Int. J. Plant Sci. 168(5):619–625. 2007.
Ó 2007 by The University of Chicago. All rights reserved.
1058-5893/2007/16805-0010$15.00
PLASTICITY AND TIMING OF FLOWER CLOSURE IN RESPONSE TO POLLINATION
IN CHAMERION ANGUSTIFOLIUM (ONAGRACEAE)
Mary Jane Clark1 and Brian C. Husband2
Department of Integrative Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada
Flower longevity often exhibits a plastic response to pollination and may vary adaptively to maximize pollen
removal and deposition and minimize flower maintenance costs. Using Chamerion angustifolium (fireweed),
we experimentally tested two predictions that follow from this hypothesis: (1) flower closure should be
triggered early, before fertilization, if it is to reduce costs and adjust to maximize pollen receipt; and (2) flower
closure should respond differently to different kinds (outcross, self) and amounts (none, low, high) of pollen. In
C. angustifolium, flower closure was initiated 4 h after pollination, well before any ovules were fertilized
(25 h). The addition of pollen to stigmas significantly shortened the time to closure compared with
unpollinated controls. Flower closure times were reduced with increasing pollen loads, up to 700 pollen grains,
as might be expected if closure was adaptive. Flower closure was slower in self-pollinations than outcross
pollinations at 24 and 48 h after pollination but not across the whole experiment. These results show that
flower closure is highly plastic in fireweed and consistent with an adaptive response in that it promotes seed
production and outcrossing over a range of pollination conditions.
Keywords: flower longevity, flower closure, pollination, fertilization, Chamerion angustifolium.
Introduction
shortened in response to various external stimuli. Indeed, floral senescence has been induced by temperature change (van
Doorn and van Meeteren 2003), light (van Doorn and van
Meeteren 2003; Vesprini and Pacini 2005; He et al. 2006),
predators (Wright and Meagher 2003), and pollination (Motten
1986; Porat et al. 1995; but see Gori 1983). Pollinationinduced flower closure is well documented in a large number
of species (van Doorn 1997), and researchers have hypothesized that plasticity in flower longevity is an adaptation to
maximize pollen dispersal and receipt while minimizing the
energy costs of maintaining flowers (Primack 1985; Webb
and Littleton 1987; van Doorn 1997). However, the adaptive
significance of pollination-induced floral longevity has not
been fully tested. Specifically, it is not clear whether adjustments in floral duration occur before or as a consequence of
fertilization. If plasticity in floral life span functions adaptively to adjust pollen receipt and hence the quantity and
diversity of offspring produced, then it should respond to
pollination early, before all ovules are committed. Some studies have evaluated the chemical signals produced in response
to pollination that cause perianth senescence and thus influence floral longevity (reviewed in O’Neill 1997). These signals can be produced early and in multiple steps throughout
the pollination-fertilization process. The question remains as
to the timing of flower closure with respect to fertilization, as
evaluated by direct observation of ovules. Furthermore, there
has been limited attention to the possibility that floral duration is sensitive to either the kinds or the quantities of pollen
applied (Proctor and Harder 1995; Sato 2002).
In this study, we used the herbaceous perennial fireweed
(Chamerion angustifolium L. Holub) (Onagraceae) to evaluate the hypothesis that plasticity in floral life span can function adaptively to regulate the quantity and quality of pollen
Floral life span, the length of time a flower remains open
and functional (i.e., receives pollinator visits; disperses and
receives pollen), is highly variable among flowering plants
(Ashman and Schoen 1994; van Doorn 1997; Ishii and Sakai
2000). Among species, the duration of flower opening can range
from a few hours to days or months (Kerner von Marilaun
1895; Ashman and Schoen 1996). The significance of such
variation is not fully understood; however, researchers have
reported associations between flower longevity and taxonomic group, breeding system, and habitat (Primack 1985).
Floral life span is believed to be adaptive (Kerner von Marilaun
1895; Primack 1985; Ashman and Schoen 1996) because of
its potential fitness consequences. It can influence the quantity and diversity of pollen a plant receives (Primack 1985).
At the same time, maintaining flowers in a functional state
requires resources that could otherwise be allocated to processes such as seed development, production of new flowers,
or vegetative growth (Webb and Littleton 1987; van Doorn
1997). Therefore, floral longevity is likely a compromise between selection to enhance pollen dispersal and receipt and
to minimize further investment in existing flowers (Schoen
and Ashman 1995; Ashman and Schoen 1996).
