Plant CellPhysiol. 37(3): 239-247 (1996)
JSPP © 1996
Mini Review
Photoinhibition of Photosystem I: Its Physiological Significance in the
Chilling Sensitivity of Plants
Kintake Sonoike
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Hongo, Bunkyo-ku, Tokyo, 113 Japan
Photoinhibition was denned originally as the decrease
in photosynthetic activity that occurs upon excess illumination. The site of photoinhibition has generally been considered to be located in PSII. However, a novel type of
photoinhibition has recently been characterized in chillingsensitive plants. This photoinhibition occurs under relatively weak illumination at chilling temperatures and the main
site of damage is in PSI. The photoinhibition of PSI is initiated by the inactivation of the acceptor side, with the subsequent destruction of the reaction center and the degradation of the product of the psaB gene, which is one of the
two major subunit polypeptides of the PSI reaction center
complex. Chilling and oxidative stress (the presence of reactive species of oxygen) are characteristic requirements for
the photoinhibition of PSI in vivo.
Key words: Chilling sensitivity — Environmental stress —
Photoinhibition — Photosynthesis — Photosystem I — Reactive species of oxygen.
Light, an essential requirement for photosynthesis, is
harmful to the photosynthetic apparatus when illumination
is excessive, and the term, photoinhibition was originally
denned by Kok (1956) as the decrease in photosynthetic activity that is induced by strong light. During the early
stages of research into this phenomenon, inactivation of
both PSI and PSII was reported (e.g., Satoh 1970a, b, c).
However, considerable evidence indicating that PSII is the
site of photodamage accumulated subsequently. Powles
(1984) concluded in his review that the main site of photoinhibition is PSII and that the extent of inhibition of PSI is
much smaller or negligible when compared to that of PSII.
Nowadays, the term photoinhibition is often used as a
Abbreviations: DBMIB, 2,5-dibromo-3-methyl-6-isopropyl-/?-benzoquinone; DMPO, S^'-dimethyl-l-pyrroline-Z^-oxide;
EPR, electron paramagnetic resonance; FNR, ferredoxin-NADP
reductase; Fm, Fs, maximal and steady-state level of Chi fluorescence, respectively; LHCI, LHCII, light-harvesting Chi a/b
protein complexes associating with photosystem I and II, respectively; 4-POBN, a-(4-pyridyl-l-oxide)-JV-f-butylnitrone; SOD, superoxide dismutase; TMPD, N,N,/V,Af'-tetramethyl-p-phenylenediamine.
synonym for "photoinhibition of PSII", since there have
been no reports of photoinhibition of PSI in vivo until
very recently, with the exception of a few studies in algae
(Harvey and Bishop 1978, Gerber and Burris 1981). The activity of PSI has usually been reported to be less strongly
affected than that of PSII, and it has even been reported
to be enhanced after photoinhibitory treatment (Tjus and
Andersson 1993). Photoinhibition of PSII has attracted the
interest of many investigators because of the high turnover
of the Dl protein of the reaction center, or its relationship
with the xanthophyll cycle. Such studies have, moreover,
been stimulated by the development of sensitive methods
for the assessment of PSII activities by monitoring the fluorescence of Chi and by using an oxygen electrode. The photoinhibition of PSI in vitro, reported by Inoue et al. (1986,
1989), tended to be taken as an artifactual phenomenon.
The physiological significance of the photoinhibition of
PSI was first recognized upon the discovery of the selective
inhibition of PSI activity in higher plants in vivo (Terashima et al. 1994, Havaux and Davaud 1994, Sonoike and
Terashima 1994). In this article, the various features of photoinhibition of PSI in algal cells, leaves of higher plants,
isolated thylakoid membranes, and preparations of purified PSI are summarized. The physiological significance of
the photoinhibition of PSI in chilling and oxidative stress is
also discussed in comparison with that of the photoinhibition of PSII.
Recognition of the photoinhibition of PSI in early
studies—Kok (1956) was the first to examine the mechanism of photoinhibition. With his colleagues, he determined
the activity of electron transport in PSI separately from the
activity of whole-chain electron transport after photoinhibitory treatments of isolated spinach chloroplasts. They concluded that PSI is inactivated to a much smaller extent than
PSII upon illumination with high-intensity light (Kok et al.
