Diatoms are unicellular algae with plastids acquired by secondary endosymbiosis. They are responsible for approximately 20% of global carbon fixation. We report the 34 million-base pair draft nuclear genome of the marine diatom Thalassiosira pseudonana and its 129 thousand-base pair plastid and 44 thousand-base pair mitochondrial genomes. Sequence and optical restriction mapping revealed 24 diploid nuclear chromosomes. We identified novel genes for silicic acid transport and formation of silica-based cell walls, high-affinity iron uptake, biosynthetic enzymes for several types of polyunsaturated fatty acids, use of a range of nitrogenous compounds, and a complete urea cycle, all attributes that allow diatoms to prosper in aquatic environments.

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Diatom cell walls are regarded as a paradigm for controlled production of nanostructured silica, but the mechanisms allowing biosilicification to proceed at ambient temperature at high rates have remained enigmatic. A set of polycationic peptides (called silaffins) isolated from diatom cell walls were shown to generate networks of silica nanospheres within seconds when added to a solution of silicic acid. Silaffins contain covalently modified lysine-lysine elements. The first lysine bears a polyamine consisting of 6 to 11 repeats of the N-methyl-propylamine unit. The second lysine was identified as epsilon-N,N-dimethyl-lysine. These modifications drastically influence the silica-precipitating activity of silaffins.

Silaffins are uniquely modified peptides that have been implicated in the biogenesis of diatom biosilica. A method that avoids the harsh anhydrous hydrogen fluoride treatment commonly used to dissolve biosilica allows the extraction of silaffins in their native state. The native silaffins carry further posttranslational modifications in addition to their polyamine moieties. Each serine residue was phosphorylated, and this high level of phosphorylation is essential for biological activity. The zwitterionic structure of native silaffins enables the formation of supramolecular assemblies. Time-resolved analysis of silica morphogenesis in vitro detected a plastic silaffin-silica phase, which may represent a building material for diatom biosilica.

Biomineralizing organisms use organic molecules to generate species-specific mineral patterns. Here, we describe the chemical structure of long-chain polyamines (up to 20 repeated units), which represent the main organic constituent of diatom biosilica. These substances are the longest polyamine chains found in nature and induce rapid silica precipitation from a silicic acid solution. Each diatom is equipped with a species-specific set of polyamines and silica-precipitating proteins, which are termed silaffins. Different morphologies of precipitating silica can be generated by polyamines of different chain lengths as well as by a synergistic action of long-chain polyamines and silaffins.T he biological formation of inorganic structures, termed biominerals, is a widespread phenomenon in nature (1). Among the most famous examples are unicellular algaediatoms-that possess a cell wall composed of silica and organic molecules or macromolecules (2). Most interestingly, diatom biosilica displays a dazzling variety of species-specific silica patterns that are structured on a nanometer-to-micrometer scale (3-6). Elucidating the mechanism that controls production of nanostructured biosilica is a fascinating biochemical problem and, in addition, is of great interest in materials chemistry. Biomimetic approaches are believed to allow the production of advanced materials at ambient temperature and with high precision, which are expected to exhibit superior properties in a wide range of applications (7,8). Reaching this goal, however, requires a detailed knowledge of the organic molecules that govern silica biomineralization.Recently, cationic polypeptides (called silaffins) isolated from purified cell walls of the diatom Cylindrotheca fusiformis were shown to generate networks of silica nanospheres within seconds when added to a solution of silicic acid (9). The silaffins are tightly associated with the biosilica so that they can only be solubilized after dissolution of the cell wall in hydrogen fluoride (HF). Silaffin-1 contains a previously undescribed type of protein modification, a polyamine consisting of 6-11 repeats of the N-methyl-propylamine unit (a mass increment of 71 Da per repeat) covalently attached to specific lysine residues (9). It is probably this structural element that accelerates silicic acid polymerization and promotes production of nanosphere networks. Previously, ultrastructural work on cell wall biogenesis in a number of diatoms led to the conclusion that silica appears to be deposited in different forms during valve morphogenesis. Especially evident was the participation of silica spheres ranging up to 100 nm in diameter (4, 10). If silaffins are indeed involved in the process of pattern formation that creates the amazingly rich variety of diatom shell silica structures, then different diatom species are expected to contain different types of silaffins or silaffin-like molecules. We therefore analyzed the HFextractable organic cell wall components from a wide range of diatom species. Surprisingly, th...