Floral longevity can also be plastic in its expression within
a plant. The duration of a given flower may be extended or
1 Current address: Department of Plant Agriculture, University of
Guelph, Guelph, Ontario N1G 2W1, Canada; telephone 519-824-4120
ext. 58271; e-mail [email protected].
2 Author for correspondence; telephone 519-824-4120 ext. 54790;
e-mail [email protected].
Manuscript received August 2006; revised manuscript received January 2007.
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received by a flower. Flowers are relatively large, protandrous
(male function before female), and insect pollinated. They
are mostly cross-fertilizing (Husband and Schemske 1995,
1997); however, self-pollination (via transfer between flowers
on the same inflorescence) and outcross pollen deposition are
both possible. In this study, we addressed the following questions. (1) Does flower closure occur before or after fertilization? (2) Do the kind and quantity of pollen applied to
stigmas influence flower closure rates? (3) Is the effect of the
pollen treatments consistent with an adaptive response? As
stated above, we would expect flower life span to respond to
pollination early, before all ovules are fertilized. With respect
to pollen load size, an adaptive response at the flower level
would be to prolong flower longevity with light loads, since
this would increase the likelihood of further pollen deposition. In terms of pollen quality, we would expect flowers to
prolong longevity with the addition of self pollen. In fireweed, self-pollinated offspring are only 5% as fit as crosspollinated offspring (Husband and Schemske 1995), so fitness
would be enhanced by increasing the opportunity for outcross pollen to be deposited and to fertilize ovules.
Material and Methods
Plant Material
The study species, Chamerion angustifolium, is widely distributed throughout the Northern Hemisphere in open and
disturbed habitats. Plants are ca. 2 m tall, with indeterminate
inflorescences bearing from 10 to 15 open flowers at a time,
each with four pink petals (Husband and Schemske 2000).
We used 30 open-pollinated seed families collected from the
Fortress Mountain (K38) population, located at the Fortress
Mountain Ski Resort in Kananaskis Country, Alberta, Canada.
Seeds from each family were cleaned of their comose, sown
onto moist filter paper in petri dishes, and left to germinate
for 1.5 wk. Seedlings at the cotyledon stage were transplanted into 72-celled trays containing a substrate mixture of
ca. 40 parts Premier Promix BX soilless mix : 1 part Turface
Athletics MVP turface : 1 part Therm-O-Rock perlite. When
plants reached the six-leaf stage, they were transplanted
into 6-in-round plastic pots and grown in the University of
Guelph Axelrod greenhouses (temperature 20°–25°C; 15 h
light). Plants were watered with deionized water; side branches
were pruned and plants fertilized biweekly with Plant-Prod
soluble fertilizer (20-20-20). Plants were randomly assigned
to treatments after the first flower bud opened.
Timing of Flower Closure and Fertilization
To determine whether flower closure occurred before or after fertilization, we applied excess (ca. 2800 grains) outcross
pollen to intact stigmas and assessed flower closure and ovule
fertilization at eight different time intervals (0, 4, 8, 12, 16,
20, 24, and 48 h after pollination). Because of a lack of sufficient flowers on a single plant, each time treatment was randomly assigned to flowers among three plants for each of 24
maternal families. Hence, only two or three time treatments
were applied per plant for a total of eight per family. Excluding the first open flower (because of frequent abnormalities),
only fully open (style erect, stigma lobes fully expanded)
flowers received a pollen treatment. In addition, flowers with
damaged petals, partially closed petals, or thrips were also
excluded. Selected flowers were tagged with string, giving a
total of 24 flowers per treatment (n ¼ 192 crosses).
For each pollinated flower, we measured initial flower
openness as the distance separating the tips of the bottom
two petals. In a preliminary analysis, we found this measurement to be straightforward to measure and highly correlated
with the distance between petals and stigma (rs ¼ 0:92,
P < 0:0001). Each pollen application used pollen from four
anthers, each from a different maternal family. Pollen from
one anther was applied to each of the four stigma lobes. All
pollinations were completed before 12:30 p.m. At the designated treatment time, final flower openness was measured (as
described above), and the flower/immature fruit was collected. Flowers were also noted as closed when two or more
petals were touching the stigma or style. Petals and sepals
were removed from the flower, and a narrow slit was cut
along the middle two-thirds of the ovary wall. The fruit was
then immersed in 1 mL of formalin–acetic acid (FAA; 1 part
formalin : 1 part acetic acid : 1 part 50% ethanol) fixative in
a 1.5-mL microcentrifuge tube. Fresh FAA stock solution
was prepared every 3 d. Tubes were labeled and stored for
later ovule staining and clearing.
