MARINE ECOLOGY PROGRESS SERIES Vol. 183: 73-86.1999 1 Published July 6 Mar Ecol Prog Ser
Photoinhibition and photoprotection in symbiotic dinoflagellates from reef-building corals
Ove Hoegh-Guldberg*, Ross J. Jones
School of Biological Sciences, Building A08. The University of Sydney, New South Wales 2006. Australia
ABSTRACT: Pulse-amplitude-modulation fluorometry and oxygen respirometry were used to investi- gate die1 photosynthetic responses by symbiotic dnoflagellates to light levels in summer and winter on a high latitude coral reef. The symbiotic dinoflagellates from 2 species of reef-building coral (Porites cylindnca and Stylophora pistillata) showed photoinhibitory decreases in the ratio of variable (F,) to maximal (F,) fluorescence (F,/F,,,) as early as 09:00 h on both summer and winter days on the reefs associated wlth One Tree Island (23"30'S, 1.52" 06' E; Great Barrier Reef, Australia). This was due to decreases in maximum, F,, and to a smaller extent minimum, F,, chlorophyll fluorescence. Complete recovery took 4 to 6 h and began to occur as soon as light levels fell each day. Chlorophyll fluorescence quenching analysis of corals measured during the early afternoon revealed classic regulation of photo- system I1 (PSII) efficiency through non-photochemical quenching (NPQ). These results appear to be similar to data collected for other algae and higher plants, suggesting involvement of the xanthophyll cycle of symbiotic dinoflagellates in regulating the quantum efficiency of PSII. The ability of symbiotic dinoflagellates to develop significant NPQ, however, depended strongly on when the symbiotic dinoflagellates were studied. Whereas symbiotic dinoflagellates from corals in the early afternoon showed a significant capacity to regulate the efficiency of PSII using NPQ, those sampled before sun- rise had a slower and much reduced capacity, suggesting that elements of the xanthophyll cycle are suppressed prior to sunrise. A second major finding of this study is that the quantum efficiency of PSII in symbiotic dinoflagellates is strongly diurnal, and is as much as 50 % lower just prior to sunrise than later in the day. When combmed with oxygen flux data, these results indicate that a greater portion of the electron transport occurring later in the day is likely to be due to the increases in the rate of carbon fixation by Rubisco or to higher fluxes through the MeNer-Ascorbate-Peroxidase (MAP) cycle.
KEYWORDS. Corals . Symbiotic dinoflagellates . Chlorophyll fluorescence - Photoinhibition - Pocillo- porin Colour morphs
INTRODUCTION 400 nm) has been shown to have a variety of destruc- tive effects on marine organisms (Jokiel 1980), includ- Coral reefs thrive in the shallow sunlit seas of the ing corals and their symbiotic dinoflagellates (Shick et tropics. The high levels of solar radiation that are char- al. 1996). Effects of ultraviolet (UV) radiation on cul- acteristic of this environment are important both as an tured symbiotic dinoflagellates include decreased energy source and as a physiological challenge to the growth rate, cellular chlorophyll a, carbon:nitrogen organisms that live there (Shick et al. 1996). Reef- ratios, photosynthetic oxygen evolution and ribulose building corals are no exception, with light being bisphosphate carboxylase/oxygenase (Rubisco) activ- essential to their dinoflagellate symbionts (Symbio- ity (Banazak & Trench 1995, Lesser 1996). Similar dinium spp.), yet posing a potential problem to both effects have been reported for symbiotic dinoflagel- host and symbiont. Short wavelength radiation (290 to lates living within cnidarian tissues (Jokiel & York 1982, Lesser 1989, Shick et al. 1991, 1995, Gleason 1993, Gleason & Wellington 1993, Kinzie 1993, Banazak & Trench 1995). Both host and symbiont have been reported to have a range of protective mecha-
0 Inter-Research 1999 Resale offull article not permitted 7 4 Mar Ecol Prog Ser
nisms to counteract the direct and indirect influences Early studies of photosynthetic activity using mea- of UV radiation. These include the production of surements of oxygen flux and/or incorporation of 14C mycosporine-like amino acids for which a UV absorb- identified a decrease in the maximum photosynthetic ing function has been inferred, and a range of active rate (P,,,,,) at high photon flux rates. Decreases in P,,,, oxygen scavenging systems (for a review, see Shick et at high PAR, consequently, have been used by eco- al. 1996). physiologists as an indication of photoinhibition in a Photosynthetically active radiation (PAR) is also wide variety of studies (Falkowsh et al. 1994). While potentially detrimental to photosynthetic organisms. photoinhibition has been found in many photosynthetic Photosynthetic processes saturate rapidly at relatively organisms, curiously, it has rarely been seen in the low levels of flux. Above these levels, light energy is many studies of the photosynthetic activity of symbiotic potentially damaging to the photosynthetic machinery dinoflagellates within the tissues of cnidarians (Long et (Walker 1992, Foyer et al. 1994, Long et al. 1994, al. 1994, but see Shick et al. 1996 for a rare exception). Osmond 1994). Classically, photoinhibition has been Photoinhibition at high PAR, for example, was noted interpreted in terms of 'damage' sustained to photo- for symbiotic dinoflagellates isolated from the sea synthetic systems and was measured originally as a anemone Aiptasia pulchella, but not for the symbiotic long-lasting decrease in photosynthetic rate (oxygen dinoflagellates when studied within the tissues of the flux) at high irradiances (Kok 1956). This view of host (Muller-Parker 1984). A similar example is noted photoinhibition has been developed during the past 40 by Goiran et al. (1996) for the symbiotic dinoflagellate yr to represent one in which damage is primarily asso- coral Galaxea fasciculans. Other studles of the pho- ciated with photosystem I1 (PSII), particularly with the toynthetic response curves of symbiotic cnidarians increased loss of the D1 protein of the PSII reaction mostly lack the otherwise characteristic decrease in centre (Walker 1992, Styring & Jegerschold 1994, P,,, at the highest PAR (Falkowski & Dubinsky 1980, Telfer & Barber 1994). Recent work, however, has also Wyman et al. 1987, Chalker et al. 1988, Hoegh-Guld- associated changes in photosynthetic conversion effi- berg & Smith 1989a,b, Davies 1991, Harland & Davies ciencies with an array of photoprotective phenomena 1995). The apparent absence of chronic photoinhibition (Osmond 1994) that make the question of what is during experiments with symbiotic cnidarians has pro- damage as opposed to the down-regulation of photo- voked suggestions that the specialised environment synthetic systems equivocal (Long et al. 1994, Osmond within the cells of symbiotic dinoflagellates and their 1994). The latter has been referred to as 'dynamic' symbiotic cnidarians might act to reduce or eliminate (down regulation) as opposed to 'chronic' (damage) photoinhibition (Downton et al. 1976, Long et al. 1994). photoinhibition (Osmond 1994, Osmond & Grace 1995). In this regard, the high concentrations of active oxygen The photoprotective mechanisms associated with scavenging systems (superoxide dismutase, SOD; cata- dynamic photoinhibition span a wide range of phe- lase, CAT; ascorbate oxidase. ASPX) in symbiotic dino- nomena that are reversible over a range of physiologi- flagellates and the surrounding host tissues are particu- cal time scales. Energy-dependent quenching of PSII, larly provocative (Lesser 1996). for example, may take seconds to come into play and is This paper investigates photoinhibition and associ- thought to achieve photoprotection by changing the ated phenomena in symbiotic dinoflagellates from 2 aggregation of light harvesting complex I1 (LHCII) par- species of reef-building coral. By using the techniques ticles, or by reversibly deactivating individual PSII of pulse-amplitude-modulation (PAM)fluorometry and