Mediterranean Marine Science

Vol. 11, 2010

The Effects of UV Exposure on the Antioxidant Enzyme Systems of Anemones

CAPARKAYA D. Dokuz Eylul University, Faculty of Science, Department of Chemistry, Tinaztepe Campus, Izmir CENGIZ S. Mehmet Akif Ersoy University, Faculty of Arts and Sciences, Department of Chemistry, Burdur DINCEL B. Dokuz Eylul University, Faculty of Science, Department of Chemistry, Tinaztepe Campus, Izmir DEMIR S. Dokuz Eylul University, Faculty of Science, Department of Chemistry, Tinaztepe Campus, Izmir CAVAS L. Dokuz Eylül University, Faculty of Science, Department of Chemistry, Biochemistry Division, 35160, Kaynaklar Campus, İzmir http://dx.doi.org/10.12681/mms.76

To cite this article:

CAPARKAYA, D., CENGIZ, S., DINCEL, B., DEMIR, S., & CAVAS, L. (2010). The Effects of UV Exposure on the Antioxidant Enzyme Systems of Anemones. Mediterranean Marine Science, 11(2), 259-276. doi:http://dx.doi.org/10.12681/mms.76

http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 02/08/2019 16:05:00 | Mediterranean Marine Science Research Article Indexed in WoS (Web of Science, ISI Thomson) The journal is available on line at http://www.medit-mar-sc.net

The Effects of UV Exposure on the Antioxidant Enzyme Systems of Anemones

D. CAPARKAYA, S. CENGIZ, B. DINCEL, S. DEMIR and L. CAVAS

Dokuz Eylul University, Faculty of Science, Department of Chemistry, Tinaztepe Campus, Izmir-Turkey

Corresponding author: [email protected]

Received: 23 October 2009; Accepted: 8 April 2010; Published on line: 22 October 2010

Abstract

Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) which are housekeeping enzymes protect cells from the harmful side effects of reactive oxygen species (ROS). Anemonia sulcata var. smaragdina and Anemonia rustica are widely distributed along the Turkish coastlines of the Aegean Sea. Recent studies have shown that environmental stresses such as elevated temperature, ultraviolet light, pathogen infection and decreased salinity might cause well- known bleaching effects in Anemonia species. The effect of UV-light on antioxidant enzyme activities such as SOD, CAT, GSH-Px, protein levels and secondary pigments were determined in A. sulcata var. smaragdina and A. rustica. SOD, CAT, GSH-Px activities, protein levels and secondary pigments of these morphotypes were observed in both tentacles and columns separately. According to studies on bleaching, elevated UV radiation may cause the bleaching event in anemones as a stress factor. However, in the present study no bleaching event was observed in anemone samples even after they were subjected to 5 hours of UV-exposure intervals. Moreover, UV exposure did not change antioxidant systems remarkably. However, more investigations are still needed to obtain the complete picture of the effects of UV-light on the cellular pathways of cnidarian–algal symbiosis.

Keywords: Anemonia sulcata; Anemonia rustica; Antioxidant enzymes; Bleaching; Lipid peroxidation.

Introduction nia viridis have been previously named as Anemonia sulcata var. smaragdina and Anemo- The sea anemone Anemonia viridis nia rustica, but these names are accepted (Forsska l) (Cnidaria, Anthozoa) is a com- as synonyms of A. viridis nowadays. The colour mon species for the Mediterranean Sea and differences among these morphotypes are it is abundant along these shores. It is char- caused by the green fluorescent protein (GFP)- acterised by a short column and long green like proteins and pink pigments in Anemo- tentacles (PICTON and MORROW, 2007). nia sulcata var. smaragdina (WIEDENMANN Two well-known morphotypes of Anemo- et al., 2000; LUKYANOV et al., 2000; SHAGIN

