Mechanisms on inhibition of photosynthesis in Kandelia obovata due to extreme cold events under climate change
© The Author(s). 2016
Received: 24 August 2016
Accepted: 7 November 2016
Published: 25 November 2016
The Erratum to this article has been published in Ecological Processes 2017 6:3
Mangroves that grow at the latitudinal extremes of their distribution are susceptible to extreme cold events. Successive enhancement of low temperature stress (seLTS) is a typical characteristic of extreme cold events. Low temperature stress can inhibit mangrove photosynthesis, which often inhibits the growth and development of mangroves. However, the possible reasons for impairment to photosynthesis of mangroves due to extreme cold events remain unclear.
Kandelia obovata seedlings in a growth chamber were exposed to 5°C/−2°C (day/night) for 36 h (−2°C for 16 h) with 12 h light per day at 600 μmol m−2 s−1 photosynthetic photon flux density (PPFD) (a low temperature stress, aLTS), then the plants were transferred to the control condition (15°C/10°C (day/night) and allowed to recover for 5 days (R1). The other seedlings were subjected to low temperature treatment with a day/night temperature of 5°C/−1°C in a growth chamber for 24 h. Then these plants were transferred to 5°C/−2°C (day/night) under the same light and climate conditions for 36 h (two low temperature stresses, tLTS). Following the successive enhancement of low temperature treatment, these plants were returned to 15°C/10°C (day/night) for another 5-day recovery period (R2).
Results showed that aLTS treatment significantly reduced leaf net photosynthetic rate (P n) and stomatal conductance (G s), while increased intercellular CO2 concentration (C i). Photosynthetic activity of leaves quickly recovered after the plants were returned to control temperature for 5 days (R1). However, decreases in leaf P n were more obvious under tLTS treatment than those under aLTS treatment. This reduced P n might be attributed to stomatal and non-stomatal limitations. Moreover, non-stomatal limitation played a major role in reducing P n during tLTS treatment, as proven by reduced ribulose 1,5-biphosphate carboxylase (RuBPCase) activity. This limitation also enhanced lipid peroxidation in chloroplasts, decreased sucrose-metabolizing enzymes and ratios of both chlorophyll a/chlorophyll b (Chl a/b) and chlorophyll/carotenoids (Chl/Car), and increased protease senescence. Damages of tLTS treatment to photosynthesis were insufficiently alleviated even after the plants were returned to control temperature for 5 days (R2).
Successive enhancement of low temperature depressed successful recovery of leaf photosynthesis of K. obovata seedlings mainly by impairing pigment synthesis, antioxidant metabolism, and sucrose translocation, as well as accelerating senescence of endopeptidase. Furthermore, our results suggest that decreases in sucrose content in leaves might also account for increases in reactive oxygen species (ROS) in chloroplasts.
Low temperature is a major environmental constraint for plants as a result of the injury provoked on leaf photosynthesis. Chloroplasts are the site of photosynthesis in cells and are a rich source of reactive oxygen species (ROS). ROS production is dramatically enhanced in response to adverse environmental conditions, often resulting in an excessive demand of the cellular antioxidant machinery (Fahnenstich et al. 2008). In general, to limit the action of ROS, plant cells evolve anti-oxidative systems including superoxide dismutase (SOD, EC 22.214.171.124), ascorbate peroxidase (APX, EC 126.96.36.199), and peroxidase (POD, EC 188.8.131.52). Inhibition of antioxidant enzyme activities by chilling stress leads to production of ROS, which confers serious oxidative damages to Zingiber officinale (Li et al. 2014). Moreover, a significant rise in H2O2 content in the leaves of cucumber, as well as higher activities of APX and SOD in the leaves, was found under low temperature stress (Lee et al. 2004). When tomato leaves are exposed to low temperature stress, sucrose contents are determined by photosynthetic rate, and sucrose phosphate synthase (SPS) activity is determined by sugar concentration (Artuso et al. 2000). That is, variations in photosynthesis are related to stomatal closure, leaf chlorosis, chloroplast malfunction, photo-inhibition or photo-oxidation, and sucrose metabolism. However, it is unclear whether physiological metabolic processes that depress the photosynthesis of mangroves have unique characteristics, in comparison with terrestrial plants.