reaction centres (Duysens & Sweers 1963, Schreiber & oxygen respirometry, we have been able to clearly Bilger 1987). Other changes may take minutes (e.g. demonstrate that photoinhibition occurs at relatively xanthophyll cycle inter-conversions, Long et al. 1994), modest light levels, and is a prominent and normal hours (e.g. changes in the expression of oxygen characteristic of the photosynthetic processes of the scavenging components), or days (e.g. developmental symbiotic dinoflagellates in these corals. We also changes, Long et al. 1994). These changes decrease report variations In key photoprotective processes in redox pressure at various points in photosynthetic symbiotic dinoflagellates which indicate that the electron transport, and act as uncoupling mechanisms photosynthetlc activity is far more dynamic in symbio between incoming light energy and subsequent redox than has been previously thought. outcomes. The analogy to the 'Victorian governor' by Walker (1992) is particularly applicable to the current understanding of how increased redox pressure across MATERIALS AND METHODS PSII can ultimately induce negative feedback on light capturing efficiencies in the form of reduced photosyn- Collection and maintenance of corals. This research thetic efficiencies and thereby reduced damage from was conducted on expeditions to One Tree Island excessive light levels. (23" 30' S, 152" 06' E; Great Barrier Reef, Australia) in Hoegh-Guldbel-g & Jones: Photo]mhibition in reef-buildlng corals 75
March and June 1997, and involved experiments on then placed in darkness and F,/F, determined in 3 the reef-building corals Porites cylindnca and Sty- fragments at 1, 2.5, 5, 10, 20 and 40 min intervals. lophora pistillata, including both the 'brown' and AF/F, were measured for the same corals after incu- 'pink' colour morphs of the latter (pink colour morphs bating them for 1 min at each of a series of 8 irradi- contain high quantities of the animal pigment pocillo- ances that increased in steps froin 120 to 2340 pm01 porin, Dove et al. 1995). All corals were collected or m-2 s-I PAR. F and F,' were measured at the end of studied from a uniform depth of 30 to 40 cm (mean low each 1 min illumination period, AFIF, (I") from this water). Corals were either studied in situ on reefs experiment were used to calculate apparent relative immediately adjacent to the One Tree Island Research electron transport rates (ETR). This calculation was Station (OTIRS), or were brought back to the research based on the incubation irradiance (photon flux den- station and maintained in a flow-through aquarium sity, PFD) and the assumption that half the photons system. In the latter case, single finger-like branches required for the movement of electrons through the (4 to 8 cm in length) were removed from mature coral photosystems of symbiotic dinoflayellates are ab- colonies growing at the top of the inner reef slope of sorbed by PSII (Schreiber et al. 1994; ETR = Y' X PFD X the One Tree Island lagoon. The corals were immedi- 0.5).The validity of this calculation for symbiotic dino- ately transported to the laboratory where they were flagellates is supported by other research that shows mounted into small plastic racks using non-toxic that calculated ETR values are approximately linear plastic modelling clay. Generally, corals were left to with photosynthetic rates measured simultaneously recover for 2 to 3 d in the flow-through aquarium prior from oxygen flux measurements made with a respiro- to use in the experiments. meter (Hoegh-Guldberg & Jones unpubl.). ETR data Measurements of the quantum yield of PSII. The was plotted against incubation irradiance to provide changes in the fluorescence characteristics of symbi- light response curves for Porites cylindrica as a func- otic dinoflagellates in Porites cylindrica and Stylophora tion of time of day. pistillata were followed in 2 experiments that involved The experiment was repeated during the June 1997 measurements over the diel cycle. The basic parame- expedition (austral winter) using both Porites cylin- ters of chlorophyll a fluorescence associated with drica and Stylophora pistillata. In this case, the experi- PSII were measured in colonies of P. cylindnca at vari- ment was done on corals housed in the flow-through ous intervals over 36 h in the field (sampling times: seawater system at OTIRS under natural irradiances. 08:OO-09:OO h, 14:OO-15:OO h, 18:OO-19:OO h, 01:OO- The chlorophyll fluorescence characteristics were 02:OO h) during the first experiment, done over 2 d in measured at each time point on 4 pieces of P. cylindrica early March 1997 (late austral summer). Replicate, but and brown and pink colour morphs of S. pistillata. Irra- still attached, sections of each colony (3 to 5 replicate diance levels in this experiment were monitored every colonies per sample time; different colonies were used 15 min using a LICOR L1-189 photometer with a LI- at each measurement time) were wrapped in foil to 190SA quantum sensor. Temperature was measured exclude light for 30 min to dark-adapt them prior to using a mercury bulb thermometer. Measurements measurement. The minimal (Fo) and maximal (F,) fluo- were done hourly from pre-dawn to midnight over 3 rescence yields were measured using a PAM chloro- day-night cycles and included the dark-acclimation of phyll fluorometer (DIVING-PAM, Walz, Germany; see corals for 30 min prior to measuring F. and F, as Schreiber et al. 1997 for specifications). A weak pulsed described above. In this experiment, a separate set of red light (
culating water jacket. Corals were placed within each chamber over a perforated floor, underneath xvhich was a stir-bar powered by a magnetic stirrer Two 50 W quartz-halogen lights illuminated each chamber from opposite sides. Changes in the concentration of oxygen within the chambers were measured using Clark-type polarographic oxygen electrodes (Strathkelvin Instru- ments, Glasgow, UK)that were inserted into the top of each chamber. The oxygen sensors were connected via polarising units to an analogue-to-digital converter 0.20 1 , , , , (ADC-1, Remote Measurement Systems, Seattle, USA) 0 10 20 30 40 and an IBM-compatible 486-laptop computer running T~rne(min) data acquisition software (Sable Systems, Los Angeles, USA). Oxygen concentrations were measured every 20 S from an average of 2 consecutive voltage readings from each sensor. The voltage responses of the sensors were calibrated using air-saturated seawater at the incubation temperature and salinity, and oxygen purged (nitrogen bubbled) seawater (34 %0 salinity). Software (Temptabl, Oxford University Press, 1986) was used to calculate the oxygen concentrations at sat- uration for each experimental temperature and salin- ity. Photosynthetic rates (at 750 pm01 m-' S-' PAR for 20 min) and then respiratory rates (in dark for 30 min) were measured for 4 corals at each time point. Follow- Time of day (h) ing each dark measurement, F,,/Fm were measured as described above. The corals were then frozen and Fig. 1. (a) Changes in F,,/F,,, in symbiotic dinoflagellates of Stylophora pistillata (brown morphs) exposed to >l000 pm01 taken back to the University of Sydney, where the quanta m-2 S-' for 2 h during the middle of the day and then number of symbiotic dinoflagellates and surface area placed in darkness for different intervals (x-axis) prior to were measured. This was done by stripping the tissues measurement. Data are means 2 SE, n = 3 independent reph- off the underlying skeleton using a jet of recirculated cates at each time point. (b) Dark-adapted F,/F,,, of symb~ot~c dinoflagellates measured within the tissues of Porites cylin- seawater (Johannes & Wiebe 1970) and measuring the drica in early March 1997 on reefs at One Tree Island density of symbiotic dinoflagellates in the resulting (23" 30' S, 152" 06' E). Specimens were dark-adapted for homogenate using a hemacytometer (8 separate cham- 30 min prior to each measurement. Data are means * SE ber counts).
