Marianne Pedersén

The unique features of starch metabolism in red algae

Red algae (Rhodophyceae) are photosynthetic eukaryotes that accumulate starch granules outside of their plastids. The starch granules from red algae (floridean starch) show structural similarities with higher plant starch granules but lack amylose. Recent studies have indicated that the extra–plastidic starch synthesis in red algae proceeds via a UDP glucose–selective α–glucan synthase, in analogy with the cytosolic pathway of glycogen synthesis in other eukaryotes. On the other hand, plastidic starch synthesis in green cells occurs selectively via ADP glucose in analogy with the pathway of glycogen synthesis in prokaryotes from which plastids have evolved. Given the emerging consensus of a monophyletic origin of plastids, it would appear that the capacity for starch synthesis selectively evolved from the α–glucan synthesizing machinery of the host ancestor and its endosymbiont in red algae and green algae, respectively. This implies the evolution of fundamentally different functional relationships between the different subcellular compartments with regard to photosynthetic carbon metabolism in these organisms. It is suggested that the biochemical and molecular elucidation of floridean starch synthesis may offer new insights into the metabolic strategies of photosynthetic eukaryotes.

A study of the diurnal variation of biogenic volatile halocarbons

The diurnal variation of a number of naturally produced volatile halogenated organic compounds (VHOC) in a pool at Gran Canaria, Spain, is shown. The diurnal cycle was studied both in water and air samples. The highest production rates of VHOC were observed at mid-day, when the photosynthesis of the algae was assumed to be maximal. A second production period was observed after sunset, probably caused by algal respiration. The production rates were found to be equal during light and dark conditions. The measured rates correspond well to our earlier laboratory experiments. A rapid decrease in the concentration of VHOC in the water was observed, and the magnitude of the decrease is compared with the calculated emission of VHOC to the atmosphere. The diurnal cycle of chlorinated and brominated compounds, as well as for 1-iodobuthane, differed from those of iodomethane and 2-iodopropane, probably due to different formation pathways.

Effects of temperature on the production of hydrogen peroxide and volatile halocarbons by brackish-water algae

Marine algae produce volatile halocarbons, which have an ozone-depleting potential. The formation of these compounds is thought to be related to oxidative stress, involving H2O2 and algal peroxidases. In our study we found strong correlations between the releases of H2O2 and brominated and some iodinated compounds to the seawater medium, but no such correlation was found for CHCl3, suggesting the involvement of other formation mechanisms as well. Little is known about the effects of environmental factors on the production of volatile halocarbons by algae and in the present study we focused on the influence of temperature. Algae were sampled in an area of the brackish Baltic Sea that receives thermal discharge, allowing us to collect specimens of the same species that were adapted to different field temperature regimes. We exposed six algal species (the diatom Pleurosira laevis, the brown alga Fucus vesiculosus and four filamentous green algae, Cladophora glomerata, Enteromorpha ahlneriana, E. flexuosa and E. intestinalis) to temperature changes of 0-11 degrees C under high irradiation to invoke oxidative stress. The production rates, as well as the quantitative composition of 16 volatile halocarbons, were strongly species-dependent and different types of responses to temperature were recorded. However, no response patterns to temperature change were found that were consistent for all species or for all halocarbons. We conclude that the production of certain halocarbons may increase with temperature in certain algal species, but that the amount and composition of the volatile halocarbons released by algal communities are probably more affected by temperature-associated species shifts. These results may have implications for climatic change scenarios.

Patterns of macroalgal diversity, community composition and long-term changes along the Swedish west coast

This study analyses the complicated patterns of vertical distribution of the macroalgal vegetation in an area where brackish and marine waters meet and mix. Variables used to record vegetation characteristics are algal cover, species composition and diversity. The data set includes 64 diving profiles, all from sites exposed to wave action, along a ca. 260 km long coastline. The profiles belong to four categories: coastal sites in the Skagerrak (more marine), coastal sites in the Kattegat (more brackish), coastal sites in the Kattegat after a toxic phytoplankton bloom, and submerged offshore stone reefs in the Kattegat. The highest species diversity was found at the reefs, which are not affected directly by land runoff. At the reefs the 18 most common perennial species penetrate 2–11 m (on average 5.5 m) deeper than at the coastal sites. The virtual absence of sedimentation, and thus the availability of substratum, at the reefs may explain the differences so that the lower limit for the algae is determined by light penetration or by recruitment problems caused by strong currents at the reefs, whereas sedimentation limits the settlement of algae in coastal sites. Ordination analysis based on species composition reveals that the major environmental gradients structuring the algal vegetation in the Kattegat and the Skagerrak are salinity and water depth. The large data set of this study made it possible to quantify the downward dislocation of Atlantic intertidal species to the sublittoral along the Swedish west coast. For example, the mean upper limit of Corallina officinalis is 2 m in the Skagerrak but 12.5 m in the Kattegat and the mean occurrence interval of Fucus serratus is 0.9–2.7 m in the Skagerrak, but 1.1–6.3 m in the Kattegat. This downward dislocation is suggested to be the result of decreased competition when species successively disappear with lower salinity. Comparisons of the present studys results with those of previous investigations show that eight common red algal species have moved upwards compared to the situation before the large-scale eutrophication started in the 1960s, e.g. Cystoclonium purpureum and Polysiphonia elongata by ca. 8 m, Phycodrys rubens and Delesseria sanguinea by ca. 5 m. A toxic phytoplankton bloom affected macroalgal community composition on the whole only slightly, but it had a negative effect on algal cover and species richness below a water depth of ca. 5 m, the algae were visibly damaged and the lower vegetation limit temporarily moved upwards.