The latitudinal distribution of mangrove species is limited mainly by temperature-related drivers (Quisthoudt et al. 2012). Mangroves are mainly tropical tree species that grow in the intertidal zone of tropical and subtropical coasts. According to the tropical origin, climate warming may enable mangroves to extend their distribution to higher latitudes (Gilman et al. 2008; Naidoo 2016). However, mangroves that lie along the latitudinal extremes of their distribution are still vulnerable to the extreme cold events (Ellis et al. 2006; Gilman et al. 2008, Saintilan et al. 2014). In general, successive enhancement of low temperature is a typical characteristic of extreme cold events. Several researches reported that low temperature, especially freezing stress, seriously influenced mangrove development and survival (Ellis et al. 2006; Stuart et al. 2006; Liu et al. 2014). For example, a severe winter in Florida, occurred between the 1995 and 1996 growing seasons (minimum air temperature of −8°C in January 1996), killed 12% mangrove trees, and the severe freeze season occurred once approximately 8 years (Stevens et al. 2006; Ross et al. 2009). Studies have shown that frost may restrict nutrient resorption during leaf senescence. K. obovata resorbed 61% N and 42% P during normal leaf senescence (Wang et al. 2011). Peng et al. (2013) found that three CBF/DREB1 (C-repeat binding factor/dehydration responsive element-binding factor 1) genes play different roles in Avicennia marina, and the AmCBF2 might be involved in the signaling pathway of cold stress response. But little attention has been paid to physiological mechanisms on these damages to mangroves by seLTS.
K. obovata Sheue et al. (2003) is a common mangrove species along southeastern coastlines of China. This species has high tolerance against low temperature stress. However, low temperature still decreases the total number of leaves that emerge from K. obovata yearly (Analuddin et al. 2009). In 2010, a blizzard severely damaged or killed many mangrove trees in southern China. It was found that seLTS severely damaged the development of K. obovata by disturbing photosynthetic capacity in the Yueqing bay, Zhejiang Province, China (Zheng et al. 2013). Unfortunately, the possible reasons for impairment to photosynthesis of mangroves by seLTS remain unclear. In the present study, we investigated the effects of seLTS and subsequent recovery on photosynthetic characteristics, photosynthetic pigment contents, chloroplast anti-oxidation system, sucrose synthesis, and proteolysis in the leaves of K. obovata seedlings to assess harmful effects on photosynthesis and find reasons for reduction in photosynthesis. These efforts will be useful in restoring and protecting mangrove forests especially in high latitude area.
Plant materials and culture
Propagules of K. obovata were collected from Fuding (the northern limit of natural mangrove distribution in China, 27°17′N), Fujian Province, China. Undamaged propagules with 12 ± 0.5 cm length were randomly planted in plastic pots (30 cm in height and 20 cm in diameter) filled with 3.0 kg of intertidal soil and maintained in a temperature-controlled growth chamber under 25°C/20°C day/night cycles. The everyday light and climate conditions in this chamber were 12 h of light at 600 μmol m−2 s−1 photosynthetic photon flux density (PPFD) and 70% relative humidity. The pots were immersed in nutrient solution (Haines and Dunn 1976), which was replenished once a week.
The experiment was conducted in Wenzhou (28°1′N), Zhejiang Province, China. Wenzhou is the northern limit of mangroves introduced in China. In Wenzhou, extreme low temperatures sporadically occur, causing substantial frost damage to mangroves. In January, 2011, low temperature in mangrove communities steadily decreased, and the minimal air temperature dropped from −1.1°C (about 10 h in 24 h) to −3.2°C (about 4 h in 72 h). The lowest temperature occurred during flood tide and the minimal surface seawater temperature during this period was about 5.5°C. The seedlings of K. obovata were therefore protected by warmer seawater. But seedlings were exposed to freezing temperature (−2.0°C) for about 16 h in 72 h. That is, there was a rare cold event in 2011, resulting in badly damaged mangroves (Zheng et al. 2016). To simulate this cold event, an experiment was performed as follows: The seedlings were maintained at 15°C/10°C day/night (near minimum monthly mean temperature in Wenzhou) with 12 h light per day at 600 μmol m−2 s−1 PPFD for 15 days. Then, they were divided into three batches after the tenth pair of leaves expanded. One batch was in a chamber at a day/night temperature of 15°C/10°C (control plants). Another batch in a growth chamber was exposed to −2°C from 8:00 pm to 8:00 am in the next morning in the dark and then the temperature was shifted to 5°C from −2°C for 12 h with light at 600 μmol m−2 s−1 PPFD. Afterwards, the seedlings continuously grew at −2°C for 4 h in the dark and at 5°C for 8 h with light at 600 μmol m−2 s−1 PPFD. This treatment was designated as a low temperature stress (aLTS). After aLTS treatment, the plants were transferred to the control condition and allowed to recover for 5 days (R1). The last batch was subjected to low temperature treatment with a night temperature of −1°C in a growth chamber from 8:00 pm to 8:00 am in the next morning in the dark, then the day temperature was 5°C with light at 600 μmol m−2 s−1 PPFD by setting the control from 8:00 am to 8:00 pm. Then, these plants were allowed to continue processing according to the methods of aLTS treatment (designated as two low temperature stresses (tLTS)). Following the successive enhancement of low temperature treatment, these plants were returned to 15°C/10°C (day/night) for another 5-day recovery period (R2). In total, three treatments were established. Physiological parameters were determined by using the second pair of fully expanded leaves. The experiment was replicated in five separate plants for each treatment.