imen and the optical cone of the fluorometer (Schrei- ber et al. 1997). The quenching characteristics of the RESULTS symbiotic dinoflagellates in each coral was investi- gated during each incubat~on in response to actinic Changes in F,, F,,, and dark-adapted F,IF, light of 800 pm01 m-2 S-' PAR. Photochemical [qP = (F,,,'- F)/(F,'- Fo')] and non-photochemical [NPQ = Fv/Fmof the symbiotic dinoflagellates in fragments of (Fm- F,,,')/F,,,'] quenching components were calculated Stylophora pistillata (brown colour morphs) exposed to in addition to maximum F,/F,,,, maximum AFIF,, F,, > 1000 pm01 quanta m-' S-' for 2 h during the middle of F. and F,', which were measured during each experi- the day increased rapidly as a function of the length of mental run (Schreiber & Bilger 1987). F. (observed as a time m darkness (dark-adapted; Fig. la). Most change downward drift in F) was observed to undergo slow in dark-adapted F,/F, occurred in the first 5 to 10 min but significant quenching (downward drift) in symbi- of dark-adaptation, after which there was little change. otic dinoflagellates sampled prior to dawn. qP was cal- Accordingly, a 30 mm period of dark-adaptation was culated using F. corrected for the downward drift considered suitable for su.bsequent measurements of [Fo' = F. X F(Oj/F(n), where F(0) is F at the beginning dark-adapted Fv/Fm.The dark-adapted FJF,, of sym- of the Light period, and F(n) is the value of F at time = n]. biotic dinoflagellates in Porites cylindnca and both Respirometry. Photosynthetic and respiratory rates colour morphs of S. pistillata showed distinct changes were measured using a respirometer that consisted of over the course of the day during both experiments 4 separate 90 m1 acrylic chambers enclosed in a reclr- (Flgs. lb & 2a,b). During the June 1997 study, dark- Hoegh-Guldberg & Jones: Photoinhibitlon in reef-building corals 7 7
0,8 ,a Porites cylindrica F. and F,, used to calculate dark-adapted Fv/Fm, T changed as a function of light level during the die1 cycle (Fig. 3 a,b;June 1997). Both F. and F, decreased with increasing light levels and slowly returned to their pre-dawn values by approximately 20:OO h. Changes in F,, were slightly less marked (decreasing to 60% of pre-dawn values on the first day) than changes in F, (decreasing to 40% of pre-dawn values during the first day). F, also showed some tendency to drift 0, b Stylophora pistillat? T- downward during the course of the experiment, espe- cially during the first 2 days.
Quenching analysis
Chlorophyll fluorescence quenching analysis was used to investigate regulatory phenomena associated c Irradiance with symbiotic dinoflagellates that were exposed to 1600 1
I
Time of day (h)
Fig. 2. Dark-adapted F,/F,,, of symbiotic dinoflagellates mea- sured within the tissues of (a) Pontes cylindrica and (b) Sty- lophora pistillata (+ = pink colour morphs, = brown colour morphs) over the course of 3 d at One Tree Island in June 1997 Specimens were dark-adapted for 30 min before mea- surement. Data are means i SE for 4 colonies of P. cylindnca and 4 colonies of each colour morph of S. pistillata. (c) Light levels (PAR) were measured beside colonies. Temperatures within the experunents were: Day 1 = 22 * l.O°C (mean * SD), Day 2 = 22 3 + l.O°C, Day 3 = 21.1 t 0.5"C
adapted Fv/Fm dropped from 0.65 in both species to less than 0.5 by 12:OO h on the first day. Dark-adapted F,/F, generally decreased as light levels increased occurring on each day and the lowest values coincided with the highest irradiances (Fig. 2a to c). Dark- adapted Fv/Fmbegan to increase again as light levels began to decrease in the early afternoon, and had returned to their pre-dawn values by approximately 20:OO h each evening. Water temperatures within the Time of day (h) experiment did not vary by much over the 3 days Fig. 3. (a) Minimal fluorescence yields (Fo) and (b) maximal (mean + SD: Day = 22 + l.O°C, Day 2 = 22.3 + l.O°C, 1 fluorescence yields (F,) of symbiotic dinoflagellates in Pontes Day 3 = 21.1 + 0.5"C). Interestingly, dark-adapted cylindnca (A)and Stylophora pistillata (* = pink colour F,/F,,, increased to 0.7 in both species just after dawn morphs, = brown colour morphs) measured over 3 d at One (between 07:OO and 08:OO h) on the second and third Tree Island in June 1997. Colon~eswere dark-adapted for days. This coincided with light levels that were be- 30 min before measurement. Data are means * SE for 4 colonies of P. cylindrica and 4 colonies of each colour morph of 20 100 tween and pm01 m-2 S-' PAR. They had dropped S. pistillata. (c) Light levels (PAR) measured beside colonies. to 0.65 again by 09:OO h, when light levels were gen- Temperatures within the experiments were: Day 1 = 22 * l.O°C erally above 300 pm01 m-' S-' PAR. (mean * SD),Day 2 = 22.3 + l.O°C, Day 3 = 21.1 * 0.5"C 78 Mar Ecol Prog Ser 183: 73-86, 1999
Symbiotic dinoflagellates from the 2 corals sampled pre-dawn versus in the early afternoon also showed other differences in dynamics of the fluorescent tran- sient following induction by modest light levels. Per- haps the most interesting of these was the tendency for dark fluorescence (F; an approximation of Fol) to remain well above F. (measured just before the end of the dark period) for longer in the pre-dawn samples. The differences were quite marked, with F remaining above F, for at least 6 min in the pre-dawn corals while 2 4 6 dropping to F. within a few seconds in symbiotic Time (min) dinoflagellates from corals sampled in the early after- noon (Fig. 5a,b). Trends were strongest for Porites
0.4 b Stylophora pistillata cylindrica, yet were still significant (p < 0.05) though less pronounced in Stylophora pistillata. Regulatory phenomena and the ease of electron flow beyond PSII (e.g. activation of Calvin-Benson cycle enzymes) ultimately determine the quenching of exci- tations in PSII by the photochemistry of the symbiotic dinoflagellates. qP is a measure of the average oxida-
Time (min)
Fig. 4. Comparison of quenching responses (non-photochem- ical quenching [NPQ] of photosystem 11) of symbiotic dmo- flagellates in (a) Pon'tes cylindrica and (b) the brown colour morphs of Stylophora pistillata as a function of time of day in June 1997. Four corals of each species were sampled at 06:OO h [prior to sunnse) and 14:OO h (following the solar zenith). Shown are means [n = 4) and 95% confidence limits (dashed lines) Time (min)
light for different lengths of time. Th~sprocedure al- t b Stylophora pistillata lows the identification of specific control mechanisms "l5 1,; that underlie changes in quantum efficiency. Symbi- otic dinoflagellates in corals that were measured prior to sunnse had distinctly different quenching character- istics to those that had been exposed to light for an ex- tended period, over the day (compare 06:OO h with 14:OO h measurements, Fig. 4a,b; p < 0.05). Symbiotic dinoflagellates from corals that had been exposed to natural irradiances from sunrise until the early after- noon showed a classic pattern of regulation of energy flux through PSII via NPQ (compare Fig. 4a, 14:OOh measurement with Fig. 9a of Schreiber & Bilger 1987, Time (min) p. 40). Symbiotic dinoflagellates from pre-dawn corals, however, had reduced abilities to rapidly quench PSII Fig. 5. Comparison of quenching responses (difference be- using NPQ (Fig. 4a,b). This is shown most dramatically tween F and Fo) of symbiotic dinoflagellates in (a) Porites in Pontes cylindrica, which showed rapid induction of cylindrica and (b) the brown colour morphs of Stylophora pis- tillataas a function of time of day in June 1997. Four corals of NPQ in the early afternoon, but showed a much slower each specles were sampled at 06:OO h (prior to sunrise) and induction of the same phenomenon in the symbiotic di- 14:OO h (following the solar zenith). Shown are means (n = 4) noflagellates in corals sampled prior to dawn (Fig. 4a). and 95% confidence limits (dashed lines) Hoegh-Guldberg & Jones: Photoinhibition in reef-building corals 79
cn a Porites cylindrica 300 .-c 1.01
L 0 2 4 6 E am Time (rnin) aa 0
m b Stylophora pistillata .E 1.0 l pmol quanta m-2s-1PAR
Time (rnin)
Fig. 6. Cornpanson of quenching responses (photochemcal quenching [qP]) of symbiotic dinoflagellates in (a) Porites cylindrica and (b) the brown colour morphs of Stylophora pis- tillata at One Tree Island as a function of time of day in Tlme of day (h) June 1997. Four corals of each species were sampled at 06:OO h (prior to sunrise) and 14:00 h (following the solar Fig. 7. Electron transport rates (ETR) of symbiotic dinoflagel- zenith). Shown are means (n = 4) and 95% confidence limits lates in Pontes cylindrica measured in the field at One Tree (dashed lines) Island in March 1997 (a) Apparent ETR as a functlon of irra- diance at 09:OO h and at 15:OO h. Shown are means (n = 4) and 95% conf~dence limits (dashed hnes). (b) Apparent ETR at light saturation as a function of time of day. Shown are the means of duplicate measurements from individual colonies. tion state of the primary acceptor pool, Q,. The greater Trends are indicated by line of best fit the value of qP, the higher the throughput of energy to acceptors downstream of this redox pool. qP was higher (p < 0.05) initially in symbiotic dinoflagellates from light-acclimated (i.e. early afternoon) colonies of pinned by changes in AF/F,. AFIF, after exposure to both coral species (Fig. 6a,b). Notably, after 6 min of a series of light intensities (culminating in 1555 pm01 PAR irradiation, the qP of symbiotic dinoflagellates m-2 S-' PAR increased from dawn [06:00 h] to values from pre-dawn and early afternoon corals were no that are almost 2-fold higher by about 09:OO h longer statistically different. (Fig. 8a,b; both species, winter).