Fruits were cleared following Stelly et al. (1984), with
modifications by T. Sage (personal communication). FAA was
initially replaced with 50% ethanol and then changed hourly,
decreasing ethanol content 10% each time and ending with
two changes of deionized water. Fruits were then immersed
in Mayer’s hematoxylin solution (Sigma MHS-32), which
stains the nuclei and cellulose in the cell walls. After 3 d, the
stain was replaced with 3% glacial acetic acid. The following
day, the dehydration sequence was completed by replacing
the glacial acetic acid with deionized water and then increasing the ethanol content by 10% each hour until 100% ethanol was added. The solution was left overnight and replaced
with two changes of 100% ethanol the next day, leaving the
fruit immersed for 1 h in each fresh solution. The ethanol
was then replaced with a solution of 3 parts 100% ethanol
and 1 part methyl salicylate and left overnight. One part
methyl salicylate was added every hour for 3 h. After waiting
1 h, the solution was replaced with 100% methyl salicylate
for long-term storage.
Stained and cleared fruits were cut into top and bottom
halves, and ovules from each half were transferred to separate glass microscope slides containing 100% methyl salicylate. We separated fruit into halves because we expected
heterogeneity in fertilization between top and bottom due to
time required for pollen tube growth and changes in available resources. Ovules were separated from the comose and
other debris using a glass pipette and transferred to another
microscope slide. The ovules were immersed in 100% methyl
salicylate and covered with a glass cover slip. Slides were observed with a 3100 objective using a Leica DMRB brightfield
microscope. Macintosh Openlab software was used for further digital enhancement and analysis of images.
We estimated the frequency of fertilized ovules at all eight
time periods for 10 of the 24 maternal families, since the results were sufficiently uniform to address the question statistically. For each fruit, we scored fertilization in the first 10
CLARK & HUSBAND—VARIATION IN FLORAL LONGEVITY
ovules located on the slide in each of the top and bottom
halves of the fruit (in total, n ¼ 10 families 3 8 time
treatments 3 2 halves ¼ 160 frequency estimates). Ovules
were considered suitable for scoring if the central cell was
undamaged, the ovule was properly stained and cleared, and
the egg and polar nucleus were both clearly visible. Because
the egg was not always clearly discernible, an ovule was considered fertilized if the polar nucleus in the central cell was
dividing (i.e., two nuclei, two separate nucleoli present
within the nuclear membrane) (Dorken and Husband 1999).
Chamerion angustifolium has a monosporic embryo sac in
which only one polar nucleus contributes to the endosperm
(Johri et al. 1992).
ANOVA was used to compare the mean proportion of
petal closure—i.e., (initial openness openness at treatment
time)/initial openness—for flowers in each time treatment.
These values were arcsine transformed to improve normality
of the residuals; however, untransformed means were presented in the figures for ease of interpretation. A two-factorial
ANOVA was used to evaluate the influence of position (top
vs. bottom) and time since pollination on the proportion of
ovules fertilized (both treated as fixed effects). Also, the
mean time that flower closure was first observed was compared with the mean time that fertilization was first observed
using ANOVA (n ¼ 20 observations). All statistical analyses
were performed using JMP statistical software (SAS Institute,
Cary, NC).
The Effect of Pollen Type and Quantity on Floral Longevity
To determine whether flower life span is influenced by pollination conditions, we applied pollen of different types and
amounts to flower stigmas and tracked flower openness over
a 120-h period. Plants from 25 different maternal families
were used. Five pollination treatments were applied per family, each to a separate plant to avoid interactions between
pollinated flowers on an inflorescence. The pollination treatments applied were (1) no pollen, (2) ca. 2800 grains of self
pollen, (3) ca. 2800 grains of outcross pollen, (4) ca. 700 grains
of outcross pollen, and (5) ca. 100 grains of outcross pollen.
In total, 125 flowers were pollinated. The ca. 2800- and ca.