1965). Further analysis revealed that visible light affected
the photochemical activity of PSI more severely than that
of PSII (Jones and Kok 1966b), while UV light affected
PSII activity with practically no damage to PSI (Jones and
Kok 1966b). The studies by Kok's group revealed several
unique features: (i) photoinhibition of PSI proceeds even in
the presence of DCMU (Kok et al. 1965); (ii) the inhibition
is independent of the presence of oxygen (Jones and Kok
1966a); and (iii) the inhibition occurs even at — 190°C (Kok
239
240
K. Sonoike
et al. 1965). These features clearly distinguish Kok's observations from the later results discussed below. The difference might be attributable to the very high intensity of
light employed by Kok (up to 6 x 106 erg cm" 2 s~', about 15
times the intensity of full sunlight).
Photoinhibition of PSI in algal cells—Harvey and
Bishop (1978) isolated two mutants of a green alga (Scenedesmus obliquus) that were sensitive to strong light and
found that several hours of exposure to strong light
caused a decrease in photosynthetic activity in parallel with
that in PSI activity. PSII activity decreased more slowly in
one mutant and actually increased in the other. The inhibition did not occur in an atmosphere of pure nitrogen and it
was accompanied by a rapid decrease in the amount of photooxidizable P700. Gerber and Burris (1981) also reported
a similar decrease in the level of photo-oxidizable P700
upon photoinhibitory treatment of a marine diatom, Amphora sp. The decrease in the rate of photosynthesis (in
terms of fixation of 14CO2) was greater than that in the level
of P700, suggesting a more serious inactivation of some
other component(s). Upon a shift from strong to less intense light, the recovery of photosynthesis required several
hours, even though the level of photo-oxidizable P700
returned to normal within only 1 h.
Photoinhibition of PSI in leaves of higher plants—
There are only a few reports on the impairment of PSI by
photoinhibitory treatment of leaves of higher plants.
Strong illumination (3.5-5 mmol m~2 s~') of spinach leaves
under a nitrogen atmosphere decreased the amount of
EPR-detectable P700 + , and the decrease in PSI electron
transport from TMPD to methyl viologen was accompanied by the inactivation of PSII (Godde et al. 1992).
The selective photoinhibition of PSI in plant leaves
was not reported until 1994. Weak illumination (100-200
//mol m~2 s~') of cucumber leaves at 4°C for several hours
caused a decrease in the relative quantum yield of electron
transport in PSI from DAD to NADP + with almost no inhibition of PSII activity (Terashima et al. 1994). Biochemical
and biophysical analysis revealed that the acceptor side of
PSI was inactivated first, with subsequent destruction of
P700 and degradation of a subunit of the reaction center
(Sonoike and Terashima 1994). The photosensitive subunit was then identified as the product of the psaB gene
(Sonoike 1996). EPR measurements showed that three
iron-sulfur centers, F A /F B and F x of PSI, were destroyed
by the photoinhibitory treatment (Sonoike et al. 1995b).
This type of photoinhibition of PSI was only observed at
chilling temperatures in the presence of oxygen (Terashima
et al. 1994). Similar photoinhibition of PSI was observed
in another chilling-sensitive plant, Phaseolus vulgaris
(Sonoike et al. 1995a). The extent of inhibition vaxied,
depending on the irradiance at which the plants were
grown: the weaker the growth irradiance, the more sensitive were the plants to photoinhibition of PSI (Sonoike et
al. 1995a).
Photoinhibition of PSI at chilling temperature was
also observed by Havaux and Davaud (1994) in potato
leaves, which are less sensitive to chilling stress than cucumber leaves. Exposure to strong light (900-3,700 ^molm" 2
s~') at 3°C decreased the maximal quantum yield of electron transport in PSII, as determined by pulse-modulated
fluorometry. However, the decrease was insufficient to explain the larger decrease in the rate of oxygen evolution
measured under light-limiting condition. The amount of
photooxidized P700, determined by monitoring absorption
at 820 nm in vivo, decreased, while the Emerson enhancement of oxygen evolution increased, suggesting that PSI is
a rate-limiting step in electron transport (Havaux and
Davaud 1994). These results resemble those of Terashima
et al. (1994), obtained with chilling-sensitive cucumber, in
particular in terms of the requirement for oxygen.