UPTAKE OF INORGANIC CARBON BY CLADOPHORA GLOMERATA (CHLOROPHYTA) FROM THE BALTIC SEA1

Carbon uptake in the green macroalga Cladophora glomerata (L.) Kütz. from the brackish Baltic Sea was studied by recording changes in pH, alkalinity, and inorganic carbon concentration of the seawater medium during photosynthesis. The use of specific inhibitors identified three uptake mechanisms: 1) dehydration of HCO3− into CO2 by periplasmic carbonic anhydrase, followed by diffusion of CO2 into the cell; 2) direct uptake of HCO3− via a 4,4′‐diisothiocyanato‐stilbene‐2,2′‐disulfonate‐sensitive mechanism; and 3) uptake of inorganic carbon by the involvement of a vanadate‐sensitive P‐type H+‐ATPase (proton pump). A decrease in the alkalinity of the seawater medium during carbon uptake, except when treated with vanadate, indicated a net uptake of the ionic species contributing to alkalinity (i.e. HCO3−, CO32−, and OH−) from the medium, where OH− influx is equivalent to H+ efflux. This would suggest that the proton pump is involved in HCO3− transport. We also show that the proton pump can be induced by carbon limitation. The inducibility of carbon uptake in C. glomerata may partly explain why this species is so successful in the upper littoral zone of the Baltic Sea. Usually, carbon limitation is not a problem in the upper littoral of the sea. However, it may occur frequently within dense Cladophora belts with high photosynthetic rates that create high pH and low carbon concentrations in the algas microenvironment.

Active carbon uptake in Laminaria digitata and L . saccharina (Phaeophyta) is driven by a proton pump in the plasma membrane

Mechanisms for inorganic carbon acquisition in Laminaria digitata (Hudson) Lamour and L.xa0saccharina (L.) Lamour from the Swedish west coast were studied in pH-drift experiments, using several inhibitors for different types of carbon uptake across the cell membrane. Throughout the experiments total carbon decreased in concert with a pH increase while alkalinity stayed relatively stable. Addition of acetazolamide had a strong inhibitory effect on the carbon uptake rate in L.xa0digitata and the anion exchange protein inhibitor DIDS had a small inhibitory effect above pH 9.5. These results indicate that carbon uptake in L.xa0digitata depends on the presence of an external carbonic anhydrase while direct bicarbonate uptake may contribute at high pH. These two mechanisms have previously been shown to occur in L.xa0saccharina. We show that two inhibitors of H+-ATPases, vanadate and erythrosin B, also decreased carbon uptake rates in both Laminaria species. The effect of erythrosin B was immediate and it probably acts on the outside of the cell membrane. Contrarily, vanadate needs to be transported into the cell, where it competes with the phosphate from ATP for the aspartic acid phosphorylation site on the plasma membrane P-type H+-ATPase. Therefore, 1–2xa0h of pH drift were usually required before an inhibitory effect became apparent. Additional experiments with P-enriched and P-starved L.xa0saccharina corroborated this process. Based on these results we suggest that the investigated Laminaria species, besides external carbonic anhydrase and DIDS-sensitive anion exchange, also possess a mechanism for the active uptake of carbon, which is dependent on plasma membrane P-type H+-ATPase activity. This paper also reports on the buffering capacity of the inhibitors when used in natural seawater, an aspect that has been neglected in previous studies.