Gas exchange assay
Net photosynthetic rate (P n), stomatal conductance (G s), and intercellular CO2 concentration (C i) were measured on the youngest fully expanded leaves using a Li-6400 portable photosynthesis system (Li-Cor, Lincoln, NE, USA). A leaf chamber equipped with a red/blue LED light source was used. All measurements were taken at a constant flow rate of 500 mL min−1 and CO2 concentration of 390 μmol mol−1 under a photosynthetically active radiation of 600 μmol m−2 s−1.
Leaf chlorophyll content assay
The youngest fully expanded leaf sample (0.1 g) was sliced and incubated in 50 mL of pigment extraction solution containing acetone and anhydrous ethanol (1:1, v/v) in the dark for 24 h at 25°C. Chlorophyll contents were determined as described by Arnon (1949).
Ribulose 1,5-biphosphate carboxylase activity assay
RuBPcase activity was determined according to the method of Sui et al. (2005) by spectrophotometry. In brief, the reaction mixture was comprised of 5 mM NADH, 50 mM ATP, 50 mM phosphocreatine, 0.2 mM NaHCO3, 100 mM Tris–HCl buffer (pH 7.8), 12 mM MgCl2, 0.4 mM EDTA–Na2, 160 U mL−1 3-phosphoglycerate kinase, glyceraldehyde-3-phosphate dehydrogenase, creatine phosphokinase, and 0.1 mL enzyme extract solution. The changes of A340 in 1 min of the assay mixture were measured with a UV-VIS recording spectrophotometer (UV-160A, Shimadzu, Japan).
Chloroplasts were isolated using the method described by Seigneurin-Berny et al. (2008). Fresh leaves were homogenized in ice-cold isolation buffer (0.4 mM sorbitol, 20 mM Tricine-KOH, 10 mM EDTA, 10 mM NaHCO3, and 0.15% bovine serum albumin (BSA)). The homogenate was filtered through four layers of gauze, and the filtrate was centrifuged at 1465×g for 90 s (including deceleration). The pellets were resuspended in 2 mL of buffer (0.4 mM sorbitol, 20 mM Hepes–KOH, 2.5 mM EDTA, 5 mM MgCl2, 10 mM NaHCO3, and 0.15% BSA), placed into a tube containing 8 mL of resuspension medium with 40–80% (v/v) Percoll and then centrifuged at 13300×g for 10 min. The chloroplasts were collected at the 40%/80% interface. All procedures were performed at 0–4°C.
Assays of malondialdehyde content, hydrogen peroxide release, superoxide anion release rate, and antioxidant enzyme activity in leaf chloroplasts
Malondialdehyde (MDA) content was measured in terms of the total content of thiobarbituric acid (TBA) reactive substances as described by Hodges et al. (1999). Hydrogen peroxide (H2O2) content was assayed using the method by Moloi and van der Westhuizen (2006). Release rate of O2 − was measured as previously described by Sui et al. (2007).
Superoxide dismutase (SOD) activity was assayed following the method from Giannopolitis and Ries (1977), while peroxidase (POD) activity was measured as described by Lee and Lin (1995). Ascorbate peroxidase (APX) activity and soluble protein (SP) content were determined according to Nakano and Asada (1981) and Bradford (1976), respectively.
Total soluble sugar, sucrose, and free amino acid assays
Approximately 0.1 g of dried leaf sample powder was extracted with 8 mL of 80% (v/v) ethanol. The extract was incubated at 80°C for 30 min and then centrifuged at 10000×g for 10 min. The supernatant solution was collected. The extraction procedure was repeated thrice to ensure the complete extraction of sugars and amino acid. The total soluble sugar content was measured as previously described by Zhang et al. (2011). The sucrose content was determined by a similar method from Hendrix (1993) with minor modifications. Ethanol extract (0.4 mL) was mixed with 0.2 mL of 2 M NaOH, heated in a boiling water bath for 5 min and then allowed to cool. The reactants were mixed with 2.8 mL of 30% HCl and 0.8 mL of 0.1% resorcin (0.1 g of resorcin in 100 mL of 95% aqueous ethanol). The mixture was incubated at 80°C for 10 min. The reaction mixture was then measured at 480 nm absorbance.