Photoinhibition of PSI in isolated thylakoid membranes—Satoh (1970a, b, c) found that, in thylakoid membranes from spinach, both PSI and PSII were inactivated
under illumination at an intensity that just saturated photosynthesis in the chloroplast. In contrast to the observations
of Kok and his associates, the inactivation of PSI showed a
clear dependence on temperature: the higher was the temperature, the larger was the extent of inhibition (Satoh
1970a). It was demonstrated that photoinhibition of PSI
was sensitive to both anaerobic conditions and DCMU,
while that of PSII was independent of oxygen (Satoh
1970b). Ferredoxin, FNR and plastocyanin were shown not
to be the site of inhibition (Satoh 1970c), which was suggested to be located inside the reaction center complex of
PSI. Barenyi and Krause (1985) also reported inhibition of
the activities of both PSI and PSII by strong illumination
of isolated spinach thylakoids. The inhibition was suppressed by the addition of electron acceptors (CO2 in the case of
intact chloroplasts, methyl viologen in the case of broken
chloroplasts).
Removal of oxygen usually protects PSI from photoinhibition. However, another type of photoinhibition seems
to occur in the absence of oxygen (Satoh and Fork 1982a,
b). The activities of both PSI and PSII decreased when intact chloroplasts from Bryopsis were illuminated under
anaerobic conditions. The decrease in PSI activity was
smaller (13%) than that in PSII activity (27%; Satoh and
Fork 1982a). Satoh and Fork (1982b) concluded that the reducing equivalents that accumulated between ferredoxin
and P700 destroyed the reaction centers and that the presence of oxygen, which serves as an electron acceptor, suppressed the photoinhibition.
Inoue et al. (1986, 1989) examined the site of photoinhibition in PSI complexes under both aerobic and anaerobic conditions. They observed that illumination (30,000
lux) of thylakoid membranes from spinach under aerobic conditions decreased the photoreducing activity of
Photoinhibition of PSI
NADP + to less than 10% of the control while 40% of
the iron-sulfur centers remained intact. Furthermore, the
amount of P700 + oxidized by illumination in the presence
of methyl viologen was decreased by only 20%. From these
results, they concluded that the sites of aerobic photoinhibition are the iron-sulfur centers, which limit the photoreduction of NADP + (Inoue et al. 1986). By contrast, when
the photoinhibitory treatment (3.5 mmol m~2 s"1) was applied under strong reducing conditions, photoreduction of
NADP + decreased without any decrease in the levels of
iron-sulfur centers, phylloquinone and P700 (Inoue et al.
1989), results that suggest that the site of photoinhibition
might be located between Ao and F x .
The selective photoinhibition of PSI in isolated thylakoid membranes from spinach by very weak light was
reported recently (Sonoike 1995). The presence of oxygen
and the flow of electrons from PSII were required for the inhibition, as they were in the earlier studies by Satoh (1970b,
c). However, the intensity of illumination required for the
photoinhibition was very much weaker in the more recent
study. A photon flux density of 10-20^molm~ 2 s~' was
sufficient to decrease the amount of photooxidizable P700
by 50% with little change in the activity of PSII (Sonoike
1995). These observations contrast sharply with all the
earlier reports that showed a significant decrease in PSII activity concomitant with photoinhibition of PSI. In the
studies of isolated thylakoid membranes from spinach, two
characteristic features of the photoinhibition of PSI became apparent. The first was that the product of thepsaB
gene was degraded in vitro (Sonoike 1996), just as occurred in the photoinhibition of PSI in vivo (Sonoike and
Terashima 1994). The second was that the addition of
scavengers of reactive species of oxygen prevented the inhibition of the electron-transfer activity, as well as the degradation of the product of the psaB gene, suggesting the involvement of reactive species of oxygen in this process
(Sonoike 1996).
Photoinhibition of PSI in preparations of purified PSI
—There have been only a few studies of photoinhibition in
preparations of purified PSI. This is not surprising when
we recall that photoinhibition of PSI has been reported to
require the flow of electrons from PSII. There are, however, several reports of the bleaching of Chi in PSI (Miller
and Carpentier 1991, Purcell and Carpentier 1994). A concomitant decrease in the level of chemically determined
P700 was also reported (Purcell and Carpentier 1994).
