Distinct patterns of nitrate reductase activity in brown algae: Light and ammonium sensitivity in Laminaria digitata is absent in Fucus species

Fucus and Laminaria species, dominant seaweeds in the intertidal and subtidal zones of the temperate North Atlantic, experience tidal cycles that are not synchronized with light:dark (L:D) cycles. To investigate how nutrient assimilation is affected by light cycles, the activity of nitrate reductase (NR) was examined in thalli incubated in outdoor tanks with flowing seawater and natural L:D cycles. NR activity in Laminaria digitata (Huds.) Lamour. showed strong diel patterns with low activities in darkness and peak activities near midday. This diel pattern was controlled by light but not by a circadian rhythm. In contrast, there was no diel variation in NR activity in Fucus serratus L., F. vesiculosus (L.) Lamour., and F. spiralis L. either collected directly from the shore or maintained in the outdoor tanks. In laboratory cultures, transfer to continuous darkness suppressed NR activity in L. digitata, but not in F. vesiculosus; continuous light increased NR activity in L. digitata but decreased activity in F. vesiculosus. Furthermore, 4 d enrichment with ammonium (50 mu mol . L-1 pulses), resulted in NR activity declining by > 80% in L. digitata, but no significant changes in F. serratus. Seasonal differences in maximum NR activity were present in both genera with activities highest in late winter and lowest in summer. This is the first report of NR activity in any alga that is not strongly regulated by light and ammonium. Because light and tidal emersion do not always coincide, Fucus species may have lost the regulation of NR by light that has been observed in other algae and higher plants.

Ca(2+) signals coordinate zygotic polarization and cell cycle progression in the brown alga Fucus serratus

Zygotes of the fucoid brown algae provide excellent models for addressing fundamental questions about zygotic symmetry breaking. Although the acquisition of polarity is tightly coordinated with the timing and orientation of the first asymmetric division-with zygotes having to pass through a G1/S-phase checkpoint before the polarization axis can be fixed -the mechanisms behind the interdependence of polarization and cell cycle progression remain unclear. In this study, we combine in vivo Ca(2+) imaging, single cell monitoring of S-phase progression and multivariate analysis of high-throughput intracellular Ca(2+) buffer loading to demonstrate that Ca(2+) signals coordinate polarization and cell cycle progression in the Fucus serratus zygote. Consistent with earlier studies on this organism, and in contrast to animal models, we observe no fast Ca(2+) wave following fertilization. Rather, we show distinct slow localized Ca(2+) elevations associated with both fertilization and S-phase progression, and we show that both S-phase and zygotic polarization are dependent on pre-S-phase Ca(2+) increases. Surprisingly, this Ca(2+) requirement cannot be explained by co-dependence on a single G1/ S-phase checkpoint, as S phase and zygotic polarization are differentially sensitive to pre-S-phase Ca(2+) elevations and can be uncoupled. Furthermore, subsequent cell cycle progression through M phase is independent of localized actin polymerization and zygotic polarization. This absence of a morphogenesis checkpoint, together with the observed Ca(2+)dependences of S phase and polarization, show that the regulation of zygotic division in the brown algae differs from that in other eukaryotic model systems, such as yeast and Drosophila.

A tip-high, Ca2+-interdependent, reactive oxygen species gradient is associated with polarized growth in Fucus serratus zygotes

We report the existence of a tip-high reactive oxygen species (ROS) gradient in growing Fucus serratus zygotes, using both 5-(and 6-) chloromethyl-2',7'-dichlorodihydrofluorescein and nitroblue tetrazolium staining to report ROS generation. Suppression of the ROS gradient inhibits polarized zygotic growth; conversely, exogenous ROS generation can redirect zygotic polarization following inhibition of endogenous ROS. Confocal imaging of fluo-4 dextran distributions suggests that the ROS gradient is interdependent on the tip-high [Ca2+](cyt) gradient which is known to be associated with polarized growth. Our data support a model in which localized production of ROS at the rhizoid tip stimulates formation of a localized tip-high [Ca2+](cyt) gradient. Such modulation of intracellular [Ca2+](cyt) signals by ROS is a common motif in many plant and algal systems and this study extends this mechanism to embryogenesis.

Arsenic is not stored as arsenitephytochelatin complexes in the seaweeds Fucus spiralis and Hizikia fusiforme

Environmental context Seaweeds hyperaccumulate the toxic metalloid arsenic, but seemingly achieve detoxification by transformation to arsenosugars. The edible seaweed hijiki is a notable exception because it contains high levels of toxic arsenate and arsenite. Terrestrial plants detoxify arsenic by forming arsenitephytochelatin complexes. The hypothesis that seaweeds also synthesise phytochelatins to bind arsenite as a means of detoxification before arsenosugar synthesis is tested in this investigation. Abstract Phytochelatins (PCs), generic structure [-Glu-Cys]n-Gly, are peptides synthesised by terrestrial plants to bind toxic metal(loid)s such as cadmium and arsenic. Seaweeds are arsenic hyperaccumulators, seemingly achieving detoxification via arsenosugar biosynthesis. Whether seaweeds synthesise PCs to aid detoxification during arsenic exposure is unknown. Hizikia fusiforme (hijiki) and Fucus spiralis were used as model seaweeds: the former is known for its large inorganic arsenic concentration, whereas the latter contains mainly arsenosugars. F. spiralis was exposed to 0, 1 and 10mgL ^-1 arsenate solutions for 24h, whereas hijiki was analysed fresh. All samples contained As ^III, glutathione and reduced PC ₂, identified using HPLC-ICP-MS/ES-MS. Although hijiki contained no As ^IIIPC complexes, arsenate exposed F. spiralis generated traces of numerous arsenic compounds that might be As ^IIIGS or As ^IIIPC ₂ complexes. As ^IIIPC complexes seem not to be a principal storage form for long-term arsenic storage within seaweeds. However, 40 times higher glutathione concentrations were found in hijiki than F. spiralis, which may explain how hijiki deals with its high inorganic arsenic burden. © 2011 CSIRO.