700-grain treatments were achieved by applying pollen from
four and one anthers, respectively, over the surfaces of four
stigmatic lobes. For the ca. 100-grain treatment, a 1-cm
length of plastic fishing line was cut, held with forceps, and
drawn through an anther to collect adhering pollen. The approximate number of pollen grains applied in each of the
three pollen load size treatments (2800, 700, 100 grains) was
determined by estimating the number of grains in a single anther and on a 1-cm length of fishing line using 13 different
plants. Pollen was counted using a Beckman Coulter Multisizer 3 particle counter.
Treatments were randomly assigned to a fully open flower
on the recipient plant. Initial openness was recorded (as before) before applying the pollination treatments. Flowers
were pollinated between 9:30 p.m. and 12:00 a.m. Openness
was measured 8 h after pollination and at 4-h intervals thereafter for the first 24 h, followed by 8-h intervals until the
flower closed (when two or more petals were touching the
stigma or style).
621
From initial and final openness measurements, the mean
proportion of petal closure—i.e., 1 ½ðopenness at treatment
time final openness)/(initial openness final openness)]—
was calculated for each treatment at each time interval. This
measurement differs from the previous experiment in that it
is expressed relative to the observed total flower closure and
gives a value of 1 when the flower is fully closed. These
values were arcsine transformed to improve normality of the
residuals and compared among treatments using a repeatedmeasures ANOVA (124 df overall; 4 df for treatment effect,
120 df for error). Orthogonal contrasts (each 1 df) were used
to conduct a set of a priori means comparisons (with vs. without pollen, 2800 self grains vs. 2800 outcross grains, 100 outcross grains vs. 700 outcross grains, 700 outcross grains vs.
2800 outcross grains). Untransformed means were presented
in the figures for ease of interpretation. Because the repeatedmeasures analysis can mask differences in the time course of
flower closure among pollination treatments, we also compared treatments at four times (12, 24, 48, 96 h) after pollination. The specific times, which are a logarithmic series,
were chosen to cover the full time course of the experiment
but with emphasis on early stages, when changes in flower
closure are most evident.
Results
Timing of Flower Closure and Fertilization
The proportion closure of flowers differed significantly
among time intervals (F6; 63 ¼ 19:26, P < 0:0001). Flower
closure was first observed 4 h after pollination, significantly
earlier than the beginning of fertilization (X ¼ 25:2 h,
SE ¼ 1:2) (F1; 18 ¼ 312:11, P < 0:0001) (fig. 1). Mean time
to 50% flower closure (ca. 16 h) was also earlier than the
first record of fertilization. The proportion of ovules fertilized differed significantly with time after pollination
(F7; 144 ¼ 196:43, P < 0:0001) (fig. 1). A Tukey’s HSD means
comparison showed fertilization at 48 h was higher than at
Fig. 1 Comparison of the mean proportion (6SE) of ovules
fertilized (diamonds) to the mean proportion (6SE) of petal closure
(squares) for Chamerion angustifolium flowers at 4-h intervals after
pollination with ca. 2800 grains of outcross pollen.
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INTERNATIONAL JOURNAL OF PLANT SCIENCES
all other times. Fertilization was first observed 20 h (mean
time to first fertilization ¼ 25:2 h) after pollination; by 48 h,
an average of 80% of ovules were fertilized.
The proportion of ovules fertilized was significantly higher
in the top of the ovary than in the bottom (F1; 144 ¼ 38:94,
P < 0:0001) (fig. 2). However, there was a position 3 time interaction (F7; 144 ¼ 30:81, P < 0:0001), reflecting the lack of
a position effect before 20 h. A significant difference between
fertilization in the top compared with the bottom of the fruit
was shown only at the 48-h time treatment when analyzed
using a Tukey’s HSD analysis.