Baba et al. (1995) noted that very strong illumination (11
mmol m~2 s"1) of PSI complexes resulted in destruction of
the acceptor side of PSI without any decrease in the level of
chemically determined P700. They also observed that photoinhibitory treatments induced degradation of the reaction center proteins and that such degradation could be suppressed by the presence of histidine, a scavenger of singlet
oxygen (Baba et al. 1995). Thus, apparently, very strong
241
light can cause photoinhibition of PSI even in preparations
of purified PSI.
Photoinhibition of PSI and chilling stress—As summarized above, the selective photoinhibition of PSI in vivo is
only observed at chilling temperatures (Terashima et
al. 1994, Havaux and Davaud 1994). In chilling-sensitive
plants, such as cucumber, tomato and common bean, the
rate of photosynthesis (monitored in terms of the oxygen
evolution) decreases sharply below a specific threshold temperature of around 10°C (Powles et al. 1983, Hodgson et
al. 1987, Raison and Brown 1989, see Fig. 1A). This sudden decrease in the rate of photosynthesis is not observed
in chilling-resistant plants, such as spinach (Hodgson et al.
1987; see Fig. 1A, open circles) or in intermediate species,
such as maize (Long et al. 1983). In maize, the rate of photosynthesis decreases as the temperature is lowered, but the
relationship is close to linear (Long et al. 1983). Several factors were examined as possible causes of the strong dependence of the rate of photosynthesis on temperature. For example, dark-chilling treatments decrease the rate of oxygen
evolution in PSII via the liberation of extrinsic PSII proteins and manganese (Shen et al. 1990). Light-chilling treatments caused uncoupling of the ATPase (Terashima et al.
1991a, b). However, both types of damage were reversible,
and neither could explain the irreversible damage to the
photosynthetic machinery by chilling treatment (Terashima
et al. 1991a). The primary reason for the decrease in the
rate of photosynthesis caused by chilling treatment turned
out to be the photoinhibition of PSI (Terashima et al.
1994). The dependence on temperature of the decrease in
the rate of photosynthesis and that of photoinhibition of
PSI are strongly correlated (Fig. IB, closed circles). Moreover, it turns out that recovery from photoinhibition of
PSI in common bean leaves proceeds very slowly for several days (Sonoike et al. unpublished), as might be anticipated from the irreversible nature of the damage to the
photosynthetic machinery. The relative quantum yield of
PSII was also decreased at chilling temperatures, but the dependence on temperature was almost linear, without a specific threshold temperature (Fig. IB, open circles). This
result agrees well with the earlier report that the dependence on temperature was linear, even in chilling-sensitive
plants such as cucumber, when photoinhibition was monitored in terms of the Chi fluorescence of PSII (Hetherington et al. 1989). It is evident that PSI, and not PSII,
limits the overall photosynthetic activity after chilling
treatment of cucumber leaves in the light. Enzymes such
as stromal fructose 1,6-bisphosphatase and sedoheptulose
1,7-bisphosphatase are oxidatively inactivated during chilling treatment in the light, and it has been proposed that
this inactivation is the cause of the damage to the photosynthetic machinery (Wise 1995). However, inactivation of
these enzymes is reversible in vitro (Sassenrath et al. 1990),
and it could, alternatively, be explained by the photoinhibi-
242
K. Sonoike
tion of PSI since these stromal enzymes are subjected to oxidation by oxygen and reducing power must be supplied by
PSI.
Photoinhibition of PSI has also been observed in
potato, which is less sensitive to chilling than cucumber
(Havaux and Davaud 1994). Although the dependence
on temperature of the activity of PSI was not determined,
the decrease in the quantum yield of non-cyclic electron
transport, estimated in terms of (Fm—Fs)/Fm, showed a
gradual decrease between 2°C and 25°C, with no sharp
threshold temperature (Fig. 1C). Such a response is typical
of a chilling-tolerant plant. Although PSI was photoinhibited at chilling temperatures in cucumber and potato
similarly, the role of chilling temperature might be quite
different in the two species.