The content of free amino acids was measured by a similar method from Moore and Stein (1954) with minor modifications. Ninhydrin reagent (3 mL, pH 5.4), distilled water (1 mL), 0.1% ascorbic acid, and ethanol extract (1 mL) were mixed. The mixture was heated in a boiling water bath for 15 min and then allowed to cool in tap water. The content of free amino acids was spectrophotometrically determined at 570 nm.
Sucrose phosphate synthase and sucrose synthase assays
Enzyme extraction and assay were according to Hubbard et al. (1989). Approximately 0.5 g of fresh leaves were ground and homogenized in a prechilled mortar and pestle with 5 mL of extraction buffer [50 mM Mops–NaOH (pH 7.5), 5 mM MgCl2, 1 mM EDTA, 2.5 mM dithiothreitol (DTT), 0.05% (v/v) Triton X-100, and 0.5 mg mL−1 BSA]. The homogenate was centrifuged at 10,000×g for 30 min, and the supernatant was immediately used to analyze activities of sucrose synthase (SS) and sucrose phosphate synthase (SPS). All extraction procedures were carried out at 0–4°C.
The assay mixture contained 50 mM Mops–NaOH buffer (pH 7.5), 15 mM MgCl2, 5 mM fructose-6-phosphate, 15 mM glucose-6-phosphate, 5 mM UDP–glucose, and 0.1 mL of enzyme extract. The mixture was incubated at 25°C for 30 min, and the reaction was terminated by adding 0.1 mL of 2 M NaOH. The tubes were placed in boiling water bath for 10 min and then allowed to cool. The mixture was added with 1 mL of 0.14% (w/v) anthrone in 13.8 M H2SO4 and then incubated in 40°C water bath for 20 min. The tubes were allowed to cool, and sucrose content was spectrophotometrically determined at 620 nm absorbance. SS assay was the same as above, except that the reaction mixture contained 10 mM fructose instead of fructose-6-phosphate and was devoid of glucose-6-phosphate.
Endopeptidase and carboxypeptidase assays
Enzymes were extracted according to the procedure described by Mehta and Mattoo (1996). Fresh leaves (1 g) were homogenized in 5 mL of refrigerated grinding medium (50 mM Tris–HCl buffer, 4 mM DTT, 1 mM EDTA, and 1% PVP). The extract was centrifuged at 15000×g for 30 min, and the supernatant was collected to estimate activities of endopeptidase and carboxypeptidase.
Carboxypeptidase activity was assayed as described by Wang et al. (2003). The reaction mixture (0.4 mL of enzyme supernatant, 0.4 mL of 100 mM acetate buffer, and 0.2 mL of 0.05 M bovine hemoglobin) was incubated at 38°C for 1 h. Then, the reaction was terminated by adding 1 mL of 10% TCA. After centrifugation at 4000×g for 10 min, the suspension was used for ninhydrin reaction.
The reaction mixture for endopeptidase assay was similar to that for carboxypeptidase assay except that 0.3% (w/v) hippuryl-L-phenylalanine was used instead of 0.05 M bovine hemoglobin. Activities of endopeptidase and carboxypeptidase were expressed in μmol α-NH2 g−1 FW h−1.
Data of each parameter were normal and reported as mean + standard deviation (SD) of five replicates. The one-way ANOVA was used to test any significant differences among the treatments. Pearson’s correlation was used to analyze relationships among physiological indices. Significant level was at 0.05. All statistical analyses were performed by using the SPSS software (SPSS Ver. 19.0, Chicago, IL, USA).
Leaf gas exchange
Hydrogen peroxide and malondialdehyde contents, O2 − release rate, and anti-oxidative enzyme activities in chloroplasts
Contents of MDA and H2O2 and release of O2 − in chloroplasts of the K. obovata leaves were significantly increased under aLTS and tLTS treatments (Fig. 4a, b and f). Only the contents of MDA and H2O2 and the release rate of O2 − in the chloroplasts recovered within 5 days of recovery after aLTS treatment. Significant differences in the contents of MDA and H2O2 and release rate of O2 − were still observed in the chloroplasts between tLTS treatment and the control within 5 days of recovery after tLTS treatment.
