TRANSLOCATION OF PHOTOSYNTHATE IN THE BROWN ALGA NEREOCYSTIS

Translocation of photosynthates was found to occur when blades of Nereocystis were illuminated in the presence of C¹⁴ bicarbonates. Rates of translocation averaging 37 cm/hr in the laboratory were observed. Samples from the epidermis, cortex, and medulla of the stipes of plants with blades exposed to tracer showed that the radioactivity in organic compounds was confined to medulla where sieve filaments occurred. Girdle preparations of blades, interrupting the mucilage ducts and leaving the blade medulla intact, allowed translocation to take place. These data indicate that conduction of photosynthate takes place in the medulla. Similarities between the anatomy of algal sieve filaments and angiosperm sieve tubes, coupled with the continuity of the sieve filaments from blade medulla to stipe medulla suggested indirectly that the sieve filaments were conducting elements. Further support of this hypothesis was provided from collections of radioactive exudate from cells in the medulla of the lower stipe that were continuous with the sieve filaments. Tracer applied to the blades was partially recovered as organic material in a clear fluid that collected inside the pneumatocyst. Continued accumulation of radioactivity in this fluid was dependent on living blades; fluid with low specific activity that did not increase during the experiment accumulated slowly if blades were killed with ethanol after an exposure to tracer. It is possible that the system that produced the stipe fluid was part of (or a side effect of) the system responsible for maintaining volume in the conducting system. It may also provide an alternate route (other than the sieve filaments) for delivery of photosynthates to the base of the plant. Carbon-14 applied to blade tips as bicarbonate was recovered in part as radioactive mannitol in the translocation stream.     

REACTIVE OXYGEN METABOLISM IN INTERTIDAL FUCUS SPP. (PHAEOPHYCEAE)

Our previous research suggests that interspecific variation in stress tolerance in intertidal Fucus spp. (Phaeophyceae) is partially mediated by differences in the production of, or ability to detoxify, reactive oxygen. Here we report on the content of antioxidants (ascorbate, glutathione, carotenoids, and tocopherols) and protective enzymes (catalase, superoxide dismutase, ascorbate peroxidase, and glutathione reductase) involved in reactive oxygen metabolism in three species of intertidal brown algae— Fucus spiralis L., F. evanescens C. Ag., and F. distichus L.—that differ in stress tolerance and position in the intertidal zone. Contents of the major antioxidants were similar in the three species and were not correlated with stress tolerance. The least stress tolerant species, F. distichus, had the lowest activity of reactive-oxygen-scavenging enzymes, although F. spiralis, the species with the highest stress tolerance, and F. evanescens contained similar activities of antioxidant enzymes on a fresh-weight basis. However, the activities of superoxide dismutase and ascorbate peroxidase in F. evanescens are lower than those of F. spiralis when expressed on the basis of chlorophyll. These data show that the ratio between reactive oxygen protection and production might be more important than the absolute content of antioxidants and protective enzymes. It also shows the importance of localization of detoxifying mechanisms and avoidance of oxidative stress.     

SEASONALITY AND THERMAL ACCLIMATION OF REACTIVE OXYGEN METABOLISM IN FUCUS VESICULOSUS (PHAEOPHYCEAE)