The strong dependence on temperature observed in
chilling-sensitive plants implies the involvement of phase
transitions of lipids in the inactivation process. In fact, the
phase separation of thylakoid lipids from chilling-sensitive
plants occurs at higher temperatures than that of lipids
Co-100
10
'55
a>
|
I
2o
common bean
a. BO
"B
1
CD
I
CD
rx
0
10
20
Temperature during treatment (°C)
0
10
20
Temperature during treatment (°C)
100-
0
10
20
Temperature during treatment (°C)
3-100-
0
10
20
Temperature during treatment (°C)
Fig. 1 Dependence of photosynthetic activity on the temperature at which photoinhibitory treatments were applied.
Panel A: The
rate of photosynthesis in slices of spinach and cucumber leaf [open circles and closed circles, respectively, taken from Fig. 2 of Hodgson
et al. (1987)], in slices of tomato leaf [closed triangles, taken from Fig. lb of Raison and Brown (1989)] and of attached leaves of common bean [closed squares, taken from Fig. 4B of Powles et al. (1983)]. Panel B: The relative quantum yield of electron transfer through
PSI and PSII (DAD to NADP + and H2O to 2,6-dimethyl-p-quinone, respectively) in thylakoid membranes from photoinhibited cucumber leaves [closed circles and open circles, respectively, taken from Fig. 1 of Terashima et al. (1994)]. Panel C: The quantum yield of
linear electron transfer [determined as (Fra—Fs)/Fm] in potato leaves [open squares, taken from Fig. 2 of Havaux and Davaud (1994)].
Panel D: The rate of photosynthesis in wild-type tobacco plants (open lozenges) and tobacco plants transformed with a gene for
glycerol-3-phosphate acyltransferase of squash (open triangles). Data are redrawn from Figure 4 A-C (points after incubation for 10 h)
of Moon et al. (1995). The broken lines in panels B-D are the same curve as the solid line in panel A.
Photoinhibition of PSI
from chilling-tolerant plants (Fork et al. 1981). The inhibition of photosynthesis starts at temperatures at which the
polar lipids from thylakoid membranes undergo a phase
separation (Hodgson et al. 1987). A correlation between
the chilling sensitivity of plants and the extent of unsaturation of fatty acids in phosphatidylglycerol from leaves
has been suggested (Murata et al. 1982), and the genetic
manipulation of the unsaturation of fatty acids has been
reported to alter the chilling sensitivity of plants (Murata et
al. 1992, Kodama et al. 1995). Introduction of a gene for
glycerol-3-phosphate acyltransferase from squash, which is
a chilling-sensitive plant, into tobacco plants resulted in increased damage upon chilling treatment in the light (Moon
et al. 1995). The dependence on temperature of photosynthesis was almost linear in both wild-type and transgenic
plants, and it was only the slope of the line that increased in
transgenic plants (Fig. ID). Apparently, the linear dependence on temperature observed with decreased rate of photosynthesis in chilling-tolerant or intermediate species can be
closely correlated with the lipid composition of the plant
membranes. The dependence on temperature of photoinhibition of PSII in chilling-sensitive plants can also be explained by the lipid composition. However, the question
still remains as to the primary factor responsible for the
sudden decrease in the rate of photosynthesis at a certain
specific temperature in such chilling-sensitive plants as cucumber or tomato.
Selective photoinhibition of PSI, observed in isolated
thylakoid membranes, provides a clue to the answer to the
above question. Inhibition is observed at room temperature, as well as at chilling temperatures, in this case, and inhibition can be observed even with thylakoid membranes
from spinach, a chilling-tolerant plant (Sonoike 1995).
This observation suggests the following. First, PSI is readily photoinhibited in vitro irrespective of temperature during treatment. Second, some mechanism exists to protect
PSI in vivo, and the mechanism is inactivated during the
isolation of thylakoid membranes. Third, the protective
mechanism is chilling-sensitive factor in chilling-sensitive
plants. Apparent dependence on temperature of the damage to PSI in vivo must be brought about indirectly by
the dependence on temperature of protective mechanism.
Thus, the protective component, which might be associated
with thylakoid membranes, is supposed to be the chillingsensitive factor in chilling-sensitive plants and might be responsible for the strong dependence on temperature.
Photoinhibition of PSI and reactive species of oxygen
—In many cases, the photoinhibition of PSI has been reported to require oxygen and the flow of electrons from
PSII, both in vivo (Terashima et al. 1994, Havaux and
Davaud 1994) and in vitro (Satoh 1970b, Inoue et al. 1986,
Sonoike 1995). The major site of superoxide production in
chloroplasts is the reducing side of PSI (Asada 1994), and
enzymes that scavenge reactive species of oxygen, such as
243
SOD and ascorbate peroxidase, are localized at or near the
PSI reaction center (Miyake and Asada 1992, Ogawa et al.