Compared with the control, treatment aLTS significantly increased the activities of SOD and POD in the chloroplasts (Fig. 4c, d). Both activities recovered within 5 days recovery treatment. However, treatment tLTS significantly decreased these activities, compared with the control plants. Subsequent recovery only slightly improved the activities of SOD and POD in chloroplasts; thus, significant differences were still observed between the treatments. Compared with the control, aLTS and tLTS treatments significantly decreased activity of APX (Fig. 4e). Activity of APX was recovered within only 5 days of recovery after aLTS treatment.
TSS and sucrose contents and sucrose synthase and sucrose phosphate synthase activities in leaves
Soluble protein and free amino acid contents and endopeptidase and carboxypeptidase activities
Correlation coefficient among physiological indices
Correlation coefficients between net photosynthetic rate (Pn) and physiological indices in K. obovata
Stomatal conductance (G s)
Intercellular CO2 concentration (C i)
Chlorophyll a/chlorophyll b (Chl a/b) ratio
Chlorophyll/carotenoid (Chl/Car) ratio
Ribulose 1,5-biphosphate acrboxylase (RuBPcase)
Ascorbate peroxidase (APX)
Superoxide dismutase (SOD)
Superoxide anion (O2 −)
Hydrogen peroxide (H2O2)
Total soluble sugar (TSS)
Sucrose synthase (SS)
Sucrose phosphate synthase (SPS)
Soluble protein (SP)
Free amino acid (FAA)
P n positively correlated with ratios of Chl a/b and Chl/Car in leaves (Table 1). This suggested that decreases in ratios of Chl a/b and Chl/Car in leaves limit photosynthesis under low temperature stress. In the present study, decreases in ratio of Chl a/b in leaves under low temperature stress and subsequent recovery resulted from more increases in Chl b than in Chl a. One explanation is that, when Chl a preferential assembling reaction centers are limited, the reaction centers no longer incorporate the majority of Chl a. Thus chlorophyllide a oxygenase synthesizes Chl b in significant amounts (Wu et al. 2007). In addition, low ratios of Chl/Car in leaves under low temperature stress and subsequent recovery could be attributed to the faster decreases in Chl than in Car. These indicated that aberration of Chl synthase function decreases Chl a synthesis. Moreover, ratios of Chl a/b and Chl/Car in leaves under tLTS treatment were lower than those of aLTS treatment, indicating that seLTS more severely affected pigment components, in accordance with the findings from Zheng et al. (2013).
Reduction in P n under low temperature stress is associated with increase in ROS in the chloroplasts of cucumber, and low temperature increases activities of SOD and APX in chloroplasts (Hu et al. 2008). In the present study, although activities of APX and POD increased, decreases in SOD activity would still probably lead to the accumulation of H2O2 and O2 − in chloroplasts for the first low temperature injury. This is in accordance with the findings of Peng et al. (2015). And levels of H2O2, O2 −, and MDA in chloroplasts recovered after the first recovery period of 48 h under control conditions (Fig. 3a, b, and d). However, treatment tLTS aggravated damage extent and then depressed activities of SOD, POD, and APX and caused massive accumulation of H2O2, O2 −, and MDA in chloroplasts. This indicated that seLTS induced oxidative bursts because of ROS overproduction in chloroplasts, thereby reduced utilization of the absorbed light energy for CO2 assimilation. Correlation coefficients between P n and activities of APX, SOD, and POD, and levels of H2O2, O2 −, and MDA in chloroplasts agreed with this proposition. After the plants were subjected to two freezing injuries, levels of H2O2, O2 −, and MDA and activities of SOD, POD, and APX did not recover even when they were transferred to control conditions for 5 days.
Sucrose metabolism is crucial to carbohydrate translocation and accumulation, and low temperature stress might impair sucrose translocation (Artuso et al. 2000). Photosynthesis rate also determines sucrose content in sugarcane leaves exposed to low temperature stress (Du and Nose 2002). Therefore, positive relationship between P n and sucrose content indicated that decreases in P n restrained sucrose synthesis under low temperature. Sucrose synthesis and/or degradation referred to SPS and SS. Genetically, SPS catalyzes a step in sucrose synthesis. In addition, protein SS is preferentially expressed during cold stress, suggesting that the enzyme might be involved in plant responses to low temperature stress (Crespi et al. 1991). SS participates in sucrose cleavage or sucrose synthesis. In the present study, SS activity in leaves increased after aLTS treatment, but recovered after treatment R1. Meanwhile, treatment aLTS significantly increased contents of TSS and sucrose but decreased activity of SPS. These suggested that SS activity in leaves during aLTS treatment participated in sucrose synthesis and that sucrose accumulation induced by low temperature injury principally depended on increases in SS activity. Treatment tLTS significantly reduced content of sucrose and activities of SS and SPS but significantly increased content of TSS. These parameters did not recover even after 5 days recovery treatment, suggesting that sucrose metabolism in K. obovata leaves was deteriorated after seLTS treatment. It was also found that sucrose acted as a direct ROS scavenger (Van den Ende and Valluru 2009). Therefore, decreases in sucrose content in leaves might also account for increases in ROS in chloroplasts.
