CARBONIC ANHYDRASE IN THE MARINE DIATOM THALASSIOSIRA WEISSFLOGII (BACILLARIOPHYCEAE)1

The zinc metalloenzyme carbonic anhydrase plays a critical role in inorganic carbon acquisition in marine diatoms, thus conferring on zinc a key role in oceanic carbon cycling. As a first step in determining the location and function of carbonic anhydrase (CA) in Bacillariophyceae, we purified and partially sequenced CA from T. weissflogii (Gru) Fryxell et Hasle (TWCA1) and cloned the corresponding cDNA (twca1). The twca1 sequence is different from other known algal carbonic anhydrase genes, and encodes a protein of roughly 34 kDa. The amino terminal amino acids sequenced from purified TWCA1 are 72 residues downstream of the putative starting methionine predicted by twca1. This difference may be due to the presence of a short-lived signal sequence designed to guide the enzyme to the correct cellular location. The absence of any homology between TWCA1 and previously sequenced CAs from Chlorophyceae may indicate either convergent evolution or that carbon acquisition represents a fundamental physiological difference among algal phyla.     

PROTEINACEOUS AND IMMUNOCHEMICAL DISTINCTIONS BETWEEN THE OVAL AND FUSIFORM MORPHOTYPES OF PHAEODACTYLUM TRICORNUTUM (BACILLARIOPHYCEAE)1

Two morphotypes, fusiform and oval, were isolated from a single clone of the diatom Phaeodactylum tricornutum Bohlin and maintained as subclones by culturing in liquid and solid substrates, respectively. Salinity of the medium, from brackish to marine, had no effect on expression of the phenotypes. The oval cell is generated endogenously within a “transformed”fusiform cell upon transfer from liquid medium to agar plates. With the light microscope, normal and “transformed”fusiform cells, prior to giving rise to oval cells, can be discriminated by means of their staining response to toluidine blue. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of protein extracts from lysed cells revealed slight differences in polypeptide composition between fusiform and oval types. A phenotype-restoration experiment from oval to fusiform demonstrated that the oval type readily reestablished not only fusiform morphology but also the protein pattern characteristic for the fusiform type. Immunochemical analyses (western blots) using antisera raised against whole and lysed cells of both morphotypes revealed antigenic alterations of the oval morphotype. Several antigenic determinants restricted mainly to the surface of oval cells were detected. Results indicate that environmentally induced phenotypes of Phaeodactylum may be not only the consequence of specific gene expression but also the result of significant, general post-translational modifications.     

LOCATION AND EXPRESSION OF FRUSTULINS IN THE PENNATE DIATOMS CYLINDROTHECA FUSIFORMIS, NAVICULA PELLICULOSA, AND NAVICULA SALINARUM (BACILLARIOPHYCEAE)

A detailed immunocytochemical and biochemical study of the location and expression of frustulins, a family of proteins associated with the frustules of diatoms, has been performed for Cylindrotheca fusiformis Reimann et Lewin, Navicula pelliculosa (Brébisson et Kützing) Hilse, and Navicula salinarum (Grunow) Husted. Immunocytochemistry revealed that frustulins, which share homologous epitopes but are different in size, were predominantly located in the organic casing. Based on timed immunolocalization experiments and Western blotting analysis of cell extracts obtained sequentially after repleting silicate to Si-synchronized cells, the continuous presence of the frustulins in the mature and parental organic casing of the examined species was observed. The frustulins of N. pelliculosa appeared as proteins similar to those of C. fusiformis, sharing identical epitopes. The extractions, however, yielded a markedly lower abundance of frustulins in N. pelliculosa. Peak concentrations of extracted frustulins appeared to be expressed just ahead of the silicification process in C. fusiformis, whereas the level of expression in N. pelliculosa increased along with maturation of the new valves. For N. salinarum, the presence of the frustulins could not be confirmed properly by Western blotting, most probably because of the small sample volumes, inefficient extraction, and a lower amount of homologous frustulins in the casing of this species. It is concluded that the frustulins of these species are not associated with the silicalemma of the newly formed silica deposition vesicles and therefore do not seem to be involved in the silicification process itself. Overall, the results imply a structural role of the frustulins in the casing of diatoms rather than a regulation of the silicification process.     

Absolute configuration of heteroxanthin and diadinoxanthin

The absolute configurations of heteroxanthin ((3S,5S,6S,3′R)- 7′,8′-didehydro-5,6-dihydro-β,β-carotene-3,5,3′,6′-tetrol) ex Euglena gracilis and of diadinoxanthin ((3S,5R,6S,3′R)-5,6-epoxy-7′,8′-didehydro-5,6-dihydro-β,β-carotene-3,3′-diol) from the same source have been established by chemical reactions, hydrogen bonding studies, ¹H NMR and CD. Two previously unknown carotenoids (artefacts?) from Trollius europaeus, assigned the structures (3S,5S,6S,3′S,5′R,6′R)-6,7-didehydro-5,6,5′,6′-tetrahydro-β,β -carotene-3,5,6,3′,5′-pentol and its 5R epimer, served as useful models.     