1995). It has also been reported that the generation of superoxide by xanthine/xanthine oxidase in darkness results
in a decrease in iron-sulfur centers and PSI activity in isolated thylakoid membranes (Inoue et al. 1986). It seems reasonable to assume that PSI is susceptible to the attack by reactive species of oxygen. Selective photoinhibition of PSI
in isolated thylakoid membranes (Sonoike 1995) enabled
us to determine the effects of scavengers of reactive species
of oxygen on the photoinhibition of PSI. Two spin-trap
reagents, DMPO and 4-POBN, and n-propyl gallate, a
scavenger of hydroxyl radicals, were effective in suppressing the photoinhibition of PSI (Sonoike 1996). Photoinhibition of PSI was not induced when the iron-sulfur centers
were oxidized by the addition of methyl viologen (Sonoike
1996). It was also reported that hydroxyl radicals were produced through the interaction of reduced ferredoxin with
hydrogen peroxide (Asada and Takahashi 1987). Thus, we
can speculate that hydroxyl radicals, produced at reduced
iron-sulfur centers in PSI, immediately destroyed these
centers once hydrogen peroxide escapes from the system
that scavenges reactive species of oxygen.
When reactive species of oxygen are scavenged in thylakoid membranes, electrons flow from H2O to H2O (Asada
1994, see also Fig.3A). In this pathway (the so-called
Asada pathway), oxygen accepts electrons from PSI and
produces superoxide, which is scavenged by SOD and ascorbate peroxidase, being converted to H2O in a reaction that
involves reduced ferredoxin, with no net change in oxygen
and electron. Therefore, the presence of oxygen facilitates
the dissipation of excess light energy, provided that the
scavenging system is intact. The results by Satoh and Fork
(1982b) who demonstrated the enhanced photoinhibition
of PSI in very intact chloroplasts from Bryopsis in the
absence of oxygen, can be explained by the decreased
dissipation of energy via the Asada pathway which requires
the presence of oxygen. By contrast, in all the studies that
demonstrated the photoinhibition of PSI in the presence of
oxygen, experiments were performed with broken chloroplasts in which the scavenging system for reactive species
of oxygen may no longer have been intact (Satoh 1970b,
Inoue et al. 1986, Sonoike 1995, 1996). In these preparations, the anaerobic conditions protected PSI from photoinhibition, rather than enhancing it. The Asada pathway
did not function for the dissipation for energy in these
cases, and the presence of oxygen promoted photoinhibition through the generation of reactive species of oxygen.
In intact cucumber leaves, photoinhibition of PSI was only
observed at chilling temperatures, implying that some components) of the Asada pathway might be the chilling-sensitive factors.
Proposed mechanism for photoinhibition of PSI—A
hypothetical scheme for the mechanism of photoinhibition
K. Sonoike
244
of PSI is shown in Figure 2. The first proposed event is the
temperature-dependent loss of the protective mechanism.
If reducing equivalents are accumulated under unprotected
conditions, oxygen is reduced on the acceptor side of PSI
to produce superoxide and other reactive species of oxygen. These reactive species of oxygen attack the iron-sulfur
centers, F A /F B and F x (Sonoike et al. 1995b). A, might also
be attacked (Sonoike et al. 1995b), and P700 is destroyed
as well under stronger light (Terashima et al. 1994). Inactivation of these components triggers the specific degradation of the product of the psaB gene, one of the subunits of
the PSI reaction center (Sonoike 1996). The turnover of the
protein might be necessary for the recovery from photoinhibition because restoration of PSI activity requires several
days (Sonoike et al. unpublished).
Photoinhibition of PSI exhibits some similarities to
"acceptor side" photoinhibition of PSII (Aro et al. 1990).
In both cases, inactivation of the acceptor side is induced
first and is followed by damage to the reaction center and
degradation of a subunit of the reaction center. Although
the reaction centers of both PSI and PSII are composed of
two homologous subunits, only one of them is selectively
degraded upon photoinhibition: the product of the psaB
gene in PSI and that of thepsbA gene (Dl protein) in PSII.