Electron transport rate and maximum effective yields Oxygen flux measurements at saturation and as a function of time of day The gross rate of photosynthesis (Pg)per dinoflagel- Light response curves (ETR vs irradiance) showed a late cell (range 1.1 to 1.8 pg O2 cell-' h-') in colour consistent pattern of change over the course of the day morphs of Stylophora pistillata did not vary as a func- for both species. In all cases, the capacity of symbiotic tion of time of day (Fig. 9a). Dark-adapted F,/F, mea- dinoflagellates for electron transport increased from sured at the same time showed the characteristic vari- sunrise to midday and then decreased gradually to ation with light level seen previously in this study sunset (Fig. ?a,b). These changes in ETU are under- (Fig. 9b; see also Fig. 2b). 80 Mar Ecol Prog Ser 183: 73-86, 1999
a Porites cylindrica 1987, Falkowski et al. 1994). In Porites cylindnca and l the 2 colour morphs of Stylophora pistillata, relatively minor PAR fluxes (200 to 400 pm01 m-* S-' PAR) resulted in significant reductions (p < 0.01) in dark- adapted Fv/Fm(Figs. 1 & 2). The lowest values of dark- adapted F, IF, occurred during the highest light levels and trends were identical in both coral species. Conse- quently, the symbiotic dinoflagellates in the 2 species of reef-building coral studied here clearly exhibited photoinhibition. 0600 1200 1800 2400 The quantity of incoming PAR had a large influence Time of day (h) on dark-adapted F,,/Fm on the second and third days of the study (Fig. 2). Light levels on the second day were b Stylophora pistillata 0.207 strongest in the morning and were reduced by cloud during the afternoon. Dark-adapted F,IF,,, in this case dropped quickly to approximately 0.5 by midday and then rose slowly back to 0.65 a few hours after light levels had decreased in the early afternoon. Opposite trends were evident for the third day. In this case, clouds reduced light levels in the morning and the nor- mal decrease in dark-adapted Fv/Fm began later, but
"."U , , . 'l 0600 1200 1800 2400 Time of day (h)
Fig. 8. Maximum effective yields (ANF,) at the end of a series of eight 1 min irradiances that increased and culminated in 1 rnin of irradiation at PAR of 1555 pm01 photon m-' S-' for symbiotic dinoflagellates from (a) Porites cylindrica and (b) the brown colour morphs of Stylophora pistillata as a function of time of day at One Tree Island in June 1997. Shown are means (n = 4)a.nd 95 D/oconf~dence lmits (dashed lines)
DISCUSSION
The results of this study clearly show that symbiotic Time of day (hour) dinoflagellates experience photoinhibition in symbio, and are similar to other marine plants that have been examined (Falkowski et al. 1994, Franklin et al. 1996). Our studies also highlight the dynamic nature of the photosynthetic machinery of sym.biotic dinoflagellates of corals and indicate that photoinhibitory modifica- tions of the quantum efficiency occur even on winter days for corals growing on a high latitude reefs.
Photoinhibition in symbiotic dinoflagellates
Photolnh.ibition is defined as any decrease in the Time of day(hour) capacity of a photosystem to capture and process pho- tons that is caused by incoming light (Long et al. 1994, Fig. 9. (a) Gross photosynthetic rates of colonies of the brow, Osmond 1994). In studies of PSI1 chlorophyll fluores- (e)and pink (m)morphs of Stylophora pistillata as a function of time of day. (b) Maximum potential quantum yield (F,/F,) cence this is synonymous with light-related decreases of the same colonies. Specimens were dark acclimated for in the dark-adapted Fv/Fm, a measure of the maximum 30 min prior to each measurement. Data are means * SE for potential quantum yield in plants (Schreiber & Bilger 4 colonies of each morph Hoegh-Guldberg & Jones: Phol :oinhibition in reef-building corals 81
eventually reached a value in the early afternoon that thin in marine algae (Franklin et al. 1992, 1996). Simi- was similar to that seen earlier on the first day. Similar lar decreases in F, occur as the ratio of DT to DD changes have been seen for higher plants (e.g. increases in both diatoms (Arsalane et al. 1994, Cheeseman et al. 1997), macroalgae (Henley et al. Olaizola & Yamamoto 1994) and free-living dinoflagel- 1992, Hanelt et al. 1993) and other microalgae lates (Johnson & Sakshaug 1993). Changes in the ratio (Falkowski et al. 1994, Vassiliev et al. 1994). It is of DT to DD in symbiotic dinoflagellates of the coral notable that the changes in dark-adapted F,IF,,, Goniastrea aspera have recently been correlated with reported here for symbiotic dinoflagellates in corals the light environment of corals growing intertidally in occurred under the low light conditions of a cloudy Thailand (Ambarsari et al. 1997). These results further winter day on a high latitude (23" 30's) coral reef. suggest that xanthophyll cycle inter-conversions are Changes in dark-adapted F,/F,, reflect the energy likely to be central to the acclimative and adaptive conversion efficiency or capacity of PSII to process abilities of symbiotic dinoflagellates in the face of high captured light photochemically (Schreiber & Bilger PAR conditions and stresses such as those during ther- 1987, Long et al. 1994, Schreiber et al. 1996) and are mally related bleaching (Jones et al. 1998). due to changes in the composition and organisation of the light harvesting complex associated with PSII (Demmig-Adams et al. 1989, Krause & Weis 1991, Variability in quantum efficiency and Walker 1992, Horton & Ruban 1994, Long et al. 1994, non-photochemical quenching Bukhov et al. 1996, Young & Frank 1996). Almost iden- tical responses in dark-adapted F,IF,,, F, and F. for Symbiotic dinoflagellates sampled in the early after- symbiotic dinoflagellates in corals at One Tree Island noon (after most of a day of sunlight) were able to more have been reported for macroalgae at the same loca- rapidly engage NPQ of PSII excitations when undergo- tion and time of year (Franklin et al. 1996). In this case, ing a transition from dark to light (Fig. 4a,b). The sig- decreases in dark-adapted F,,/F,, and F. were shown nificantly slower decay of the baseline fluorescence to be correlated with the conversion of violaxanthin (F-F,, Fig. 5a,b) In the pre-dawn dinoflagellates when into zeaxanthin by the enzyme violaxanthin epoxidase. exposed to moderately high actinic light (800 pm01 m-2 These 2 carotenoids were prominent in the xantho- S-' PAR) also indicates a much slower induction of phyll cycle of most of the macroalgae studied by energy-dependent quenching. The efficient engage- Franklin et al. (1996). An increased ratio of zeaxanthin ment of NPQ requires a sufficiently large pool of epox- to violaxanthin is thought to increase the rate of ther- idase (Demmig-Adams et al. 1989). The present study mal dissipation away from PSII, thereby decreasing the indicates that epoxidase levels are probably depleted amount of energy flowing towards the reaction centre (or deactivated) during the night such that the engage- (Demmig-Adams et al. 1989, for review, see Young & ment of NPQ was much slower just prior to dawn than Frank 1996).The avoidance of over-reduction of reac- in the