The Effect of Pollen Type and Quantity on Floral Longevity
In a repeated-measures ANOVA, mean flower closure in
control flowers (no pollen added) was significantly later than
all other pollination treatments combined (F1; 120 ¼ 57:31,
P < 0:0001). Control flowers reached 50% closure at 48 h
compared with 17.5 h in treatments with pollen. In the
repeated-measures analysis, the course of flower closure after
self-pollination did not differ from that in the outcross pollination (F1; 120 ¼ 2:98, P ¼ 0:087) (fig. 3). Selfed flowers
closed at a rate similar to that of outcrossed flowers during
the first 16 h (50% closure self ¼ 15:5 h; outcross ¼ 17 h)
but appeared to slow after 16 h. This was confirmed by the
orthogonal contrasts conducted at four individual time periods, which showed significant differences between self and
outcross flowers at 24 and 48 h (table 1). However, we noted
that some of the pollen used in the self-pollination treatments
had an unusual texture and was difficult to remove from the
anthers. When we repeated the repeated-measures ANOVA
without those individuals (data not shown), flower closure
rates in self- and outcross pollinations showed less of a difference (F ¼ 0:88, P ¼ 0:351) than the original analysis, although the trends were similar.
The effect of quantity of outcross pollen on flower closure
was statistically significant (fig. 4). In the repeated-measures
ANOVA, the 100-grain treatment had slower flower closure
than the 700-grain treatment (F1; 120 ¼ 6:10, P ¼ 0:015). Orthogonal contrasts showed a difference at 24 h but not at 12,
48, or 96 h (table 1). There was no difference between 2800-
Fig. 2 Comparison of the mean proportion of fertilized ovules
(6SE) in the top (diamonds) and bottom (squares) of Chamerion
angustifolium fruits at eight time intervals after pollination with ca.
2800 grains of outcross pollen.
Fig. 3 Time course of flower closure in Chamerion angustifolium
after pollination with different kinds of pollen. Flowers received no
pollen (diamonds), ca. 2800 grains of outcross pollen (triangles), or
ca. 2800 grains of self pollen (squares).
and 700-grain treatments in the repeated-measures ANOVA
(F1; 120 ¼ 0:24, P ¼ 0:624), and this result was corroborated
by the orthogonal contrasts (table 1). Time to 50% closure
was 22.2 h for the 100-grain treatment compared with 17 and
15.5 h for the 2800- and 700-grain treatments, respectively.
Discussion
Flower longevity is a highly plastic trait that may function
adaptively in response to a variable pollination environment
to minimize flower maintenance costs and maximize fertilization and outcrossing rates (Primack 1985; Webb and Littleton
1987). In this study of Chamerion angustifolium, we tested
two predictions that follow from this hypothesis: (1) flower
closure should be triggered by cues before fertilization and
(2) flower closure must be able to respond differentially to
different kinds and amounts of pollen. We found that flower
closure was initiated 4 h after pollen deposition and was well
advanced before any ovules were fertilized. In addition,
flower longevity was reduced with increasing pollen loads
and responded weakly to self versus outcross pollen. In all
cases, plasticity in flower longevity was consistent with an
adaptive response.
Flower closure in C. angustifolium occurred ca. 16 h before fertilization was first observed. This result indicates that
the trigger for flower closure is not fertilization itself, as has
been implied in the literature (Ishii and Sakai 2000; Goodwillie
et al. 2004). Rather, triggers of flower closure are being transmitted earlier, coincident with pollination. To our knowledge, no other studies have directly examined the timing of
flower closure in relation to timing of fertilization. Stead and
Moore (1983) showed that pollination-induced flower senescence in Digitalis was associated with ethylene production
occurring at the top of the style, far from the ovary, and only
1 h after pollination, well before any fertilization likely
would have occurred. O’Neill (1997) reviewed other studies
that find ethylene production, which has been linked to
flower senescence, preceding pollen tube growth, although
fertilization was never measured directly. These results as
CLARK & HUSBAND—VARIATION IN FLORAL LONGEVITY
Table 1
Comparisons of Proportion Closure of Flowers among Pollination
Treatments in Chamerion angustifolium
Whole model
Orthogonal contrasts
Time
(h)
MS
F
C vs.
all
S vs.
C2800
C2800 vs.
C700
C700 vs.
C100
12
24
48
96
.23
6.10
2.60
.14
1.7
23.3***
15.3***
2.3
3.9*
69.9***
54.3***
7.8**
.8
5.1*
5.0*
.9
2.2
.4
.0
.0
1.8
17.8***
1.1
.5
Note. Results are given for the whole model (mean squares and
F values; 4 and 120 df for numerator and denominator variances,
respectively) and four planned contrasts (F values) at each of four
times (12, 24, 48, 96 h) after pollination. C ¼ control, S ¼ selfpollinated, C2800 ¼ cross-pollinated ca. 2800-grain application, C700 ¼
cross-pollinated ca. 700-grain application, C100 ¼ cross-pollinated
ca. 100-grain application.