Decreases in SP and increases in free amino acids might be related with activities of endopeptidase and carboxpeptidase (Cohen and Demaggio 1986). Endopeptidase is a protease that catalyzes the cleavage of peptide bonds within a polypeptide or protein, and carboxypeptidase is another protease enzyme that hydrolyzes (cleaves) a peptide bond at the C-terminal end of a protein or peptide, which has a stronger preference for free amino acids. In the present study, aLTS and tLTS treatments improved the accumulation of SP and FAA and activity of carboxypeptidase but decreased activity of endopeptidase, indicating that carboxypeptidase was critical to cleave peptide bonds for FAA. In addition, the principal reason for protein degradation under low temperature stress is the accumulation of ROS that promotes oxidation of proteins and lipids and inhibits activity of endopeptidase (Prasad 1996). Protein degradation in K. obovata was completely recovered after R1, but proteolysis was insufficiently recovered after R2. Furthermore, FAA content positively correlated with carboxypeptidase activity but negatively correlated with endopeptidase activity, indicating that low temperature stress induced senescence of endopeptidase but improved efficiency of carboxypeptidase to synthesize FAA.
In conclusion, tLTS treatment decreased P n in the leaves of K. obovata seedlings because of stomatal closure and non-stomatal factors. Increase in C i might be partly explained by declined ratios of Chl a/b and Chl/Car, enhanced peroxidation in chloroplasts, weakened activities of sucrose-metabolizing enzymes, and accelerated senescence of endopeptidase. Therefore, photosynthetic capacity could not completely recover after tLTS treatment.
This study was funded by the projects of Special Funding for Research of National Oceanic Public Service Industry (201505028), Zhejiang Provincial Natural Science Foundation (LQ13C030002), Science and Technology Plan of Zhejiang Province (2012C12017-3, 2013C25096, and 2014F50003), China and Israel Cooperation Project (2015), and Wenzhou Municipal Science and Technology Plan Project (S20110007).
WL, YY, CZ, and JT designed the study and collected and analyzed the data. CZ, JC, JQ, and XP performed the experiment work and analyzed the data. CZ and JT drafted the manuscript. All authors read and approved the final manuscript.
The authors declare that they have no competing interests.
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- Analuddin KS, Sharma S, Suwa R, Hagihara A (2009) Crown foliage dynamics of mangrove Kandelia obovata in Manko wetland, Okinawa Island, Japan. J Oceanogr 65:121–127View ArticleGoogle Scholar
- Arnon D (1949) Copper enzymes in isolated chloroplasts. Polyphenoloxidase in Beta vulgaris. Plant Physiol 24:1–5View ArticleGoogle Scholar
- Artuso A, Guidi L, Soldatini GF, Pardossi A, Tognoni F (2000) The influence of chilling on photosynthesis and activities of some enzymes of sucrose metabolism in Lycopersicon esculentum Mill. Acta Physiol Plant 22:95–101View ArticleGoogle Scholar
- Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254View ArticleGoogle Scholar
- Cohen H, Demaggio AE (1986) Biochemistry of fern spore germination: protease activity in ostrich fern spores. Plant Physiol 80:992–996View ArticleGoogle Scholar
- Crespi M, Zabaleta E, Pontis H, Salerno G (1991) Sucrose synthase expression during cold acclimation in wheat. Plant Physiol 96:887–891View ArticleGoogle Scholar
- Du Y, Nose A (2002) Effects of chilling temperature on the activity of enzymes of sucrose synthesis and the accumulation of saccharides in leaves of three sugarcane cultivars differing in cold sensitivity. Photosynthetica 40:389–395View ArticleGoogle Scholar
- Ellis WL, Bowles JW, Erickson AA, Stafford N, Bell SS, Thomas M (2006) Alteration of the chemical composition of mangrove (Laguncularia racemosa) leaf litter fall by freeze damage. Estuar Coast Shelf Sci 68:363–371View ArticleGoogle Scholar