early afternoon. Similarly, it appears that epoxi- tion centres and electron transport components dase levels increase (or existing epoxidase is reacti- reduces the production of active forms of oxygen vated) in symbiotic dinoflagellates as they experience (Asada & Takahashi 1987), and thereby reduces the elevated light levels. In concert with changes in the risk of subsequent oxidative damage. Dinoflagellates ability to engage NPQ were the expected differences and diatoms have different but comparable com- in qP. In this case, symbiotic dinoflagellates sampled pounds (monoepoxide diadinoxanthin, DD, and dia- prior to light exposure had lower qP than those sam- toxanthin, DT) which appear to serve the same roles as pled after most of a day of sunlight (Fig. 6a,b). These violaxanthin and zeaxanthin and within the light har- differences indicate that a greater proportion of the vesting structures of these organisms (Olaizola & acceptor pool (Q,) was available in symbiotic dinofla- Yamamoto 1994, Olaizola et al. 1994, Young & Frank gellates that had been exposed to light for some time. 1996, Rhiel et al. 1997). The decreases in F. during the qP indicates the proportion of captured light energy present study are also indicative that the de-epoxida- that is used for processes requiring electrons down- tion of xanthophyll pigments like DD are involved in stream of PSII. By the analysis so far, the prediction is increasing the thermal dissipation of energy away that the transport of electrons beyond PSII would be from PSII. F. is a measure of the initial energy distrib- depressed earlier in the day. This was indeed the case ution within PSII and of the efficiency of energy trap- when the rate of apparent ETR was investigated as a ping by P680 (Schreiber & Bilger 1987). Decreases in function of time of day. Maxinlunl effective yields or Fo, consequently, indicate that the increased dissipa- ETR for symbiotic dinoflagellates in both Porites cylin- tion of energy within the light harvesting antennae are dnca and Stylophora pistillata measured at light-satu- probably associated with the formation of DT as they ration (>l000 pm01 m-2 S-' PAR) varied significantly are strongly correlated with the formation of zeaxan- (p < 0.05)during the course of the die1 cycle (Figs. 7 & 8). 82 Mar Ecol Prog Ser 183:73-86, 1999
Just prior to dawn, light-saturated ETR were lower by The second possible explanation for the change in comparison to later in the day by as much as 50%. ETR in the 2 species over the day is that Rubisco I1 Light-saturated ETR increased until maxima occurred could function more as an oxygenase (photorespira- in the early afternoon in concert with the declines tion) as opposed to a carboxylase. Oxygenase activity measured in F,/F, discussed previously. Rates de- as a possible means of photoprotecting photosystems clined slowly overnight, however, indicating that the has been seen in some higher plant studies (for review, processes involved took about 12 h to recover from see Walker 1992, Bichler & Fock 1996, Heber et al. light exposure during the day. The increases in the 1996). Lovelock & Winter (1996), for example, esti- AF/F,,, (after exposure to high light levels) during the mated that excess excitations were mainly processed middle of the day indicate that Q, is less reduced (and through this pathway in light-exposed, outer-canopy electron flow beyond PSI1 more efficient) for a given leaves of several species of tropical rainforest trees. photon influx than earlier in the day. There are several The contribution of other pathways like the Mehler- possible reasons for the more efficient processing of Ascorbate Peroxidase cycle (MAP cycle) was estimated excitations by symbiotic dinoflagellates during the to represent only 10 to 20% of the CO2-independent early afternoon. The first possibility is that light expo- electron transport. The oxygen respirometry data col- sure evokes the up-regulation of processes that are lected during the present study allows some crucial related to carbon fixation by Rubisco. Increased insight into whether added oxygenase activity by enzyme activity could relate to the increased expres- Rubisco I1 can explain the light-related increases in sion or activation-state of existing enzymes, or to the electron transport of symbiotic dinoflagellates. changes in their activity caused by local availability of Although electron transport is almost double in the substrates such as carbon dioxide. Changes in the middle of the day as opposed to just prior to sunrise, availability of substrates and the actlvity of enzymes the rates of net oxygen flux measured at same time do that deliver substrates to Rubisco (e.g. carbonic anhy- not vary (Fig. 9b). This tends to mitigate the possibility drase) can also have a significant effect on the rate of that photorespiration by Rubisco I1 oxygenase activity photosynthetic fixation of inorganic carbon, as shown has been stimulated, as this would lead to higher rates for symbiotic dinoflagellates in sea anemones (Weis of oxygen consumption and hence a lower net outward 1993). The light-dependent modulation of Rubisco oxygen flux in the light. To achieve the doubling of activity has also been established in terrestrial plant electron transport observed, the respiratory fluxes studies (for review, see Geiger & Servaites 1994). Flux would almost completely annihilate outward oxygen through the Calvin-Benson cycle varies with irradi- fluxes. This did not happen. Some caution, however, ance through changes in the activation-state of should be exercised here. The methods used to mea- Rubisco and at least 2 other light-activated enzymes, sure oxygen fluxes have an inherent time delay and phosphoribulokinase (EC 2.7.1.19) and glyceraldehyde may fail to measure the responses measured by PAM 3-phosphate dehydrogenase (EC 1.2.1.13). Changes fluorometry. In the latter case, measurements are are of the order of magnitude that would match the made over time resolutions on the order of seconds, changes seen in the symbiotic dinoflagellates over the while it takes at least 30 min to obtain a flux measure- course of the day. For example, in the study of Sage et ment. As changes in the maximum quantum yield are al. (1993), the ratio of initial to fully activated Rubisco likely to be dynamic on shorter time scales, the meth- activity declined by 40 to 50% within 60 min of light ods used to measure oxygen flux may lead to the fail- reduction in 2 species of higher plants. These changes ure to measure the changes seen by the PAM fluoro- were due to the decarbamylation of Rubisco as a regu- meter. latory response to the reduction in Light. Measuring The third possibility is that light-adapted symbiotic changes in the activation-state of Rubisco in symbiotic d.inoflagellates may have greater ETR through non- dinoflagellates is exceptionally difficult given the Rubisco catalysed pathways (Foyer et al. 1994). In this inherent instability of the unusual Rubisco I1 found in case, pathways in which oxygen IS the ultimate elec- symbiotic dinoflagellates (Whitney & Yellowlees 1995). tron acceptor such as the MAP cycle could be stimu- However, initial studies over the diel cycle indicate lated in response to the greater need to dissipate that the expression of Rubisco I1 does not change in energy away from the photosystems under high light symbiotic dinoflagellates from Tridacna gjgas (D.Yel- conditions (Schreiber & Neubauer 1990, Neubauer & lowlees, James Cook University, pers. comm.) How- Yamamoto 1992, Henning et al. 1993, Cheeseman et ever, nothing is known as to the activation state of the al. 1997, Gee1 et al. 1997). In the MAP cycle, oxygen enzyme over diel cycle or how putative carbon concen- reduction results in superoxide and H20z production, trating mechanisms (Yellowlees pers. comm.) might catalysed by superoxide dismutase (SOD). H20zis then affect the flux of energy through the Calvin-Benson reduced by ascorbate in a reaction catalysed by ascor- cycle. bate peroxidase (ASPX). The monodehydroascorbate Hoegh-Guldberg & Jones: Phcbtoinhibition in reef-building corals 83