P < 0:05.
P < 0:01.
P < 0:001.
well as our observations suggest that plasticity in flower closure occurs sufficiently early to allow investments in flower
function to be minimized when ample outcross pollen is available and to regulate patterns of fertilization and hence the
number and quality of offspring by adjusting availability of
the stigma for pollination.
Compared with unpollinated controls, flower longevity
was reduced by more than half through the addition of pollen. The influence of pollen deposition on floral life span
has been studied in many species (Primack 1985; Webb and
Littleton 1987; Porat et al. 1995; Proctor and Harder 1995;
Ashman and Schoen 1996; Clayton and Aizen 1996; Ishii
and Sakai 2000; Evanhoe and Galloway 2002; Sato 2002;
Stpiczynska 2003; Giblin 2005; Harder and Johnson 2005)
and was reviewed by van Doorn (1997). In many cases (but
not all), pollination accelerates flower closure and reduces total flower life span. These studies confirm that flower longevity is not always genetically fixed. Rather, it is highly plastic
and varies in response to external stimuli. Thus, it will be
particularly important for theoretical models of flower life
span to extend beyond optimality arguments for species
(Schoen and Ashman 1995) and to consider the effects of
temporal and spatial heterogeneity in pollen availability and
the fitness benefits of plasticity in floral longevity.
Although flower closure is coincident with pollen deposition, it is not clear from these results what aspect of the
pollination process is triggering flowers to close: pollen deposition, pollen germination, or pollen tube growth. Studies
of ethylene-induced flower senescence indicate that in some
species the deposition of pollen may be sufficient for triggering flower closure (O’Neill 1997). In C. angustifolium, pollen
tube growth to the top of the ovary has been observed as
early as 3 h after pollination (J. Tindall and B. C. Husband,
unpublished data). However, because flower closure was initiated at 4 h or later, it is not possible to separate the effects
of pollen deposition from germination or tube growth. In addition, we did not examine the physiological or molecular
mechanisms signaling flower closure in this species. However,
623
ethylene production is increasingly implicated in flower closure of other species. Changes observed in pollinated flowers
are similar to those seen in flowers exposed to exogenous
ethylene (van Doorn 1997, 2002). Moreover, a comparison
among families shows that families containing species with
pollination-induced flower closure are also the families that
are sensitive to exogenous ethylene (van Doorn 1997).
If flower longevity functions to regulate pollination, then
we expect flower closure to be affected by the amount and
composition of pollen received. Pollen quantity had a significant and negative effect on flower longevity. Increasing the
number of pollen grains deposited from 100 to 700 caused a
significant decrease in flower life span, whereas the increase
from 700 to 2800 grains had no detectable effect. Similarly,
Evanhoe and Galloway (2002) demonstrated a negative effect of pollen number on morphological female phase longevity, and Ishii and Sakai (2000) found that flower life span in
Erythronium japonicum decreased with increasing number
of grains added to the stigma but with diminishing effects.
These results are consistent with the expectation that flowers
should remain open when they receive insufficient pollen but
that the fitness benefits of keeping a flower open would decrease as the number of grains on the stigma approaches
some threshold, presumably related to the number of grains
necessary to fertilize all ovules. It is interesting that in C. angustifolium, the addition of pollen above 700 grains had no
effect on flower life span; this threshold is roughly similar to
the number of ovules (mean ¼ 390) found in diploid plants
(Husband et al. 2002).
Flower longevity was weakly affected by the kind of pollen
(self or outcross) placed on the stigma. Assessed across the
entire observation period, self pollen did not affect flower
longevity differently than outcross pollen; however, the degree of flower closure at 24 and 48 h after pollination was
significantly less with self pollen. This pattern is consistent
with an adaptive response. Chamerion angustifolium exhibits
high inbreeding depression, and selfed offspring are only 5%
as fit as outcrossed offspring (Husband and Schemske 1995).
Therefore, keeping selfed flowers open longer may increase
the opportunity for outcross pollen to be deposited on the
Fig. 4 Time course of flower closure in Chamerion angustifolium
after pollination with different quantities of outcross pollen. Flowers
received no pollen (diamonds), ca. 2800 grains (triangles), ca. 700
grains (squares), or ca. 100 grains (circles) of outcross pollen.