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http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 02/08/2019 16:05:00 | et al., 2004; WIEDENMANN et al., 2004; formation is done by catalase (CAT). CAT OSWALD et al., 2007). Due to having these decomposes hydrogen peroxide to molec- proteins and pink pigments, green tentacles ular oxygen and water. Organic hydroper- with pink tips are specific to Anemonia sul- oxides are decomposed by glutathione per- cata var. smaragdina, while in Anemonia rus- oxidase (GSH-Px). However, the overpro- tica they are an ordinary brown colour duction of ROS results in deleterious ef- (GOSSE, 1860; LEUTENEGGER et al., fects such as lipid peroxidation (LPO) of 2007). Both morphotypes contain an alga polyunsaturated fatty acids (PUFA), DNA commonly known as zooxanthellae in a sym- strand breaks and protein oxidation biotic form. This association is considered (HALLIWELL and GUTTERIDGE, 1999). to be mutualistic as the anemone provides The role of ROS in the bleaching phe- a hosting place, a protection from herbivory nomenon of anemones has been investi- and inorganic nutrients required for pho- gated by few researchers so far (RICHIER tosynthesis, while the alga provides oxygen et al., 2006; LEUTENEGGER et al., 2007). and carbohydrate-based products to the The antioxidant enzyme activities, lipid per- anemone as a product of photosynthesis oxidation levels and also photosynthetic pig- (MUSCATINE et al., 1979; FALKOWSKI ments were determined dependent on UV et al., 1984; MULLER-PARKER et al., 1990; exposure time in the present study. FURLA et al., 1998; GROVER et al., 2002; SABOURAULT et al., 2009). Environmental Materials and Methods stresses such as strong light, UV radiation, elevated sea temperature and pollution cause Sample collection and preparation well-known bleaching effects in Anemonia The specimens were collected from shal- species. Bleaching effects result in whiten- low waters (0.2- 0.5 m) of Gümüldür coast- ing of cnidarian symbiotic tissues and in- line. The coordinates were 38Æ 02' 39.79" N duce symbiont photoinhibition. RICHIER and 27Æ 00' 51.93" E. The habitats of sea et al. studied the effects of temperature and anemones were rocky and no microclimatic increased UV radiation in the symbiotic habitats were observed in the collection site cnidarian Anthopleura elegantissima tran- as mentioned by LEUTENEGGER et al. scriptome. The results of this study cor- (2007). The samples were transferred to lab- roborate that the stress response results oratory in natural seawater in plastic bags. in overproduction of the ROS in the aero- The tentacles from three individuals were bic metabolism (RICHIER et al., 2006; immediately cut at the base. Both tentacles RICHIER et al., 2008). and column samples were stored at -80 ÆC The ROS are overcome by antioxidants until used. These samples were labelled as under normal conditions. Enzyme-based the control group. UV-exposed experimen- antioxidants play important roles in scav- tal groups were created on the basis of dif- enging ROS (HALLIWELL & GUTTE- ferent exposure time and each of them con- RIDGE, 1999). Superoxide dismutase (SOD) tained three individuals. These groups were catalyses the dismutation reactions of the exposed to UV-Lamb (Slyvania, G30W). The superoxide radical into hydrogen peroxide. stress periods were set as 5 hours and sam- The formed hydrogen peroxide is also a dan- ples were collected at hourly intervals. 0.10 gerous molecule for the cell and must be gram samples were put into eppendorf tubes converted to a harmless molecule. This trans- which contained 1 mL of phosphate buffer