The intertidal brown macroalga Fucus vesiculosus L. acclimates its defense against reactive oxygen in response to both (1) growth at different temperatures in laboratory culture and (2) seasonal changes in environmental conditions. Fucus vesiculosus was grown in seawater at 0° C, 20° C, and at 0° C with a 3-h daily emersion at −10° C. Algae grown at low temperature, both with and without freezing, produced less reactive oxygen after severe freezing stress than those grown at 20° C. These differences were correlated with growth temperature-induced changes in activities of superoxide dismutase (SOD), glutathione reductase, and ascorbate peroxidase. The contents of tocopherols increased with increased cultivation temperature, whereas the activity of catalase and the content of glutathione and ascorbate did not change. Growth at 0° C increased the resistance of photosynthesis to freezing and reduced photoinhibition in high light at 5° C; the latter effect was further increased in algae subject to daily freezing. These data suggest that elevated activity of reactive oxygen scavenging enzymes, especially SOD, increases the resistance to photoinhibition, at least at low temperature, as well as being important for freezing tolerance. Seasonal changes in reactive oxygen metabolism showed a similar pattern to those elicited by temperature in laboratory culture. Summer samples had lower activities of most reactive oxygen scavenging enzymes than algae collected in autumn and winter when water temperatures were lower. In contrast to the laboratory experiments, ascorbate content did change and was lower during the winter than summer, whereas the content of glutathione was not influenced by season. Overall, the data not only indicate that temperature plays an important role in the regulation of stress tolerance and reactive oxygen metabolism but also suggest that other factors are also involved.     

IN VIVO MEASUREMENT OF ACTIVE OXYGEN PRODUCTION IN THE BROWN ALGA FUCUS EVANESCENS USING 2′,7′-DICHLOROHYDROFLUORESCEIN DIACETATE1

Intracellular production of active oxygen in the brown alga Fucus evanescens C. Ag. was studied by measuring the capacity for in vivo conversion of 2′,7′-dichlorohydrofluorescein diacetate (DCFH-DA) to the fluorescent dye 2′,7′-dichlorofluorescein (DCF), both in emersed and immersed seaweeds. Algae were incubated in seawater containing DCFH-DA under a range of conditions, and it was also possible to load algae with DCFH-DA and then follow subsequent DCF production in emersed tissue. DCF formation was linear for at least 2 h in both darkness and light, with the rate of formation increasing with the light level. DCF formation was temperature dependent. It also increased when algae were treated with H₂O₂ or methyl viologen (paraquat), which disrupts photosystem 1 electron transport and increases O^?₂ production. Exogenous catalase reduced in vivo DCF production, presumably by lowering cellular concentrations of H₂O₂. Hydrogen peroxide was released into the seawater by illuminated algae resulting in external dye conversion to DCF. However, this does not interfere with in vivo measurement of DCF by loaded, washed algae because DCF leakage appeared to be negligible. Internal DCF did not affect photosynthetic oxygen production relative to untreated controls. Overall, our data suggest that DCFH-DA is a potentially very useful probe for studying active oxygen metabolism in seaweeds subjected to environmental stresses.     

LIPID COMPOSITION OF THE BROWN ALGA FUCUS VESICULOSUS1,2

The lipids of the brown alga Fucus vesiculosus consist of approximately 3.6% hydrocarbons (including carotenes) and sterol esters, 11.8% triglycerides, 4.2% free fatty acids, 10.6% sterols, 15.1% chlorophylls and xanthophylls, 29.9% glycolipids, and 6.5% phospholipids. Diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, and phosphatidylinositol appear to be present and phosphatidylcholine is absent.     