SEXUAL REPRODUCTION IN STEPHANODISCUS NIAGARAE (BACILLARIOPHYTA)1

We observed sexual reproduction in a clonal culture of Stephanodiscus niagarae Ehrenb. and used light and scanning electron microscopy to absent flagellated male cells, auxospore growth, initial valve structure and production, and subsequent daughter cell division. Free auxospores were spherical and nonsiliceous throughout growth, producing hemispherical initial valves devoid of spines and with nonfasciculate striae. Pregametangial cells averaged 43% of the diameter of the daughter cell population and were 1/9 the biovolume of initial, cells. This paper is the first confirmed report of sexual reproduction in S. niagarae, although it appears that specimens of Actinocyclus niagarae H. L. Smith, described from Lake Erie in 1878, are actually initial valves of S. niagarae.     

EFFECTS OF SILICON DEFICIENCY ON LIPID COMPOSITION AND METABOLISM IN THE DIATOM CYCLOTELLA CRYPTICA1

The effects of silicon deficiency on the metabolism and composition of lipids in Cyclotella cryptica T13L Reimann, Lewin, and Guillard were examined. Silicon-deficient cells had higher levels of neutral lipids (primarily triacylglycerols) and higher proportions of saturated and monounsaturated fatty acids than silicon-replete cells. After 4 h of silicon deficiency, the percentage of newly assimilated NaH¹⁴CO₃ partitioned into lipids increased from 27.6% to 54.1%, whereas the percentage partitioned into chrysolaminarin decreased from 21.6% to 10.6%. In addition, pulse-chase experiments with NaH¹⁴CO₃ indicated that the amount of ¹⁴C in the total cellular lipid fraction increased by 32% after 12 h of silicon deficiency despite the absence of additional photoassimilable ¹⁴C. Therefore, the accumulation of lipids in response to silicon deficiency appears to be due to two distinct processes: (a) an increase in the proportion of newly assimilated carbon partioned into lipids, and (2) a slow conversion of previously assimilated carbon from non-lipid compounds into lipids     

EFFECT OF SILICATE LIMITATION ON VALVE MORPHOLOGY IN THALASSIOSIRA AND COSCINODISCUS (BACILLARIOPHYCEAE)1

Coscinodiscus radiatus Ehrenb. and Thalassiosira eccentrica (Ehrenb.) Cleve were grown in a silicate-limited chemostat at silicate concentrations below 1 μg-atoms · l^?1. The resulting abnormal valves of C. radiatus lacked a thickened ring around the foramina; their pore membranes were thinner and their loculi shallower than those in normal cells. Abnormal valves of T. eccentrica had a fasciculate areolae pattern; they lacked a silica covering over the foramina and some tangential areolae walls. Neither abnormal valve could be termed a new species.     

Observations on Aulacoseira nipponica from Lake Biwa, Japan, and Aulacoseira solida from North America (Bacillariophyceae)

A recent diatom, Aulacoseira nipponica (Skvortzow) Tuji comb. et stat. nov., is described from Lake Biwa, Japan, where they had been identified previously as Aulacoseira solida (Eulenstein) Krammer. These forms are compared with populations of A. solida from North America. The Japanese species differs from the North American specimens in characteristics related to the density of striae and form of the rimoportulae.     

PELAGIC AND BENTHIC ENVIRONMENTAL CONTROLS ON THE SPATIAL DISTRIBUTION OF A VIABLE DIATOM PROPAGULE BANK ON THE SWEDISH WEST COAST1

The distribution of viable diatom resting stages in sediments on the Swedish west coast was assessed by the most probable number (MPN) culture technique. Multivariate analyses correlated benthic and pelagic environmental factors to the observed spatial variations in the size and taxonomic composition of the propagule bank. Viable diatom resting stages were plentiful (0.2–4.8 million cells·g ^{? 1} 1 Received 11 September 2001. Accepted 12 June 2002.
dry weight) and were dominated by the genera Skeletonema, Detonula, Chaetoceros, and Thalassiosira. Size of the propagule bank was primarily related to planktonic biomass (measured as chl a) and was highest in the Orust‐Tjörn fjord system. Species composition in this fjord system was dominated by D. confervacea (Cleve) Gran and T. nordenskioeldii Cleve in contrast to stations on the outer coast, which contained more cells of T. minima Gaarder, Asterionellopsis glacialis (Castracane) Round, and Leptocylindrus danicus Cleve. These taxonomic variations were principally influenced by deep water oxygen concentrations and water column stability. Benthic resting cells of S. costatum (Greville) Cleve were abundant all along the coast but showed reduced viability in low oxygen environments. Calculations based on MPN values estimated that resuspension of sediment could provide a sizable inoculum to the plankton, although the development of planktonic blooms will also depend on forces of hydrography and weather. Although benthic resting stages may not be absolutely necessary for survival of all diatoms, these cells may be important in determining species cycles, succession, and the spatial distribution of diatoms.