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http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 02/08/2019 16:05:00 | (pH 7.2, 50 mM). The samples were ho- H2O2 (AEBI, 1985). 1 IU/mL of CAT ac- mogenized with Ultra-Turrax T8 IKA-Werke tivity was expressed as decomposition of 1 homogenizator in 15-seconds intervals, for mL of hydrogen peroxide (10 mM) in one 30 seconds as homogenization time. The ho- minute. The absorbance at 240 nm was meas- mogenates were centrifuged in a refriger- ured in a UV-VIS 1601 Schimadzu Spec- ated centrifuge (Hettich 32R) at 10,000 rpm trophotometer at 25ÆC. for 10 min at +4ÆC to remove the cell de- bris. Then supernatants were used for meas- Glutathione peroxidase (GSH-Px) activity uring protein content and enzyme activities. determination GSH-Px activity was assayed by using a Protein Assay commercial kit produced by RANDOX (RS Total protein concentrations in super- 505). GSH-Px catalyses the oxidation of glu- natants were determined through the method tathione (GSH) by cumene hydroperoxide. reported by BRADFORD (1976). Bovine In the presence of glutathione peroxidase serum albumine (BSA) was used as the stan- and NADPH the oxidized glutathione dard for calibration curve. (GSSG) is immediately converted to the re- duced form by glutathione reductase (GR) Superoxide dismutase (SOD) activity deter- with a concomitant oxidation of NADPH mination to NADP+. The decrease in absorbance SOD activity was assayed by using a com- at 340 nm is measured. mercial kit produced by RANDOX (SD 125). The assay principle is based on the inhibition Lipid peroxidation (LPO) level determina- of occurrence formazan dye formed in the tion reaction of a superoxide radical. Superoxide The method described by YAGI (1994) radical anions are formed by the xanthine- was used to estimate the lipid peroxidation xanthine oxidase system. The formed super- level in the samples. It was expressed as oxide radicals attack INT (2-(4-iodophenyl)- nmol MDA/g wet weight. 3-(4-nitrophenol)-5-phenyltetrazolium) to According to this method the samples give a formazan dye. As SOD is an inhibitor are homogenized in KCl to aggregate the pro- of this reaction, the SOD enzyme activity is teins. The membrane lipids are dissolved in expressed via its inhibition rates. One unit of SDS (sodium dodecyl sulfate) and a peroxi- SOD is that which causes a 50% inhibiton of dation reaction achieved by acetic acid. The the rate of reduction of INT under the con- coloured complex which was measured at 532 ditions of the assay. The absorbance of oc- nm was formed with thiobarbituric acid. curred formazan dye was measured at 505 nm against air in UV-VIS 1601 Schimadzu Chlorophyll a-b and total carotenoids deter- Spectrophotometer at 37Æ C. minations Chlorophyll a-b and total carotenoids Catalase (CAT) activity determination levels were determined via the method pub- CAT activity was determined by the lished by LICHTENTALER and WELL- AEBI method (1985, which measures the BURN (1985) and DERE et al. (1998), with decrease in absorbance of H2O2 at 240 nm. slight modifications. A 0.1 g sample was ho- The reaction mixture (1mL) contained 50 mogenized by using an Ultra-Turrax T8 mM phosphate buffer, pH 7.0 and 10 mM IKA-Werke homogenisator. After addition

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http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 02/08/2019 16:05:00 | of 100% acetone into the samples, these the control groups (in tentacles) show a sta- samples were vortexed vigorously for one tistical difference in the protein levels minute. The homogenates were centrifuged (p<0.05). Similar results are also observed for 5 min at 2500 rpm, 20ÆC in a refriger- in control groups and 1 - 2 hour UV-exposed ated centrifugator (Universal 32R, Hettich experimental groups for the protein levels Zentrifuggen). The spectrum scanning was of columns (Fig. 1, a and b) (p<0.05). done between 350-800 nm in UV-VIS 1601 Figure 2a and b show the SOD activi- Schimadzu Spectrophotometer. The max- ties in tentacles of A.sulcata var. smaragdi- imum absorbances were recorded at 470, na and A. rustica. According to these fig- 645 and 662 nm which were in association ures, there is a statistical difference between with the total carotene, chlorophyll b and the SOD activities of A. sulcata var. smarag- chlorophyll a, respectively. The chlorophyll dina and A. rustica which was exposed UV a, b and total carotenoids levels were esti- light for 1 hour (p<0.05). It was interesting mated by using the following formulas: to note that there were no statistical dif- Chlorophyll a = 11.75 A662 - 2.350 A645 ferences among SOD activities in the ten- Chlorophyll b = 18.61 A645 - 3.960 A662 tacles of the control and experimental groups. Total Carotenoids = ((1000 A470 - 2.270 SOD activities in the columns of A. sul- Chlorophyll a - 81.4 Chlorophyll b)) / 227 cata var. smaragdina and A. rustica are shown in Figure 2c and 2d. It can be said that from Statistical Analysis the results of these figures, the SOD values Comparisons were statistically analyzed in the columns are significantly different by ANOVA followed by the Tukey test. The from each other, although they show a sim- statistical significance was set at 0.05. All ilar trend. numerical data were given as mean ± S.E.M. The results related to CAT activities in The values were the means of three sepa- tentacles and columns are presented in Fig- rate experiments. The correlation test was ure 3. Volumetric CAT activities in tenta- applied for possible relationships between cles have noticeable differences for 1, 3 and species. 5 hour UV exposed groups between two morphotypes (p<0.05). A sharp decrease Results occurs by the 3rd hour for A. sulcata var. smaragdina and as a result of this, signifi- The protein concentration of both ten- cant differences between control group and tacles and columns are presented in Figure 2, 3 and 5 hour UV-exposed groups are ob- 1. A remarkable difference in the protein served (p<0.05). levels of tentacles of A.sulcata var. smarag- According to Figure 3b which shows the dina is observed between the control group specific CAT activities of tentacles, the two and the experimental groups (p<0.05). A morphotypes show the same tendency but similar difference is observed only for the A. sulcata var. smaragdina exhibits a sharp 2nd hour group for the column of A. rusti- change in the 3rd hour, creating a significant ca (p<0.05). According to the results of a difference between the two species for the statistical test which was conducted to point 3rd and 5th hours (p<0.05). out the differences between two morpho- According to Figure 3c which demon- types that were exposed to UV light with strates volumetric CAT activity in columns the same duration, it can be said that only of A. sulcata var. smaragdina and A. rusti-