HETEROMORPHIC LIFE-HISTORY STRATEGIES IN THE BROWN ALGA SCYTOSIPHON LOMENTARIA (LYNGB.) LINK1

The adaptive significance of a life-history strategy, expressed as divergent morphological forms, was examined for the heteromorphic alga Scytosiphon lomentaria. Successional studies were performed by physically clearing mature, temporally-constant intertidal communities on San Clemente I., California. At week six, after clearing, complanate thalli dominated the successional plots (mean cover = 23.5%) and began to decline as the cylindrical form became abundant. The latter attained its peak cover (82.3%) at week 13, whereupon it too began a precipitous reduction. The crustose ralfsioid form appeared surprisingly early (4–13 wks) in trace amounts but did not achieve its greatest cover (85.0%) until week 43. The ranking from high to low primary productivity (cylindrical form = 8.1 mg C · g dry. wt^-1·h^-1, complanate form = 6.5 mg, crustose form = 0.5 mg) corresponded closely with the data for photosynthetic vs. structural components (cylindrical = 92.3% pigmented, complanate = 65.3%, crustose = 32.0%). This finding indicates that selection in the crust form, which is more readily accessible to epilithic grazers, has tended to increase allocation of materials to nonpigmented structural tissue at the expense of photosynthetic tissue and reduced production rates. The results for thallus losses to urchin grazing over 48 h were complanate form = 82.7% lost, cylindrical = 81.4% and crustose = 16.2%, which correlates with the calorific contents of the three forms (i.e. complanate = 4.97 kcal · ash-free g dry wt.^-1, cylindrical = 4.46 kcal and crustose = 3.55 kcal). The crustose form had tougher thalli (26 g · mm^-2 to penetrate thallus) than either the complanate form (12 g) or the cylindrical form (15 g). It is likely that opposing selective factors have resulted in the evolutionary divergences observed in algae with heteromorphic life histories. Previous work may have overemphasized the selective role of grazing because the crustose form is also adapted to withstand physical forces (sand-scouring, burial and wave-shearing) or as an overwintering stage under physiologically stressful conditions.     

雨生红球藻抗氧化系统对活性氧的清除机制

雨生红球藻(Haematococcus pluvialis)是一种淡水单细胞绿藻,隶属绿藻门、团藻目、红球藻科、红球藻属。它受强光、氮饥饿、高盐等胁迫时积累大量的虾青素,含量可高达细胞干重的6.0%以上[1]。相关研究也显示来虾青素具有极强的抗氧化活性[2—4],它的抗氧化活性较α-生育酚强千倍[5],比维生素E高近百倍[6]。雨生红球藻产生     

CHLOROPLAST AND MITOCHONDRIAL DNA IN A BROWN ALGA EGREGIA MENZIESII

Chloroplasts and mitochondria of the brown alga Egregia menziesii were studied with the electron microscope. In both organelles, 15-25-A fibrils with DNA characteristics are found within areas of electron transparency. In each chloroplast there are two DNA-containing areas, one at each tip of the chloroplast. This localization, the shape and size of each DNA-containing area, and its close association with lamellae in a nondividing chloroplast are noted. One or occasionally two DNA-containing areas are found within the mitochondrion and they are compared with a similar structure in the chloroplast.     

大豆萌发过程的活性氧代谢

本文研究了大豆萌发过程中活性氧的产生与清除,并探讨了光因子在活性氧代谢中的作用。大豆呼吸强度、O产生速率及H2O2水平都在吸水后第四天达到高峰,然后下降,三者的变化趋势同步。SOD、POD及APX的活性随萌发过程而逐渐增强,最后趋于平稳。SOD同工酶谱中分别于萌发的第二、第三天各出现一条新的酶带。CAT在萌发的初期猛增50倍左右,之后趋于稳定。在三种清除H2O2的酶（CAT、POD、APX）中,CAT清除H2O2的能力远远高于POD与APX,CAT可能是大豆萌发过程中最主要的H2O2清除酶。光萌发时呼吸强度低于暗中萌发,但O产生速率与H2O2水平高于暗萌发,光萌发时O的产生占总耗氧量的1．1—2．7％,而暗中萌发为0．9—1．3％。光条件下SOD、APX活性明显高于暗中萌发,而POD与CAT则在光和暗条件下相差不大。     