- Farquhar GD, Sharkey TD (1982) Stomatal conductance and photosynthesis. Annu Rev Plant Physiol 33:317–345View ArticleGoogle Scholar
- Fahnenstich H, Scarpeci TE, Valle EM, Flügge UI, Maurino VG (2008) Generation of hydrogen peroxide in chloroplasts of Arabidopsis overexpressing glycolate oxidase as an inducible system to study oxidative stress. Plant Physiol 148:719–729View ArticleGoogle Scholar
- Giannopolitis C, Ries S (1977) Superoxide dismutases: I. Occurrence in higher plants. Plant Physiol 59:309–314View ArticleGoogle Scholar
- Gilman EL, Ellison J, Duke NC, Field C (2008) Threats to mangroves from climate change and adaptation options: a review. Aquat Bot 89:237–250View ArticleGoogle Scholar
- Haines BL, Dunn EL (1976) Growth and resource allocation responses of Spartina alterniflora Loisel. to three levels of NH4 +-N, Fe and NaCl in solution culture. Bot Gaz 137:224–230View ArticleGoogle Scholar
- Hendrix DL (1993) Rapid extraction and analysis of nonstructural carbohydrates in plant tissues. Crop Sci 33:1306–1311View ArticleGoogle Scholar
- Hodges D, Delong J, Forney C, Prange R (1999) Improving the thiobarbituric acid-reactive-substances assay for estimating lipid peroxidation in plant tissues containing anthocyanin and other interfering compounds. Planta 207:604–611View ArticleGoogle Scholar
- Hu W, Song X, Shi K, Xia X, Zhou Y, Yu J (2008) Changes in electron transport, superoxide dismutase and ascorbate peroxidase isoenzymes in chloroplasts and mitochndria of cucumber leaves as infulenced by chilling. Photosynthetica 46:581–588View ArticleGoogle Scholar
- Hubbard NL, Huber SC, Pharr DM (1989) Sucrose phosphate synthase and acid invertase as determinants of sucrose concentration in developing Muskmelon (Cucumis melo L.) fruits. Plant Physiol 91:1527–1534View ArticleGoogle Scholar
- Kao WY, Shih CN, Tsai TT (2004) Sensitivity to chilling temperatures and distribution differ in the mangrove species Kandelia candel and Avicennia marina. Tree Physiol 24:859–864View ArticleGoogle Scholar
- Lee MA, Chun HS, Kim JW, Lee H, Lee DL, Lee CB (2004) Changes in antioxidant enzyme activities in detached leaves of cucumber exposed to chilling. J Plant Biol 47:117–123View ArticleGoogle Scholar
- Lee T, Lin Y (1995) Change in soluble and cell wall-bound peroxidase activities with growth in anoxia-treated rice (Oryza sativa L.) coleoptiles and roots. Plant Sci 106:1–7View ArticleGoogle Scholar
- Li X, Gong B, Xu K (2014) Interaction of nitric oxide and polyamines involves antioxidants and physiological strategies against chilling-induced oxidative damage in Zingiber officinale Roscoe. Sci Hortic 170:237–248View ArticleGoogle Scholar
- Liu K, Liu L, Liu H, Li X, Wang S (2014) Exploring the effects of biophysical parameters on the spatial pattern of rare cold damage to mangrove forests. Remote Sens Environ 150:20–33Google Scholar
- Mehta R, Mattoo A (1996) Isolation and identification of ripening-related tomato fruit carboxypeptidase. Plant Physiol 110:875–882View ArticleGoogle Scholar
- Moore S, Stein WH (1954) A modified ninhydrin reagent for the photometric determination of amino acids and related compounds. J Biol Chem 211:907–913Google Scholar
- Moloi MJ, van der Westhuizen AJ (2006) The reactive oxygen species are involved in resistance responses of wheat to the Russian wheat aphid. J Plant Physiol 163:1118–1125View ArticleGoogle Scholar
- Naidoo G (2016) The mangrove of South Africa: An ecophysiological review. S Afr J Bot http://dx.doi.org/10.1016/j.sajb.2016.04.014
- Nakano Y, Asada K (1981) Hydrogen peroxide is scavenged by ascorbate-specific peroxidase in spinach chloroplasts. Plant Cell Physiol 22:867–880Google Scholar
- Peng Y, Wang Y, Chen H, Sun C, Wu P, Wang L (2013) Characterization and expression analysis of three CBF/DREB1 transcriptional factor genes from mangrove Avicennia marina. Aquat Toxicol 140–141:68–76View ArticleGoogle Scholar