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Fig. 1: Protein concentration of tentacles (a) and columns (b) of UV-exposed A.sulcata var. smaragdina and A.rustica. Error bars show ±S.E.M. The results are the means of three different experiments. All the groups are compared with control groups and experimental groups exposed at the same time. Asterisks (*) on the error bars show the statistical difference compared to control groups in A.rustica and A.sulcata (p<0.05). The letter (a) shows the statistical difference between the morphotypes which were exposed to UV light for the same length of time (p<0.05).

ca, the values observed for the 2, 3 and 5 valid for A. rustica except for 2,3 and 5 hour hour UV-exposed groups show statistical UV-exposed groups (p<0.05). Control differences between two morphotypes groups of A .sulcata var. smaragdina and A. (p<0.05). Considerable differences are ob- rustica show statistical differences for the served at 1, 2 and 3 UV-exposed groups of CAT activities of columns (Fig. 3d) (p<0.05). A. sulcata var. smaragdina from the con- The results for GSH-Px are depicted trol samples. A similar explanation is also in Figure 4. No significant change in the vol-

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Fig. 2: Superoxide dismutase (SOD) activities of UV-exposed A.sulcata var. smaragdina and A.rustica. (a) shows volumetric activities in tentacles and (b) shows specific activities in tentacles. (c) shows volumetric activities in columns and (d) shows specific activities in columns. Error bars show ±S.E.M. The results are the means of two different experiments. All groups are compared with control groups and simultaneous experimental groups. Asterisks (*) on the error bars show the statistical difference compared to control group in A.rustica (p<0.05). The letters (a) show the statistical difference between experimental groups exposed at the same time (p<0.05). umetric GSH-Px activities of tentacles is ob- Significant differences at the 1st and 3rd hours served among the control and experimental for A. sulcata var. smaragdina and at the 2nd groups (p>0.05). On the other hand, a sig- hour for A. rustica are determined by com- nificant difference between 5 hour UV-ex- paring both of these morphotypes’ experi- posed A. sulcata var. smaragdina and A. rus- mental groups to control groups (Fig. 4d). tica is determined (p<0.05). No significant LPO levels of these morphotypes correlation is found between the volumetric versus UV exposure time are plotted in Fig- activity values of the two morphotypes (r=0.11, ure 5. According to Figure 5a, there is a p>0.05) (Fig. 4a). As can be seen in Figure statistical difference between 1 hour UV- 4b; the two morphotypes show a similar trend exposed morphotypes, as they show a great up to the third hour and significant differ- discrepancy in their LPO levels. The LPO ences are observed at the 5th hours for A. sul- level at 1 hour UV-exposed A. sulcata var. cata var. smaragdina and A. rustica (p<0.05). smaragdina is higher than that of the con- In the columns, there is no difference trol group (p<0.05). LPO levels in columns in volumetric GSH-Px activity between A. of A. sulcata var. smaragdina increase sharply sulcata var. smaragdina and A. rustica (Fig- in one hour, then decrease with the increasing ure 4c). In the light of specific GSH-Px ac- UV-exposure time. On the other hand, no tivity results, it can be noted that A. rustica significant change is observed for the LPO shows a peak at the 2nd hour of UV-expo- levels of the columns of A. rustica (Fig. 5b). sure. There is a statistical difference between Photosynthetic pigments such as chloro- the control groups of these morphotypes. phyll a, b and total carotenoids are also in-