Two silica-precipitating peptides, silaffin-1A 1 and-1A 2 , both encoded by the sil1 gene from the diatom Cylindrotheca fusiformis, were extracted from cell walls and purified to homogeneity. The chemical structures were determined by protein chemical methods combined with mass spectrometry. Silaffin-1A 1 and -1A 2 consist of 15 and 18 amino acid residues, respectively. Each peptide contains a total of four lysine residues, which are all found to be post-translationally modified. In silaffin-1A 2 the lysine residues are clustered in two pairs in which the ⑀-amino group of the first residue is linked to a linear polyamine consisting of 5 to 11 N-methylated propylamine units, whereas the second lysine is converted to ⑀-N,N-dimethyllysine. Silaffin-1A 1 contains only a single lysine pair exhibiting the same structural features.

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For almost 200 years scientists have been fascinated by the ornate cell walls of the diatoms. These structures are made of amorphous silica, exhibiting species-specific, mostly porous patterns in the nano-to micrometer range. Recently, from the diatom Cylindrotheca fusiformis unusual phosphoproteins (termed silaffins) and long chain polyamines have been identified and implicated in biosilica formation. However, analysis of the role of silaffins in morphogenesis of species-specific silica structures has so far been hampered by the difficulty of obtaining structural data from these extremely complex proteins. In the present study, the five major silaffins from the diatom Thalassiosira pseudonana (tpSil1H, -1L, -2H, -2L, and -3) have been isolated, functionally analyzed, and structurally characterized, making use of the recently available genome data from this organism. Surprisingly, the silaffins of T. pseudonana and C. fusiformis share no sequence homology but are similar regarding amino acid composition and posttranslational modifications. Silaffins tpSil1H and -2H are higher molecular mass isoforms of tpSil1L and -2L, respectively, generated in vivo by alternative processing of the same precursor polypeptides. Interestingly, only tpSil1H and -2H but not tpSil1L and -2L induce the formation of porous silica patterns in vitro, suggesting that the alternative processing event is an important step in morphogenesis of T. pseudonana biosilica.During evolution many organisms (e.g. diatoms, sponges, radiolaria) have acquired the ability to use the ubiquitous monosilicic acid Si(OH) 4 for the formation of species specifically structured, silica-based exo-or endoskeletons (1). This interesting biomineralization phenomenon is mediated by cellular organic (macro-) molecules that accelerate silicic acid polycondensation and control morphogenesis of the forming silica (2). Diatoms are an extremely large group (Ͼ10,000 species) of unicellular eukaryotic algae that play a major role in biological silica cycling. Within the last few years diatom biosilica-associated proteins (termed silaffins) and long chain polyamines (LCPA) 1 have been identified and hypothesized to represent key components of the diatom biosilica-forming machinery. Silaffins and LCPA exhibit the remarkable ability to induce rapid silica deposition in vitro and to control the nanostructure of the forming silica (3). Therefore, unraveling the correlations between chemical structures, physical properties, and silica-forming activities of silaffins and LCPA will be important for understanding the molecular mechanism of species-specific biosilica nanopatterning. So far, silaffins have only been characterized from the diatom Cylindrotheca fusiformis. They are highly modified proteins/peptides rich in hydroxyamino acids (serine, threonine, hydroxyproline) and lysine residues. Silaffins natSil1A and -1B are O-phosphorylated at numerous sites and contain polyamine-modified lysine residues, features that enable these peptides to rapidly form silica nanospheres in vitr...