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INTERNATIONAL JOURNAL OF PLANT SCIENCES
stigma and to sire seeds. However, our observations must be
interpreted with caution. If self pollen is less likely to germinate on the stigma, then the ‘‘‘self versus outcross’’ effect
may result from having effectively fewer viable grains in the
self pollen treatment. Furthermore, in our study, a few plants
used in the self treatment produced pollen that appeared less
numerous and adhered to the anther sac (not confirmed
quantitatively). When we removed all pollinations involving
these plants, the effect of selfing was negligible, although
with the same trend. More research is needed on C. angustifolium and other species to evaluate whether flower life span
is sensitive to the quality of pollen deposited on stigmas.
We are aware of few other studies that have observed
flower closure in response to self-pollination. Gibbs et al.
(2004) examined flower longevity in the genus Ceiba. Compared with an unpollinated control, they found that selfing
actually extended floral life span. This differs markedly from
our own results, where selfing shortened life span relative
to no pollination, but is consistent with the idea that selfpollination is not likely to reduce floral life span. Unfortunately, these results are not fully comparable, because they
did not report the influence of cross-pollinations on floral
longevity. In addition, Sato (2002) compared flower longevity
of staminate and pistillate phases of two Impatiens varieties
under autonomous selfing and outcrossing. They found that
longevity of the pistillate stage in a mixed-mating variety
was longer in self compared with outcross pollinations. In
contrast, a selfing variety had a shorter pistillate phase upon
selfing. However, again, these results are not directly comparable to ours because selfing occurred autogamously rather
than being applied manually, nor did selfing occur at the
same time as the outcross pollination.
Our study has focused on the extent to which plants can
regulate female function—the kinds and amounts of pollen
received on their stigmas and hence fecundity and mating
system—through rapid changes in flower life span. However,
this may be an overly restrictive view in that flower closure
may affect fitness through other means. Ishii and Sakai
(2000) argued that flower closure acts primarily to facilitate
male function; specifically, a delay in flower closure allows
pollen to be removed and dispersed more completely. They
suggested that this would explain the minimum floral longevity they observed, regardless of how rapidly pollen was deposited on the stigma. However, they found pollen removal
had no direct effect on flower longevity. Other research has
found a similar lack of effect (Proctor and Harder 1995), al-
though Devlin and Stephenson (1984) and Evanhoe and
Galloway (2002) found that pollen removal did shorten the
duration of the male phase and ultimately flower life span.
We did not measure the effects of pollen removal, although
we think it is unlikely that flower closure functions entirely
to maximize male function in C. angustifolium. Chamerion
angustifolium flowers are strongly protandrous, with anthers
opening over the first 2 d before stigma receptivity. As a
result, in the field, anthers are almost always devoid of any
pollen by the beginning of female phase (B. C. Husband, personal observation). Thus, variation in flower closure in response to pollen deposition would have no measurable effect
on pollen removal.
Flower closure may affect not only the function of individual flowers; it can, in turn, affect the number of open flowers
displayed on an inflorescence. Harder and Johnson (2005)
showed that pollination enhanced flower closure but not the
opening of new flowers in the orchid Satyrium longicauda.
As a result, plants subject to intense pollination displayed
fewer flowers. In doing so, the plants were able to reduce investment in floral display when pollinators were abundant
and enhance attractiveness when they were rare. A similar
process may operate in C. angustifolium because it is also insect pollinated. Pollinator visitation rises with increased floral
display size (Routley and Husband 2003), and flower closure
is accelerated with adequate cross-pollination. However, because of its multiflowered inflorescence and potential for
high geitonogamous self-pollination in large floral displays
(Routley and Husband 2003), selection to reduce floral display size (by reducing flower longevity) may oppose selection
to increase outcrossing in the individual flower (by increasing
floral longevity). The question remains as to how selection
on floral display size interacts with selection for reproductive
function of individual flowers to affect the evolution of floral
longevity in C. angustifolium.
Acknowledgments
We thank Mike Mucci for assistance in the greenhouse,
Dr. C. Caruso for comments on the manuscript, as well as
Holly Sabara, Paul Kron, Sara Wagner, Ryan Geil, and
Graham Smith for advice and technical support. Financial
assistance was provided by a Natural Sciences and Engineering Research Council of Canada Discovery Grant and Canada
Research Chair Award to B. C. Husband.
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