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Fig. 3: CAT activities of UV-exposed A.sulcata var. smaragdina and A.rustica. (a) shows volumetric activity in tentacles and (b) shows specific activities in tentacles. (c) shows volumetric activity in columns and (d) shows specific activities in columns. Error bars show ±S.E.M. The results are the means of three different experiments. All groups are compared with control groups and experimental groups exposed at the same time. Asterisks (*) on the error bars show the statistical difference compared to control group A.sulcata and A.rustica (p<0.05). The letters (a) show the statistical difference between same time experimental groups (p<0.05). vestigated in the present study. According tion of ROS in the cell (LEUTENEGGER to Figure 6a which is related to chlorophyll et al., 2007; LESSER and SHICK, 1989; a level, these species show the same ten- DOWNS et al., 2002; TCHERNOV et al., dency with changing UV exposure but the 2004; SMITH et al., 2005). The cnidarians, level for control groups of A. sulcata var. like other aerobic organisms possess com- smaragdina is a bit higher than that of A. prehensive enzymatic and non-enzymatic rustica. According to Figure 6b there is no antioxidant defences to cope with overpro- significant difference between the chloro- duced ROS (RICHIER et al., 2008). In the phyll b levels of these morphotypes. Total present study, since no remarkable bleach- carotenoids exhibited the same trends as ing events were witnessed for the period be- chlorophyll a (Fig. 7). tween September 2006 and September 2008, we wanted to analyse and compare an- Discussion tioxidant enzyme activities in two well-known morphotypes of A. viridis; A. rustica and SOD, CAT, GSH-Px, protein concen- A. sulcata var. smaragdina from the Aegean trations, LPO levels and photosynthetic pig- Sea along Turkish coastlines. The study al- ments were investigated in the body of A. so aimed to present the antioxidant responses sulcata var. smaragdina and A. rustica col- of two variations of A. viridis against UV ra- lected from Gümüldür region of Izmir diation. (Turkey) province. Research into antho- According to our results, there is a sig- zoans has shown that several endogenous nificant negative correlation between pro- and exogenous factors cause overproduc- tein concentrations and specific SOD ac-

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Fig. 4: Glutathione peroxidase (GSH-Px) activities of UV-exposed A.sulcata var. smaragdina and A.rustica. (a) shows volumetric activities in tentacles and (b) shows specific activities in tentacles. (c) shows volumetric activity in columns and (d) shows specific activities in columns. Error bars show ±S.E.M. The results are the means of two different experiments. All groups are compared with control groups and experimental groups exposed at the same time (p>0.05). Asterisks (*) on the error bars show the statistical difference compared to the control group A.sulcata and A.rustica (p<0.05). tivity in the tentacles of A. sulcata var. smarag- synthetic pigments in A.sulcata var. smaragdina (r=0.878, p<0.05). Similarly, correla- dina and also absence of these relationships tions were found among chlorophyll a and in A.rustica might be explained by the exis- SOD-CAT activities in the tentacles of A. tence of different regulatory mechanisms sulcata var. smaragdina (r=-0893, -0.925, in these morphotypes. Significant correla- p<0.05, respectively). It was very interest- tion (r=0.908, p<0.05) between SOD and ing to note that no correlation was found CAT activities in the column of A.sulcata for the above- mentioned relationships in var. smaragdina might show that these two A. rustica. The existence of the correlations enzymes work together in the columns. among SOD-CAT activities and photosyn- Moreover, a remarkable correlation (r=0.922, thetic pigments might be explained by the p<0.05) existed between CAT and GSH- relationship of these pigments and ROS. Px activities. These results may reveal that According to photosynthetic process in high- only one peroxidase is not enough in the er plants, secondary pigments play impor- decomposition of overproduced hydrogen tant roles in scavenging overproduced ROS peroxide by SOD activities. Therefore, two (BOYER, 2006). The bonds in the photo- peroxidases, CAT and GSH-Px, may play a synthetic pigments in anemones might have synergistic role in the sea anemones. Ob- been protected by other antioxidant mol- servation of increased specific GSH-Px ac- ecules such as vitamins and glutathione etc, tivities in the tentacles of both A.sulcata var. inasmuch as no remarkable differences were smaragdina and A.rustica can be explained observed in the concentrations of photo- by possible overproduced lipid hydroper- synthetic pigments. The existence of rela- oxides. In contrast to the sea anemone tionships related to enzymes and photo- population in Gümüldür, we have recent-