Ação de polissacarídeos sulfatados de Fucus Vesiculosus na Hemostasia e no sistema complemento

Fucans are a family of sulfated homo and teropolysaccharides respectively, composed mainly of a- (1®2) and a- (1®3) linked by L-fucose residues. Properties such as the ability to act as an anti-contraceptive, to reduce cholesterol levels, and to act as an anti-tumor agent are much related. We have focused our attention on the anticoagulant properties, platelet aggregation, hemorrhagic activity and complement system in vitro of commercial fucoidan (F) and their purified fractions (F1, F2 and F3) from Fucus vesiculosus obtained from fractionation of the fucoidan with different concentrations of acetone 1, 2 and 3v. These compounds were chemically characterized and the fucoidan (F) was modified by desulfation. The anticoagulant activity of the compounds was assessment by activated partial thromboplastin time (APTT) and prothrombine time assay (PT) using citrated normal human plasma. The results of APPT test showed that F, F1 and F2 have high anticoagulants activities 240.0 s (5 µg). The F3 showed 73.7 s in the same concentrations. The results obtained with PT test to F, F1, F2 and F3 were 81.5 s, 120.0 s, 57.1 and 32.5 s respectively with 50 µg. The dessulfated polymer showed a decrease in the anticoagulant activity in these two tests. Platelet aggregation assay was measured turbidimetrically with platelet aggregometer by method of Born. The aggregation platelet with F and fractions F1, F2 and F3 exhibited a two-phase answer in the concentration of 5 mg/mL with maximum aggregation of 76.36 ± 10.3% ; 69.54 ± 9.40%; 75.94 ± 9.01%; 51.13 ± 9.59% respectively. However, was observed a hipoaggregate profile F (15.17 ± 5.2%), F1 (7.40 ± 3.04 %), F2 (19.1 ± 5.41%) and F3 (5.09 ± 3.02%) at 0.1 mg/mL. The hemorrhagic activity assay was carried in Wistar rats and showed that these compounds have low hemorrhagic effect when compared to heparin. The complement system ( alternative pathway was made using non-sensibilized rabbit red blood cells The results of complement system essay showed that F , F2 and F3 have action inhibitory in relation to the group control 0.544, 0.697, 0.622 and 0.958 respectively The results showed that these compounds have action on this system. Interaction of the polisaccharides with proteins C3 and C4 showed that the fraction F1 stimulated the activity assay hemolytic using red blood cells

Efeito do Fucoidam de Fucus vesiculosus em um modelo experimental de artrite reumatóide

Rheumatoid arthritis (RA) is systemic auto imune disorder. It is caracterized by chronic inflammation of joints leading to progressive erosion of cartilage and bone. We investigated the effect of the administration of fucoidan, sulfated polysaccharides, from algae Fucus vesiculosus in the acute (6h) in zymosan-induced arthritis (AZy). Wistar rats (180-230 g) were used for all groups experimental. Non-treated animals received just intraarticular injection of 1 mg the zymosan, control group received intraarticular injection of 50 µL the saline, groups received either fucoidan of Fucus vesiculosus (15, 30, 50 or 70 mg/Kg) or parecoxib (1 mg/Kg) 1 hour after injection of zymosan. After 6 h, the articular exudates were collected for evaluation of the cell influx and nitrite (Griess reaction) release. The sinovial membranes and articular cartilages were excised for histopathological analysis and by determination of the glycosaminoglycan (GAG), respectively. ZyA led to increased NO and cell influx into the joints. Therapeutic administration of the fucoidan or parecoxib did significantly inhibited the cell influx and the synovitis, as compared to non-treated rats (p<0,05), though being able to reduced NO release. Representative agarose gel electrophoresis of the GAGs, the content of condroitin-sulphate was observed during the process. These findings suggest that the fucoidan from Fucus vesiculosus has potential anti-inflammatory activity

Efeito de frações obtidas da fucoidana de Fucus vesiculosus em modelo experimental de artrite induzida por Zymosan

The fucoidan from Fucus vesiculosus is known for having diverse biological properties. This study analyzed the therapeutic action of populations of commercial fucoidan (F. vesiculosus) on zymosan-induced arthritis. Three populations of fucoidan were obtained after acetone fractionation; these were denominated F1 (52.3%), F2 (36.7%) and F3 (10.7%). Chemical analyses showed that F1 contained the largest amount of sulfate ion. The electrophoretic profile shows that the commercial or total fucoidan (TF), different from the other fucoidans and from glycosaminoglycan patterns, is quite polydisperse, which indicates that it is composed of a mixture of sulfate polysaccharides. On the other hand, the fucoidans obtained from TF showed only an electrophoretic band with much lower polydispersion than that observed for TF. Fucoidan F2 showed a migration between fucoidans F1 and F3. Owing to the small amount of mass obtained from F3, we used only fucoidans F1 and F2 in the induced arthritis tests. After 1 hour of induction, we administered F1 or F2 (10, 25 and 50 mg/kg i.p.) or diclofenac sodium (10 mg/kg i.p.) or lumiracoxib (5 mg/kg o.a.) or L-NAME (30 mg/kg i.p.). After 6 hours, we performed analyses of cell influx and nitrite levels in addition to histopathological analysis. Fucoidans F1 and F2 were more potent both in decreasing the number of leukocytes and the amount of nitric oxide found in the synovial fluid. This indicates that the anti-inflammatory mechanism of these fucoidans is not only related to selectin block, but also to nitric oxide synthesis inhibition