NEW PYRENOID FORMATION IN THE BROWN ALGA,SCYTOSIPHON LOMENTARIA (SCYTOSIPHONALES,PHAEOPHYCEAE)1

The Scytosiphon lomentaria (Lyngbye) Link cell characteristically has only one chloroplast with a prominent protruding pyrenoid. We observed the appearance of a new pyrenoid in each chloroplast during first mitosis in zygotes of S. lomentaria, using the freeze substitution technique. At first, a pyrenoid matrix appeared within the outermost stroma, in which thylakoid triplets and ribosomes were absent. At this time, the surface of this part remained smooth. The old pyrenoid was covered with a pyrenoid cap on the cytoplasmic side, whereas there was no pyrenoid cap on the new pyrenoid before protrusion. Irregularly shaped membranous sacs containing fine granular materials associated with the cytoplasmic side of the new pyrenoid. The sacs fused with each other and changed conformation and finally transformed into the pyrenoid cap. The new pyrenoid gradually protruded toward the cytoplasm, and the new pyrenoid cap became curved along the surface of pyrenoid. Cytokinesis occurred, and each chloroplast had two prominent protruding pyrenoids in two‐celled zygotes. We examined immunolocalization of β‐1,3‐glucans within the pyrenoid cap with a monoclonal antibody, using EM. Gold particles indicating localization of β‐1,3‐glucans were detected in vacuoles but never in the pyrenoid cap. This observation suggests that the pyrenoid cap in brown algae contains no photosynthetic products such as polysaccharide.     

SOME FINE STRUCTURAL OBSERVATIONS ON DEVELOPING AND MATURE SIEVE ELEMENTS IN THE BROWN ALGA LAMINARIA SACCHARINA

The differentiation of sieve elements from inner cortical cells of the stipe of Laminaria saccharina (L.) Lamour. involves the development of a well-structured protoplast and an end wall possessing evenly spaced pores which are visualized by electron microscopy. The protoplast consists of organelles which are commonly found in brown algal cells, including nuclei, cup- or horseshoe-shaped chloroplasts, dictyosomes, mitochondria, and ER. Mitochondria and clusters of small vacuoles, presumably redistributed by the surging effect which occurs in sieve elements, were routinely observed in the vicinity of the end wall. Chloroplasts were seen in progressively degenerated states in older sieve elements, yet nuclei were determined to be non-necrotic. Numerous pores along the end walls interconnect adjacent sieve elements. Each pore is traversed by a strand of cytoplasm and surrounded by plasmalemma. The pores are open and possess no callose. In this paper the sieve element ultrastructures of L. saccharina are compared to those in L. groenlandica, Alaria marginata, Nereocystis lutkeana and Macrocystis pyrifera, and a possible phylogenetic specialization of sieve elements is presented in table form and discussed.     

细胞质雄性不育水稻的呼吸作用和自由基代谢的关系初探

用呼吸电子传递细胞色素途径的抑制剂氰化钾（ＫＣＮ）与抗氰呼吸途径的抑制剂水杨基氧肟酸（ＳＨＡＭ）处理水稻细胞质雄性不育系（ＣＭＳ）珍汕９７Ａ及其保持系珍汕９７Ｂ的幼穗和花药后,ＫＣＮ使不育系与保持系的超氧阴离子自由基（Ｏ２■）产生受到抑制,不育系的Ｏ２■的形成受抑制较多。ＳＨＡＭ处理则增高Ｏ２■形成,以不育系的增加较多．ＫＣＮ与ＳＨＡＭ处理后都使不育系与保持系的丙二醛（ＭＤＡ）含量升高,ＫＣＮ使保持系的ＭＤＡ含量升高较多,ＳＨＡＭ则使不育系的ＭＤＡ含量升高较多．ＫＣＮ处理后,不育系与保持系的超氧物歧化酶（ＳＯＤ）活性下降,ＳＨＡＭ处理后不育系与保持系的ＳＯＤ活性变化不明显。Ｈ２Ｏ２处理对不育系与保持系幼穗的呼吸速率影响不大．Ｈ２Ｏ２＋ＦｅＳＯ４处理后,使呼吸速率大幅度下降,表明Ｈ２Ｏ２＋ＦｅＳＯ４所形成的羟自由基（ＯＨ）比Ｈ２Ｏ２对呼吸代谢的破坏作用更大。     

OVERWINTERING OF DICTYOTA DICHOTOMA (PHAEOPHYCEAE) NEAR ITS NORTHERN DISTRIBUTION LIMIT ON THE EAST COAST OF NORTH AMERICA1