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Fig. 5: (a) Lipid peroxidation levels in tentacles of A.sulcata var. smaragdina and A.rustica. (b) Lipid peroxidation levels in columns of A.sulcata var. smaragdina and A.rustica. Error bars show ±S.E.M. The results are the means of three different experiments. Asterisks (*) on the error bars show the statistical difference compared to the control group in A.sulcata (p>0.05). The letter (a) shows the statistical difference between same time experimental groups (p<0.05).

ly observed bleached anemones which live sea anemones in the literature. Early at- in shallow waters in Dikili. According to tempts (DYKENS, 1984; DYKENS and preliminary results, this bleaching event can SHICK, 1984; SHICK et al., 1995; TYTLER be associated with the higher sea water tem- and TRENCH, 1988) at the determination perature. There are many scientific reports of antioxidant enzyme activities in sea cnidar- on the antioxidant enzyme activities of ians were carried out by using cell-free ex-

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Fig. 6: (a) Chlorophyll a amount in tentacles of A.sulcata var. smaragdina and A.rustica. (b) Chloro- phyll b amount in tentacles of A.sulcata var. smaragdina and A.rustica. Error bars show ±S.E.M. The results are the means of three different experiments.

tracts as applied in the present study. Among gen level. They found 590% and 100% in- the early attempts at antioxidant enzyme creases in the SOD and CAT activities, re- activities in sea cnidarians, DYKENS and spectively, of shade-adapted aposymbiotic SHICK (1984) investigated the antioxidant Anthopleura elegantissima (DYKENS et al., enzyme activities, SOD and CAT, in An- 1992). DYKENS et al. (1992) also under- thopleura elegantissima which was subject- lined that SOD and CAT enzyme activities ed to sunlight to generate hyperbaric oxy- in Anthopleura elegantissima are associated

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http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 02/08/2019 16:05:00 | Fig. 7: Total carotenoid amount in tentacles of A.sulcata var. smaragdina and A.rustica. Error bars show ±S.E.M. The results are the means of three different experiments. with the chlorophyll content of endosym- ROS is got rid of by sea anemones in the bionts where the SOD and CAT are lo- Aegean Sea by their antioxidant system. On calised. HAWKRIDGE et al. (2000) stud- the other hand, inasmuch as the overpro- ied the localisation of antioxidant enzymes duction of ROS in aerobic organisms is de- in the sea anemone Anemonia viridis and pendent on not only endogenous factors but the coral Goniopora stokesi. HAWKRIDGE also exogenous factors such as pollution, et al. (2000) showed that SOD localized on pathogenity and abiotic factors etc., regu- the ruptured threads and shafts of b- lar measurement of the antioxidant status mastigophore in Anemonia viridis. Existence of sea anemones is strongly warranted for of SOD and CAT in the symbiotic algae was the population health of sea anemones to also shown by researchers. However, the estimate a possible bleaching event. gold-labelling method that they used has some limitations because mammalian anti- Conclusion bodies were used for sea anemones. In a very recent study, RICHIER et al. (2006) The present study demonstrates the an- investigated the effects of thermal stress on tioxidant enzyme activities in two well-known the symbiosis breakdown in the sea anemone, sea anemones, Anemonia rustica and Anemo- Anemonia viridis. They found a link between nia sulcata var. smaragdina from the Aegean bleaching and apoptotic process in the sea Sea along Turkish coastlines. These enzyme anemone. Due to the comparable values activities may be considered as indicators among antioxidant enzyme activities in the of the health of the cnidaria-related ecosys- present study and also the absence of any tem. The antioxidant enzyme systems in sea remarkable bleaching events reported so anemones can also be defined as ROS scav- far, it can be said that the production of enging systems. The increases in these en-