(MDA)radical that is formed by the ASPX reaction also adapted F,/F,,, and oxygen evolution rates at high light acts an electron acceptor regenerating ascorbate. The intensities (for example, Hanelt et al. 1993), symbiotic MAP cycle provides a valve reaction in which O2 and dinoflagellates in symbiotic anemones (Muller-Parker H202(via MDA) serve as electron acceptors, and may 1984) and corals (Downton et al. 1976) appeared origi- also lead to down-regulation of PSI1 by causing acidifi- nally to be immune to the effects of high light. These cation of the thylakoid lumen (Schreiber & Neubauer rates are consistent with those of the present study that 1990). As with changes in Rubisco, the responses here show no decline in photosynthetic oxygen flux. The could also be related to the expression and/or activa- resolution of this feature of respirometry studies of tion-state of the enzymes, or to the concentration of algal cnidarian symbioses may lie in their complex substrates like ascorbate in the chloroplast. The exis- structure. Photosynthetic studies of symbiotic dinofla- tence of elements of the MAP cycle has been known gellates while resident in corals and other cnidarians for some time in investigations of oxidative stress in are con~plicated by the typically dense morphology of symbiotic dinoflagellates and their coral hosts (Shick & these associations. Light levels vary by as much as 50- Dykens 1985, Shick et al. 1995). SOD is the metalloen- fold from sites within a colony to sites located at the zyme responsible for catalysing the conversion of tips of the branches (Titlyanov 1991). Light levels are superoxide into H202 after the initial formation of likely to vary considerably within the coral tissues as a superoxide near PSI. The Cu and Zn containing forms result of algal self-shading, especially when the coral is of SOD are sensitive to cyanide (CN-) and are associ- in a retracted state (Lasker 1979). The consequences of ated with the chloroplast stroma (Salin 1987). CN- sen- this for whole-colony photosynthetic studies are that sitive SOD decreases with depth (from 3 to 30 m) in the rate measurements represent the integration of both symbiotic dinoflagellates of at least 1 species of Acrop- saturated and unsaturated sites within a colony or tis- ora (Shick et al. 1995). Similar decreases in the sues. Consequently, photoinhibitory decreases as light enzymes, catalase and ASPX, which are responsible levels are increased are Likely to be offset by the higher for converting H202into H20 and 02, also occurred in rates of photosynthesis stemming from the increased the same samples of symbiotic dinoflagellates. Experi- penetration of light to the more shaded sites within a ments on cultured symbiotic dinoflagellates (Symbio- colony. diniurn burrnudense, Lesser 1996) have revealed Pink colour morphs of Stylophora pistillata contain strong responses in SOD and ASPX when cultures are the hydrophilic protein-dimer pocilloporin (Dove et al. exposed to UV as opposed to being protected from it. 1995). The physiological function of pocilloporin is not These responses are increased at higher temperatures. known, although its expression within Pocillopora High culture temperature alone induced higher levels darnicornis is inducible by PAR as opposed to UV Light of SOD and ASPX activity (Lesser 1996). These results (Takabayashi & Hoegh-Guldberg 1995). It is not clear, taken together with those from the higher plant litera- however, whether pocilloporin acts as a photoprotec- ture suggest that the components of the superoxide tant (Dove et al. 1995). The present study has revealed scavenging system in symbiotic dinoflagellates are that both pink and brown colour morphs of S. pistillata likely to respond to increased PAR, and in this way displayed remarkably similar degrees of photoinhibi- provide extra capacity for electron transport via the tion and recovery. The function of pocilloponn remains MAP cycle. Changes in the activity of either carbon fix- equivocal and is currently under intense study. ation via the Calvin-Benson cycle or the MAP cycle Symbiotic dinoflagellates appear to have different pathway are good candidates for explaining why ETR abilities with regard to accommodating the large fluc- increase in symbiotic dinoflagellates from corals sam- tuations of PAR in their habitats (Warner et al. 1996). pled in the early afternoon. Warner et al. (1996) reported that symbiotic dinoflagel- lates from the Caribbean coral Siderastrea radians were better able to use NPQ to reduce the damage to Ecological implications photosystems during heat exposure than symbiotic dinoflagellates from Montastrea annularis. In the pre- Photoinhibition is a part of the daily challenges faced sent study, there are also differences between the sym- by most photosynthetic organisms (Walker 1992, Long biotic dinoflagellates from the 2 species of corals with et al. 1994). Light, while beneficial, may lead to the respect to the extent to which NPQ develops during over-reduction of components of the electron transport light exposure. It is interesting to speculate that if Fitt chain when in excess. This may lead to the increased & Warner (1995) and Warner et al. (1996) (see also production of superoxide or singlet oxygen, and even- Lesser 1996) are correct in their conclusions then sym- tually to oxidative damage. Initial studies of symbiotic biotic dinoflagellates with greater abilities to deploy dinoflagellates presented a different picture. Whereas NPQ should be more resistant to thermally related macroalgae consistently showed a decline in dark- bleaching. Energy-dependent regulation of photosyn- 84 Mar Ecol Prog Ser