The mechanism of yearly repopulation of Dictyota dichotoma (Hudson) Lamouroux in the inshore waters of North Carolina was studied in the field and in laboratory experiments. It was demonstrated that D. dichotoma is maintained throughout the year at this location and does not require yearly immigrations. A young germling stage was shown to function as the overwintering structure. Although release of aplanospores was unaffected by temperature and photoperiod, subsequent germination and establishment required warm conditions. If germlings were allowed to grow to the 7–9 cell stage under summer conditions, they could survive 21 wk of winter conditions. Transfer of 21 wk old winterized germlings to summer conditions resulted in the rapid production of erect blades. It therefore appears that D. dichotoma maintains its populations near its northern distribution limit on the East Coast of North America by undergoing dormancy after germling establishment.     

THE ROLE OF REACTIVE OXYGEN SPECIES IN COPPER TOXICITY TO TWO FRESHWATER GREEN ALGAE1

The role of reactive oxygen species (ROS) in copper (Cu) toxicity to two freshwater green algal species, Pseudokirchneriella subcapitata (Korshikov) Hindák and Chlorella vulgaris Beij., was assessed to gain a better mechanistic understanding of this toxicity. Cu‐induced formation of ROS was investigated in the two algal species and linked to short‐term effects on photosynthetic activity and to long‐term effects on cell growth. A light‐ and time‐dependent increase in ROS concentrations was observed upon exposure to environmentally relevant Cu concentrations of 50 and 250 nM and was comparable in both algal species. However, effects of 250 nM Cu on photosynthesis were different, leading to a 12% reduction in photosynthetic activity in P. subcapitata, but not in C. vulgaris. These results indicate that differences in species‐specific sensitivities measured as photosynthetic activity were not caused by differences in the cellular ROS content of the algae, but probably by different species‐specific ROS defense systems. To investigate the role of ROS in Cu‐mediated inhibition of photosynthesis, the ROS scavenger N‐tert‐butyl‐α‐phenylnitrone (BPN) was used, resulting in a reduction of Cu‐induced ROS production up to control level and a complete restoration of photosynthetic activity of Cu‐exposed P. subcapitata. This finding implied that ROS play a primary role in Cu toxicity to algae. Furthermore, we observed a time‐dependent ROS release process across the plasma membrane. More than 90% of total ROS were determined to be extracellular in P. subcapitata, indicating an efficient method of cellular protection against oxidative stress.     

NITROGEN METABOLISM AND AMINO ACID NUTRITION IN THE SOIL ALGA STICHOCOCCUS BACILLARIS (CHLOROPHYCEAE)1

Nitrate-grown cells of Stichococcus bacillaris Naeg. (UTEX 314) contained much higher activities of glutamine synthetase (GS) and NADPH-glutamate dehydrogenase (GDH) than ammonium-grown cells. Methylamine, a non-metabolizable ammonium analog, caused a decrease in GS activity in nitrate-grown cells suggesting that GS is regulated by the size of the endogenous ammonium pool. The decrease in GS observed in methylammonium-loaded nitrate-grown cells was accompanied by an increase in NADPH-GDH activity. Stichococcus bacillaris can be grown in the presence of methionine sulfoximine (MSX), a potent inhibitor of GS. However, only a fraction of a control cell population showed a requirement for glutamine or arginine for growth following MSX addition. Fully adapted MSX-grown cells were indistinguishable from control cells in their ability to photosynthesize and utilize amino acids as nitrogen sources. Alanine, arginine, asparagine, glutamine, glycine and proline were good nitrogen sources, and maximum capacity for amino acid transport was developed in cells grown on these amino acids. Compared to nitrate-grown cells the activity of GS in ammo acid-grown cells was low, whereas NADPH-GDH was very active. The activity of NADH-GDH in amino acid-grown cells was highest under heterotrophic conditions.