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http://epublishing.ekt.gr | e-Publisher: EKT | Downloaded at 02/08/2019 16:05:00 | zyme activities show the overproduced ROS C.M., 2002. Oxidative stress and seasonal level in the metabolism. Since these increases coral bleaching. Free Radical Biology & are generally caused as a result of environ- Medicine, 33 (4): 533-543. mental stresses such as UV radiation, DYKENS, J.A. & SHICK, J.M., 1984. Pho- pathogens, or elevated temperature, they tobiology of the symbiotic sea anemone, can be taken as indicators of the healthy as- Anthopleura elgantissima: defenses against sociation between anemones and zooxan- photodynamic effects and seasonal pho- thella. In the present study no bleaching to acclimatization. Biological Bulletin, event was observed in anemone samples 167: 683-687. even when they were given 5 hours of UV- DYKENS, J.A., 1984. Enzymic defenses exposure. Moreover, UV exposure did not against oxygen toxicity in marine cnidar- change antioxidant systems remarkably. ians containing endosymbiotic algae. However, more investigations are still need- Marine Biology Letters, 5: 291-301. ed in order to obtain a complete picture of DYKENS, J.A., SHICK, J.M., BENOIT, C., the effects of UV-light on the cellular path- BUETTNER, G.R. & WINSTON, G.W., ways of cnidarian–algal symbiosis. 1992. Oxygen Radical Production in the Sea Anemone Anthopleura elegantissi- Acknowledgement ma and its Endosymbiotic Algae. The Jour- nal of Experimental Biology, 168: 219-241. The authors thank ProjectAWARE for FALKOWSKI, P.J., DUBINSKY, Z., the partial financial support of the project MUSCATINE, L. & PORTER, J.W., (Grant Application number 639). 1984. Light and the bioenergetics of a symbiotic coral. Bioscience, 34: 705-709. References FURLA, P., BENAZET-TAMBUTTE, S., JAUBERT, J. & ALLEMAND, D., 1998. AEBI, H., 1985. Catalase. p. 121-126. In: Functional polarity of the tentacle of the Methods in Enzymology. Packer, L. (Ed), sea anemone Anemonia viridis: role in Academic Press, San Diego. inorganic carbon acquisition. American BRADFORD, M.M., 1976. A rapid and sen- Journal of Physiology, 274: 303-310. sitive method for the quantization of mi- GOSSE P.H., 1860. Actinologia Britanni- crogram quantities of protein utilizing ca: a history of the British sea-anemones the principle of protein-dye binding. An- and corals. Van Voorst, Paternoster Row alytical Biochemistry, 72: 248-254. London. BOYER, R., 2006. Concepts in Biochemistry, GROVER, R., MAGUER, J.F., REY- 3rd edition, John Wiley & Sons, Inc. Unit- NAUD-VAGANAY, S. & FERRIER- ed States of America. PAG S, C., 2002. Uptake of ammo- DERE, S., GUNES, T. & SIVACI, R., 1998. nium by the scleractinian coral Stylophora Spectrophotometric determination of pistillata: Effect of feeding, light, and Chlorophyll-A, B and total carotenoid ammonium concentrations. Limnology contents of some algae species using dif- & Oceanography, 47: 791-802. ferent solvents. Turkish Journal of Botany, HALLIWELL, B. & GUTTERIDGE, J.M.C, 22: 13-17. 1999. Free Radicals in Biology and DOWNS, C.A., FAUTH, J.E., HALAS, J.C., Medicine, 3rd ed. Oxford University DUSTAN, P., BEMISS, J. & WOODLEY, Press, New York, 936 pp.

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