thetic electron transport is emerging as an important Downton WJS, Bishop DG, Larkum AWL, Osmond CB (1976) process by which symbiotic dinoflagellates may reduce Oxygen inhibition of photosynthetic oxygen evolution in the impact of heat-stress-related declines in electron marine plants. Aust J Plant Physiol 3:73-79 Duysens LNM, Sweers HE (1963) The mechanism of two pho- processing wlthin the dark reactions of photosynthe- tochemical reactions in algae studied by means of fluores- sis (coral bleaching, Hoegh-Guldberg & Smith 1989b, cence. In: Studies of microalgae and photosynthetic bacte- Jones et al. 1998). The differences in susceptibility ria. University of Tokyo Press, Tokyo, p 353-372 among symbiotic dinoflagellates seen in this study are Falko~vskiPG, Dubinsky Z (1980) Light-shade adaptation of Stylophora pistillata, a hermatypic coral from the Gulf of supported by field studies which show that corals in Eilat. Nature 289:172-174 the family Poritidae are much less likely to bleach than Falkowski PG, Greene R, Kolber Z (1994) Light utilization and corals from the families Acroporidae or Pocilloporidae photoinhibition of photosynthesis in marine phytoplank- (e.g. Hoegh-Guldberg & Salvat 1995). ton. In: Baker NR, Bowyer JR (eds) Photoinhibition of pho- tosynthesis: from molecular mechanisms to the field. BIOS Scientific Publishers, Oxford, p 403-430 Acknowledgements. The authors would like to acknowledge Fitt IVK, Warner ME (1995) Bleaching patterns of four species of Caribbean reef corals. Biol Bull 187:298-307 the capable and ready help of 1Mr Mark Waugh and MS Robyn Foyer CH, Lelandais M, Kunert KJ (1994) Photooxidative Sweetapple-Waugh, who managed One Tree Island Research stress In plants Physiol Plant 92:696-717 Station during the period of this study. We also thank the com- Franklin LA, Levavasseur G, Osmond CB, Henley WJ (1992) ments of 2 anonymous reviewers and the excellent editorial Two components of the onset and recovery during pho- assistance of Dr George Humphrey. toinhibition of Ulva rotundata. Planta 186:399-'08 FranMin LA, Seaton GGR, Lovelock CE, Larkum AWD (1996) Photoinhibltlon of photosynthesis on a coral reef. Plant LITERATURE CITED Cell Environ 19:825-836 Gee1 C, Versluis W, Snel JFH (1997) Estimation of oxygen Ambarsan I, Brown BE, Barlow RG, Britton G, Cummings D evolution by manne phytoplankton from measurement of (1997) Fluctuations in algal chlorophyll and carotenoid the efficiency of Photosystem I1 electron flow. Photosynth plgments during solar bleaching in coral Gonjastrea aspera Res 51:61-70 at Phuket, Thailand. Mar Ecol Prog Ser 159:303-307 Geiger DR, Servaltes JC (1994) Dynamics of self-regulation of Arsalane W, Rousseau B, Duval JC (1994) Influence of the photosynthetlc carbon metabolism. Plant Physiol Biochem pool size of the xanthophyll cycle on the effects of light 32:173-183 stress In a diatom-competition between photoprotection Gleason DF (1993) Dlfferentlal effects of ultraviolet radiation and photoinhibition. Photochem Photobiol 60:237-243 on green and brown morphs of the Caribbean coral Porites Asada K, Takahashi M (1987) Production and scavenging of astreoides. Lirnnol Oceanogr 38:1452-1463 active oxygen in photosynthesis. In: Kyle DJ, Osmond CB, Gleason DF, Wellington GM (1993) Ultraviolet radiation and Arntzen CJ (eds) Photoinhibition. Elsevier Science Pub- coral bleaching. Nature 365:836-837 lishers, Amsterdam, p 227-287 Goiran C, Al-Moghrabi S, Allemand D, Jaubert J (1996) Inor- Banazak AT. Trench RK (1995) Effects of ultraviolet (UV) radi- ganic carbon uptake for photosynthesis by the symbiotic ation on marine microalgal-invertebrate symbioses. 1. coraVdinoflagellate association. I. Photosynthetic perfor- Response of the algal symbionts in culture and in hospite. mances of symbionts and dependence on sea water bicar- J Exp Mar Biol Ecol 194:213-232 bonate. J Exp Mar Biol Ecol 199:207-225 Bichler K, Fock H (1996) Evidence for the contribution of the Hanelt D, Huppertz K, Nultsch W (1993) Daily course of pho- Mehler-peroxidase reaction in dissipating excess elec- tosynthesis and photoinhibition in marine macroalgae trons in drought-stressed wheat. Plant Physiol 112: investigated in the laboratory and field. lMar Ecol Prog Ser 265-272 97:31-37 Bukhov NG, Wiese C, Neimanis S, Heber U (1996) Control of Harland AD. Davies PS (1995) Symbiont photosynthesis in- photosystem I1 in spinach leaves by continuous light and creases both respiration and photosynthesis in the symbi- by light pulses given in the dark. Photosynth Res 50: otic sea anemone Anemonia rriridis. Mar Biol 123:715-722 181-191 Heber U, Gerst U, Kreiger A, Neimanis S, Kobayashi Y (1996) Chalker BE, Barnes DJ, Dunlap WC, Jokiel PL (1.988) Light Coupled cyclic electron transport in intact chloroplasts and reef-building corals. Interdisc Sci Rev 13:222-237 and leaves of C3 plants: does it really exist? If so, what is Cheeseman JM, Herendeen LB. Cheesem.an AT, Clough BF ~tsfunction? Photosynth Res 46:269-275 (1997) Photosynthesis and photoprotection in mangroves Henley WJ, Lindley ST, Levavasseur G, Osmond CB, Ramus J under field conditions. Plant Cell Environ. 20:579-588 (1992) Photosynthetic response of Ulva rotundata to light Davies PS (1991) Effect of daylight variations on the energy and temperature during emersion on an intertidal sand budgets of shallow-water corals. Mar Biol 108:13?-144 flat. Oecologla 89:516-523 Dernmig-Adams B, Adams \VW, Winter K, Meyer A, Henning H, Neubauer C, Asada K, Schreiber U (1993) Intact Schreiber U, Periera JS, Kruger A, Czygan FC, Lange OL chloroplasts display pH5 optimum of 02-reduction in the (1989) Photochemical efficiency of photosystem 11, photon absence of methyl viologen: indirect evidence for a regu- yield of 0, evolution, photosynthetic capacity, and latory role of superoxide protonation. Photosynth Res 37: carotenoid composition during the midday depression of 69-80 net CO2 uptake in Arbutus unedo growing in Portugal. Hoegh-Guldberg 0, Salvat B (1995) Periodic mass bleaching Planta 177:377-387 of reef corals along the outer reef slope in Moorea, French Dove S, Takabayashi M, Hoegh-Guldberg 0 (1995) Isolation Polyne~la.Mar Ecol Prog Ser 121:181-190 and partial characterisation of the pink and blue pigments Hoegh-Guldberg 0, Smith GJ (1989a) Th.e influence of the of pocilloporid and acroporid corals. Biol Bull 189:288-297 population density of zooxanthellae and supply of ammo- Hoegh-Guldberg & Jones: Photoii nhibition in reef-building corals 85
nium on the biomass and metabolic characteristics of the Osmond CB (1994) What is photoinhibition: some insights reef corals Seriatopora hystrix (Dana 1846) and Stylophora from comparisons of sun and shade plants. In: Baker NR, pistillata (Esper 1797). iMar Ecol Prog Ser 57:173-186 Bowyer JR (eds) Photoinhibition of photosynthesis: from Hoegh-Guldberg 0, Smith GJ (1989b) The effect of sudden molecular mechanisms to the field. BIOS Scientific Pub- changes in temperature, irradiance and salinity on the lishers, Oxford, p 95-1 10 population density and export of zooxanthellae from the Osmond CB, Grace SC (1995) Perspectives on photoinhibition reef corals Stylophora pistillata (Esper 1797) and Seriato- and photorespiration in the field: quintessential inefficien- pora hystrix (Dana 1846). J Exp Mar Biol Ecol 129: cies of the light and dark reactions of photosynthesis? 