Two phylloquinone molecules are bound to the PSI complex, as are two plastoquinone molecules in PSII, but only
one of them serves as an electron acceptor (A)) in the case
of PSI. The specific degradation of the product of the psaB
gene as a consequence of the inactivation of the acceptor
side implies that this protein serves a ligand for Aj. It must
be noted, however, that there is a distinct difference between PSI and PSII. The Dl protein turns over very rapidly (Greenberg et al. 1987), and recovery from photoinhibition of PSII occurs within several hours, in contrast to the
slow turnover of PSI. Thus, in a physiological sense, photoinhibition of PSI seems to be far more harmful to plants
than that of PSII.
Photoinhibition of PSII protects PSI from photoinhibition—What are the conditions for the photoinhibition of
PSI? There are many reports that PSI is not the site of photoinhibition. The rate of electron transfer from ascorbate/
DCIP to methyl viologen in thylakoids was reported to be
unchanged by treatment with strong light of attached
leaves of cucumber (Critchley 1981) and Nerium oleander
(Powles and Bjorkman 1982), of cyanobacteria (Tytler
et al. 1984, Yokoyama et al. 1991) and of dinoflagellates
(Prezelin et al. 1986). When applied in combination
with chilling stress, photoinhibitory treatment of attached
Recovery over the
course of several days
PsaA
P700
y\.
Temperature-dependent loss of
protective mechanism
Degradation of PsaB
Light
Destruction of iron-sulfur
centers (and possibly of Ai)
Reduction of acceptor
components and oxygen
Generation of dangerous
reactive species of oxygen
Fig. 2 A hypothetical scheme for the mechanism of photoinhibition of PSI.
P700, reaction-center chlorophyll of PSI; Ao, A] and
F x , primary (chlorophyll), secondary (phylloquinone) and tertiary (iron-sulfur center) electron acceptors in PSI; F A /F B , terminal (ironsulfur centers) electron acceptors in PSI; PsaA and PsaB, products of thepsaA andpsaB genes, respectively. "Hands" in the scheme represent putative mechanism for protection of PSI.
Photoinhibition of PSI
leaves of pumpkin (Tyystjarvi et al. 1989) and of detached
leaves of spinach (Somersalo and Krause 1990) also had
only a slight effect on the rate of electron transfer to methyl
viologen. Assessment of PSI activity in vivo by a photoacoustic technique or by measurement of Emerson enhancement in Chlamydomonas cells (Canaani et al. 1989) and in
attached pea leaves (Havaux and Eyletters 1991) also gave
no indication of any damage to PSI. All the authors cited
above assigned the site of inhibition to PSII, except in the
case of the chilling treatment of pumpkin leaves in the light
in which the decrease in the rate of total photosynthesis
could not be fully explained by the decrease in PSII activity
(Tyystjarvi et al. 1989). We might ask why PSII was inactivated instead of PSI in these cases, if photoinhibition of
PSI really has physiological relevance. There are several
possible reasons. First, it is evident that the very strong
light causes inactivation of PSII. Second, the photoinhibition of PSI in vivo might not be observed at room temperature or in chilling-tolerant plants. Third, in many of the
studies, PSI activity was assessed in terms of the electron transfer to methyl viologen. At a high concentration,
methyl viologen can accept electrons from partially destroyed PSI (Fujii et al. 1990, Sonoike and Terashima
1994), so that the activity of PSI might have been overestimated in such measurements. In fact, 70% of the electron-
245
transfer activity to methyl viologen was maintained in thylakoid membranes from photoinhibited cucumber leaves in
which the rate of photoreduction of NADP + was only 2030% of the control (Terashima et al. 1994). Finally, photoinhibition of PSI should be suppressed when PSII is inactivated by strong light.