Long-term changes of macroalgal vegetation in the Skagerrak area

The algal vegetation at three rocky-shore localities on the Swedish Skagerrak coast with different environmental conditions was studied in 1960–1961 by SCUBA diving. The same localities were revisited in the summer of 1997, using the same methods for recording the vegetation. Detailed descriptions of the vegetation profiles are presented to allow the recording of future changes in these profiles, and extensive ecological herbaria are kept of both the 1960–1961 and the 1997 investigations. The abundance and depth distribution of 78 macroalgal species were recorded in 1997 and community composition was compared with that of the early 1960s. Increases of perennial red algae with delicate foliaceous thalli (Delesseria sanguinea, Phycodrys rubens) were conspicuous at the two localities most exposed to wave action, whereas perennial red algae with tougher foliaceous thalli (Phyllophora truncata and Phyllophora pseudoceranoides) prevailed at the more sheltered locality with most sedimentation. It is hypothesised that increased abundances of delicate species with a large growth potential are caused by eutrophication, but that this effect may be counteracted when eutrophication results in a high load of sedimentation. Tougher species are designed better to withstand a heavy load of sediment. Other perennial red algae with tougher thalli (Chondrus crispus, Furcellaria lumbricalis) also decreased at the exposed sites, but not at the site with most sedimentation, possibly because of lack of competition from D. sanguinea and P. rubens. The abundance of filamentous algae had increased since the early 1960s at all three localities, which may be attributed to eutrophication as well, but no changes were observed in the large perennial brown algae. Decaying loose-lying algae occurred at all localities, but algal mat formation was only abundant at the most sheltered locality. The observed changes in the attached algal vegetation of the exposed sites may be an indication of a general large-scale eutrophication of the Skagerrak.

Density-dependent patterns of thiamine and pigment production in the diatom Nitzschia microcephala

In the present study we investigate how intraspecific (density-dependent) competition for nutrients by the diatom Nitzschia microcephala affects the level of oxidative stress in the algal cells as well as their production of pigments and thiamine. N. microcephala was grown in three different densities until the stationary growth phase was reached. Throughout the experiment, growth rate was negatively related to cell density. Superoxide dismutase activity, protein thiol, and diatoxanthin concentrations indicated increasing oxidative stress with increasing cell density, which was most probably caused by nutrient depletion of the medium. Pigment contents per cell (except for diatoxanthin) decreased with increasing cell density. N. microcephala was able to synthesize thiamine and its thiamine content per cell increased in concert with cell density. In comparison, the dinoflagellate Amphidinium carterae was unable to synthesize thiamine. These results suggest that cells of N. microcephala subjected to higher competition and lower growth rates have a lower carotenoid content and a higher thiamine content. If such responses would occur in nature as well, eutrophication (higher cell densities) may alter the quality of microalgae as food items for higher trophic levels not only by species shifts in the phytoplankton, but also by changes in the cellular nutritional value within species.

Purification and characterisation of a novel starch synthase selective for uridine 5′-diphosphate glucose from the red alga Gracilaria tenuistipitata

Abstract. Red algae (Rhodophyceae) are photosynthetic eukaryotes that accumulate starch granules in the cytosol. Starch synthase activity in crude extracts of Gracilaria tenuistipitata Chang et Xia was almost 9-fold higher with UDP[U-14C]glucose than with ADP[U-14C]glucose. The activity with UDP[U-14C]glucose was sensitive to proteolytic and oxidative inhibition during extraction whilst the activity with ADP[U-14C]glucose appeared unaffected. This indicates the presence of separate starch synthases with different substrate specificities in G. tenuistipitata. The UDPglucose: starch synthase was purified and characterised. The enzyme appears to be a homotetramer with a native Mr of 580u2009kDa and displays kinetic properties similar to other α-glucan synthases such as stimulation by citrate, product (UDP) inhibition and broad primer specificity. We propose that this enzyme is involved in cytosolic starch synthesis in red algae and thus is the first starch synthase described that utilises UDPglucose inu2009vivo. The biochemical implications of the different compartmentalisation of starch synthesis in red algae and green algae/plants are also discussed.

Carbon acquisition mechanisms in Chara tomentosa

Abstract Carbon uptake mechanisms of the stonewort Chara tomentosa from the brackish Baltic Sea were studied by recording changes in pH, alkalinity and inorganic carbon concentrations of the seawater medium during photosynthesis in a closed system. The use of inhibitors identified three mechanisms: (1) a vanadate-sensitive P-type H + -ATPase (proton pump) was involved in carbon uptake. This was previously shown for perfused cells of Chara corallina , but not for living cells. (2) Periplasmic carbonic anhydrase that catalyses the dehydration of HCO 3 − into CO 2 outside the cell membrane was highly active during carbon uptake, also at high pH (>9). (3) At high pH, there was direct uptake of HCO 3 − with the help of an anion exchange protein, which previously has not been shown in Chara . We also document here the occurrence of charasomes in the cell membrane of C. tomentosa , always with mitochondria located in their direct vicinity. The simultaneous high periplasmic carbonic anhydrase and proton pump activities and the occurrence of charasomes suggest proton-pump driven H + extrusion and membrane transport of CO 2 derived from HCO 3 − as the major form of DIC acquisition in this alga. Probably, this occurs in acidic bands in C. tomentosa in which we found a banding pattern of CaCO 3 incrustations (alkaline bands). The results were compared with a similar study on the green alga Cladophora glomerata from the same area, which had very low carbonic anhydrase activity (almost negligible), no structures isolating the periplasm from the bulk seawater medium analogous to charasomes and no CaCO 3 incrustation.