- Peng Y, Wang Y, Fei J, Sun C, Cheng H (2015) Ecophysiological differences between three mangrove seedlings (Kandelia obovata, Aegiceras corniculatum, and Avicennia marina) exposed to chilling stress. Ecotoxicology 24:1722–1732View ArticleGoogle Scholar
- Prasad TK (1996) Mechanisms of chilling-induced oxidative stress injury and tolerance in developing maize seedlings: changes in antioxidant system, oxidation of proteins and lipids, and protease activities. Plant J 10:1017–1026View ArticleGoogle Scholar
- Quisthoudt K, Schmitz N, Randin C, Dahdouh-Guebas F, Robert E, Koedam N (2012) Temperature variation among mangrove latitudinal range limits worldwide. Trees 26:1919–1931View ArticleGoogle Scholar
- Ross M, Ruiz P, Sah J, Hanan E (2009) Chilling damage in a changing climate in coastal landscapes of the subtropical zone: a case study from south Florida. Global Chang Biol 20:147–157Google Scholar
- Saintilan N, Wilson NC, Rogers K, Rajkaran A, Krauss KW (2014) Mangrove expansion and salt marsh decline at mangrove poleward limits. Global Chang Biol 20:147–157View ArticleGoogle Scholar
- Seigneurin-Berny D, Salvi D, Dorne A, Joyard J, Rolland NH (2008) Percoll-purified and photosynthetically active chloroplast from Arabidopsis thaliana leaves. Plant Physiol Biochem 158:847–852Google Scholar
- Sheue CR, Liu HY, Yong JWH (2003) Kandelia obovata (Rhizophoraceae), a new mangrove species from Eastern Asia. Taxon 52:287–294View ArticleGoogle Scholar
- Stevens P, Fox S, Montague C (2006) The interplay between mangroves and saltmarshes at the transition between temperature and subtropical climate in Florida. Wetl Ecol Manag 14:435–444View ArticleGoogle Scholar
- Stuart SA, Choat B, Martin KC, Holbrook NM, Ball MC (2006) The role of freezing in setting the latitudinal limits of mangrove forests. New Phytol 173:576–583View ArticleGoogle Scholar
- Sui N, Li M, Liu XY, Wang N, Fang W, Meng QW (2007) Response of xanthophyll cycle and chloroplastic antioxidant enzymes to chilling stress in tomato over-expressing glycerol-3-phosphate acyltransferase gene. Photosynthetica 45:447–454View ArticleGoogle Scholar
- Sui X, Zhang B, Zhang Z, Mao SL, Wang L (2005) Differences of photosynthetic characteristics and low light-tolerance in seedlings of four pepper cultivars. Acta Hortic Sin 32: 222–227Google Scholar
- Van den Ende W, Valluru R (2009) Sucrose, sucrosyl oligosaccharides, and oxidative stress: scavenging and salvaging? J Exp Bot 60:9–18View ArticleGoogle Scholar
- Wang W, You S, Wang Y, Huang L, Wang M (2011) Influence of frost on nutrient resorption during leaf senescence in a mangrove at its latitudinal limit of distribution. Plant Soil 342:105–115View ArticleGoogle Scholar
- Wang XD, Yu ZW, Wang D (2003) Effect of potassium on flag leaf proteinases activity and kernel quality in wheat. Acta Agron Sin 29:285–289Google Scholar
- Wu Z, Zhang X, He B, Diao L, Sheng S, Wang J, Guo X, Su N, Wang L, Jiang L, Wang C, Zhai H, Wan J (2007) A chlorophyll-deficient rice mutant with impaired chlorophyllide esterification in chlorophyll biosynthesis. Plant Physiol 145:29–40View ArticleGoogle Scholar
- Zhang X, Jiang D, Zheng C, Dai T, Cao W (2011) Post-anthesis salt and combination of salt and waterlogging affect distributions of sugars, amino acids, Na+ and K+ in wheat. J Agron Crop Sci 197:31–39View ArticleGoogle Scholar
- Zheng C, Liu W, Qiu J, Huang L, Huang X, Chen S (2013) Comparison of physiological characteristics of Kandelia obovata at different ages in winter in the northernmost mangrove transplanted area of China. Acta Ecol Sin 33:132–138View ArticleGoogle Scholar
- Ziska LH, Seemann JR, DeJong TM (1990) Salinity induced limitations on photosynthesis in Prunus salicina, a deciduous tree species. Plant Physiol 93:864–870View ArticleGoogle Scholar
- Zheng C, Ye Y, Liu W, Tang J, Zhang C, Qiu J, Chen J (2016) Recovery of photosynthesis, sucrose metabolism, and proteolytic enzymes in Kandelia obovata from rare cold events in the northernmost mangrove, China. Ecol Process 5:9View ArticleGoogle Scholar