Photoinhibition of PSI in leaves of higher plants is induced (i) at chilling temperatures (0-10°C), (ii) in the presence of oxygen, (iii) by exposure to relatively weak light
(about 100//molm" 2 s" 1 ), (iv) for a relatively long time
(about 5 h), (v) in chilling-sensitive plants such as cucumber, and (vi) when the flow of electrons from PSII proceeds
normally. In isolated thylakoid membranes, photoinhibition of PSI can be induced at any temperature and even in
chilling-tolerant plants. In any event, weak illumination is
essential for the selective photoinhibition of PSI. Stronger
illumination induces photoinhibition of PSII and, thus,
might protect PSI. However, very strong light induces inactivation of PSI independently of PSII (as seen in the photoinhibition of PSI in preparations of purified PSI) so that
both PSI and PSII can be photoinhibited. It should be also
noted that plastoquinone and other redox components between PSI and PSII may be reduced once PSI is photoinhibited. Under such conditions, PSII is subject to the danger
of "acceptor side photoinhibition" and the light intensi-
PSI
Cyclic
electron
flow
*2H2O
Fig. 3 Mechanism for protection of PSI from photoinhibition.
A: Protection of PSI from photoinhibition by the Asada pathway.
Excess light energy is dissipated by the flow of electrons from H2O to H2O. B: Without protection, PSI is damaged by the reducing
pressure from PSII. C: PSI is protected when PSII activity is down-regulated by the gradient in pH produced by the cyclic electron flow
around PSI. D: PSI is protected when PSII is inactivated. Fd, Ferredoxin; Fdrcd, reduced ferredoxin; Asc, ascorbate; APX, ascorbate
peroxidase; MDA, monodehydroascorbate.
246
K. Sonoike
ty necessary for the photoinhibition of PSII may become
weaker than is usually necessary. Therefore, photoinhibition of PSII protects PSI from photoinhibition while photoinhibition of PSI might enhance the photoinhibition of
PSII. This phenomenon may provide another explanation
of the fact that selective photoinhibition of PSI was not
discovered until very recently.
Several mechanisms have been proposed as ways of
dissipating excess light energy: the Asada pathway (see previous section); functional dissociation of LHCII from the
PSII core by protein phosphorylation; the xanthophyll cycle; the cyclic flow of electrons around PSI; down-regulation of PSII activity by the gradient in pH across thylakoid
membranes; and even the degradation of Dl protein. The
Asada pathway may have a primary role in protecting PSI
from photoinhibition (Fig. 3A). The Asada pathway operates not only to relieve over-reduction of PSI but also to
generate a gradient in pH that leads to regulation of the
electron flow from PSII (Schreiber et al. 1995). Without
the pathway and with intact PSII, PSI is subject to photoinhibition (Fig. 3 B). Barbato et al. (1992) reported that functional dissociation of LHCI from the PSI core complex
occurred after photoinhibitory treatment of spinach thylakoid membranes. Although no such phenomenon has been
observed in cucumber leaves (Sonoike and Terashima
1994), it is possible that the inactivation of PSI might be
due to the inability to dissociate LHCI at chilling temperatures. The recent report that the amount of LHCI decreased with increases in growth irradiance in the red alga
Porphyridium cruentum (Tan et al. 1995) is of interest in
this context. Components of the xanthophyll cycle were
reported to be present in a PSI fraction, and the conversion
from violaxanthin to zeaxanthin was also observed (Thayer
and Bjorkman 1992, Lee and Thornber 1995). Although
the role of the xanthophyll cycle that operates in PSI has
been totally neglected to date, the cycle might also serve to
protect PSI. Cyclic electron flow around PSI can dissipate
the light energy absorbed by PSI, but the reducing power
should be still accumulated when the flow of electrons from
PSII is not decreased. Therefore, a decrease of PSII activity by the gradient in pH is necessary to protect PSI from
photoinhibition (Fig. 3C). The degradation of the DI protein and inactivation of PSII can also be regarded as the
mechanism for the protection of PSI (Fig. 3D). The dissipation of excess light energy has been discussed mainly in
terms of protection against photoinhibition of PSII. However, down-regulation of PSII activity might be also important for protection against the photoinhibition of PSI,
which is harder for plants to reverse and is, thus, more
harmful to them.
The author thanks Dr. I. Terashima for fruitful collaboration
throughout the studies of photoinhibition of PSI, as well as for
the encouragement to write this article and helpful discussions.
The author also thanks Dr. S. Itoh and Ms. M. Iwaki for critical
reading of the manuscript and Dr. K. Asada, Dr. M. MiyaoTokutomi, Dr. H. Sakurai, Dr. S. Katoh and Ms. Y. Hihara for
helpful discussions.
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(Received December 4, 1995; Accepted March 11, 1996)