter for helpful discussions and
to Shawn Davis and Liming Zhao for technical assistance.
| |
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Top
Abstract
Introduction