279-303 J Exp Bot 46:1351-1362 Horton P, Ruban A (1994) The role of light harvesting con~plex Rhiel E, Marquardt J, Eppard h?, Morschel E, Krumbein WE 11 in energy quenching. In. Baker NR, Bowyer JR(eds) Pho- (1997) The light harvesting system of the diatom tomhibition of photosynthesis: from molecular mechanisms Cyclotella cryptica-isolation and characterization of the to the field. BIOS Scientific Publishers, Oxford, p 11 1-128 main light harvesting complex and evidence for the exis- Johannes RE, Wiebe WJ (1970) A method for determination of tence of minor pigment proteins. Bot Acta 110:109-117 coral tissue biomass and composition. Limnol Oceanogr Sage RF, Reid CD, Moore BD, Seemann JR (1993) Long-term 21540-547 kinetics of the light-dependent regulat~onof ribulose-1,5- Johnson G, Sakshaug E (1993) Biooptical characteristics and bisphosphate carboxylase oxygenase activity in plants photoadaptive responses in the toxic and bloom-forming with and without 2-carboxyarabinitol l-phosphate. Planta dinoflagellates Gyrodin~umaureolum, Gymnodinium gala- 191.222-230 theanurn, and two strains of Prorocentrum nu'nimum. Salin ML (1987) Toxic oxygen species and protective systems J Phycol29:627-642 of the chloroplast. Physiol Plant 72:681-689 Jokiel PL (1980) Solar ultraviolet radiation and coral reef epi- Schreiber U, Bilger W (1987) Rapid assessment of stress fauna. Science 20?:1069-1071 effects on plant leaves by chlorophyll fluorescence mea- Jokiel PL, York RH (1982) Solar ultraviolet photobiology of the surements. In- Tenhunen JD et al. (eds) Plant responses to reef coral Pocillopora damicornjs and symbiotic zooxan- stress. Springer Verlag, Berlin thellae. Bull Mar Sci 32:301-315 Schreiber U, Neubauer C (1990) 02-dependent electron flow, Jones R, Hoegh-Guldberg 0, Larkum AWD, Schreiber U membrane energization and the mechanism of non-photo- (1998) Temperature induced bleaching of corals begins chemical quenching of chlorophyll fluorescence. Photo- with impairment of dark metabolism in zooxanthellae. synth Res 25.279-293 Plant Cell Environ 21:1219-1230 Schreiber U, Bilger W, Neubauer C (1994) Chlorophyll fluo- Kinzie RA (1993) Effects of ambient levels of solar ultraviolet rescence as a nonintrusive indicator for rapid assessment radiation on zooxanthellae and photosynthesis of the reef of in vivo photosynthesis. In: Schulze ED, Caldwell h4M coral R4ontipora verrucosa. Mar B101 116:319-327 (eds) Ecophyslology of photosynthesis. Springer-Verlag, Kok B (1956) On the inh~b~tlonof photosynthesis by intense Berlin, p 49-70 light. Biochim Biophys Acta 21:234-244 Schreiber U, Kuhl M, Klimant I, Reising H (1996) Measure- Krause GH, Weis H (1991) Chlorophyll fluorescence: the ment of chlorophyll fluorescence within leaves using a basics. Annu Rev Plant Physiol Plant Mol Biol 42:313-349 modified PAM fluorometer with a fibre-optic microprobe. Lasker HR (1979) Light-dependent activity patterns among Photosynth Res 47:103-109 reef corals: Montastrea annularis. Biol Bull 156:196-211 Schreiber U, Gademan R, Ralph PJ, Larkum AWD (1997) Lesser MP (1989) Photobiology of natural populations of zoox- Assessment of photosynthetic performance of Prochloron anthellae from the sea anemone Aiptasia pallida: assess- in Lissoclonium patella in hospite by chlorophyll fluores- ment of the host's role in protection against ultraviolet cence measurements. Plant Cell Physiol 38:945-951 radiation. Cytometry 10:653-658 Shick JM, Dykens JA (1985) Oxygen detoxification in algal- Lesser MP (1996) Elevated temperatures and ultraviolet radi- invertebrate symbioses from the Great Barrier Reef. ation cause oxidative stress and inhibit photosynthesis in Oecologia 66:33-41 symbiotic dinoflagellates. Limnol Oceanogr 41:271-283 Shick JM, Lesser MP, Stochaj WR (1991) Ultraviolet radiation Long SP, Humphries S, Falkowski PG (1994) Photoinhibition and photooxidative stress in zooxanthellate Anthozoa: the of photosynthesis in nature. Annu Rev Plant Physiol Plant sea anemone Phyllodiscus semoni and the octocoral Mol Biol 45633-662 Clavularia sp. Symbiosis 10:145-173 Lovelock CE, Winter K (1996) Oxygen dependent electron Shick JM, Lesser MP, Dunlap WC, Stochaj WR, Chalker BE, transport and protection from photoinhbition in leaves of Wu Won J (1995) Depth dependent responses to solar tropical tree species. Planta 198580-587 ultraviolet radiation and oxidative stress in the zooxan- Muller-Parker G (1984) Photosynthesis-irradiance response thellate coral Acropora microphthalma. Mar Biol 122: and photosynthetic periodicity in the sea anemone Aipta- 41-51 sia pulchella and its zooxanthellae. Mar Biol82:225-232 Shick JM, Lesser MP, Jokiel PL (1996) Effects of ultraviolet Neubauer C, Yamamoto HY (1992) Mehler-peroxidase reac- radiation on corals and other coral reef organisms. Global tion mediates zeaxanthin formation and zeaxanthin- Change Biol2:52?-545 related fluorescence quenching in intact chloroplasts. Styring S, Jegerschold C (1994) Light-induced reactions Plant Physiol99:1354-1361 impairing electron transfer through photosystem 11. In: Olaizola M, Yamamoto HY (1994) Short-term response of the Baker NR, Bowyer JR (eds) Photolnhlbition of photosyn- diadinoxanthin cycle and fluorescence yield to hlgh irradl- thesis: from molecular mechanisms to the field. BIOS Sci- ance in Chaetoceros muelleri (Bacillariophyceae). J Phy- entific Publishers, Oxford, p 51-74 col 30:606-612 Takabayashi M, Hoegh-Guldberg 0 (1995) The ecological Ola~zolaM, Laroche J, Kolber Z, Falkowski PG (1994) Non- and physiological differences between two colour morphs photochemical fluorescence quenching and the diadino- of the coral Pocillopora damicornis Mar Biol 123-705-714 xanthin cycle in a marine diatom. Photosynth Res 41. Telfer A, Barber J (1994) Elucidating the molecular mecha- 357-370 nisms of photoinhibition by studying isolated photosystem 86 Mar Ecol Prog Ser 183: 73-86, 1999
I1 reactlon centres In Baker NR, Bowyer JR (eds) Photoin- reef coral a novel approach Plant Cell Environ 19 hibition of photosynthesls from molecular mechanisms to 291-299 the field BIOS Scient~f~cPubhshers Oxford, p 25-50 We~sVM (1993) Effect of dissolved lnorganlc carbon concen- Titlyanov EA (1991) Light adaptation and production charac- tration on the photosynthesls of the symbiotic sea ter~st~csof branches dlffenng by age and ~llummationof anemone Alptasia pulchella (Carlgren) role of carbonic the hermatyplc coral PoclUopora verrucosa Symbiosis 10 anhydrase J Exp Mar B101 Ecol 174 209-225 249-260 Wh~tneySM, Yellowlees D (1995) Prellm~naryinvestlgatlons Vassihev IR, Pras~l0, Wyman KD, Kolber 2,Hanson AK, into the structure and actlvlty of nbulose bisphosphate Prent~ceJE, Falkowsk~PG (1994) Iniubition of PSI1 photo- carboxylase from two photosynthetlc d~noflagellates chem~stryby PAR and UV radiation in natural phyto- J Phyc0131 138-146 plankton commun~ties Photosynth Res 42 51-64 Wyman KD, Dub~nsky2, Porter JW, Falkowski PG (1987) Walker D (1992) Tansley review No 36 Excited leaves New Light absorpt~onand utihzation among hermatypic corals Phytol 121 325-345 a study in Jama~ca,West Indies Mar B101 96 283-292 Warner ME, Fitt WK, Schmdt GW (1996) The effects of ele- Young AJ, Frank HA (1996) Energy transfer react~onslnvolv- vated temperature on the photosynthetlc efflc~ency of Ing caroteno~ds-quenching of chlorophyll fluorescence zooxanthellae In hospite from four d~fferent species of J Photochem Photobiol B101 36 3-15
Editorial responsibility: George Humphrey (Contributing Submitted: May 22, 1998; Accepted: February 26, 1999 Ed~tor),Sydney, Australia Proofs received from authorjs): June 24, 1999