SILICON METABOLISM IN DIATOMS: IMPLICATIONS FOR GROWTH

Diatoms are the world's largest contributors to biosilicification and are one of the predominant contributors to global carbon fixation. Silicon is a major limiting nutrient for diatom growth and hence is a controlling factor in primary productivity. Because our understanding of the cellular metabolism of silicon is limited, we are not fully knowledgeable about intracellular factors that may affect diatom productivity in the oceans. The goal of this review is to present an overview of silicon metabolism in diatoms and to identify areas for future research. Numerous studies have characterized parameters of silicic acid uptake by diatoms, and molecular characterization of transport has begun with the isolation of genes encoding the transporter proteins. Multiple types of silicic acid transporter gene have been identified in a single diatom species, and multiple types appear to be present in all diatom species. The controlled expression and perhaps localization of the transporters in the cell may be factors in the overall regulation of silicic acid uptake. Transport can also be regulated by the rate of silica incorporation into the cell wall, suggesting that an intracellular sensing and control mechanism couples transport with incorporation. Sizable intracellular pools of soluble silicon have been identified in diatoms, at levels well above saturation for silica solubility, yet the mechanism for maintenance of supersaturated levels has not been determined. The mechanism of intracellular transport of silicon is also unknown, but this must be an important part of the silicification process because of the close coupling between silica incorporation and uptake. Although detailed ultrastructural analyses of silica deposition have been reported, we know little about the molecular details of this process. However, proteins occluded within silica that promote silicification in vitro have recently been characterized, and the application of molecular techniques holds the promise of great advances in this area. Cellular energy for silicification and transport comes from aerobic respiration without any direct involvement of photosynthetic energy. As such, diatom silicon metabolism differs from that of other major limiting nutrients such as nitrogen and phosphorous, which are closely linked to photosynthetic metabolism. Cell wall silicification and silicic acid transport are tightly coupled to the cell cycle, which results in a dependency in the extent of silicification on growth rate. Silica dissolution is an important part of diatom cellular silicon metabolism, because dissolution must be prevented in the living cell, and because much of the raw material for mineralization in natural assemblages is supplied by dissolution of dead cells. Perhaps part of the reason for the ecological success of diatoms is due to their use of a silicified cell wall, which has been calculated to impart a substantial energy savings to organisms that have them. However, the growth of diatoms and other siliceous organisms has depleted the oceans of silicon, such that silicon availability is now a major factor in the control of primary productivity. Much new progress in understanding silicon metabolism in diatoms is expected because of the application of molecular approaches and sophisticated analytical techniques. Such insight is likely to lead to a greater understanding of the role of silicon in controlling diatom growth, and hence primary productivity, and of the mechanisms involved in the formation of the intricate silicified structures of the diatom cell wall.     

Enhancing lipid production of the marine diatom Chaetoceros gracilis: synergistic interactions of sodium chloride and silicon

Silicon deficiency is a lipid-promoting stress for many oleaginous diatoms. Literature reports suggest that reduced salinity in seawater, a primary component of which is sodium chloride, may inhibit metabolism of silicon in marine diatoms. We hypothesized that lowering sodium chloride below ocean levels may thus be effective in creating silicon stress and enhancing lipid productivity. We examined the interacting effects of silicon supply (0.05, 0.1, 0.2, and 0.8 mM) and sodium chloride concentration (50, 100, and 400 mM) on growth and lipid production in Chaetoceros gracilis. This was done in batch culture to facilitate the application of severe stress. Low levels of either sodium chloride or silicon resulted in at least 50 % increases in lipid content. The synergy of simultaneous, moderate sodium chloride and silicon stress resulted in lipid content up to 73 % of dry mass and lipid productivity of 1.7 g m^?2 day^?1; with a daily integrated photosynthetic photon flux of 17.3 mol photons m^?2 day^?1, the efficiency of lipid synthesis was thus 0.1 g mol^?1 of photons. Decreased silicon also resulted in a 5 % shift in lipid chain length from C18 to C16 fatty acids. We observed a strong sodium chloride/silicon interaction on total and ash-free dry mass densities that arose because low sodium chloride concentrations were inhibitory to growth, but the inhibition was overcome with excessive silicon supply. This observation suggests that low levels of sodium chloride may have affected metabolism of silicon. The findings of this study can be used to enhance lipid production in oleaginous marine diatoms.     

Bacterial evolution and silicon

This review examines the possible role of silicon in molecular evolution. It is possible silicon participated in early molecular evolution by providing a stable mineral surface or gel structure where the assembly and replication of primitive genetic information occurred. However, as molecular evolution proceeded, silicon was not required in the evolution of C-based organisms. Silicon can be accumulated by diatoms and other living organisms such as silicoflagellates, some xanthophytes, radiolarians and actinopods and plants such as grasses, ferns, horseradish, some trees and flowers, some sponges, insects and invertebrates and bacteria and fungi. Silicon also has a role in synthesis of DNA, DNA polymerase and thymidylate kinase activity in diatoms. It is not unreasonable to examine the role of silicon in early molecular evolution as it may have been part of a micro-environment in which assembly of genetic information occurred.     

Kinetics of β-N-methylamino-L-alanine (BMAA) and 2, 4-diaminobutyric acid (DAB) production by diatoms: the effect of nitrogen

The neurotoxins β-N-methylamino-L-alanine (BMAA) and 2,4-diaminobutyric acid (DAB) are produced by cyanobacteria, diatoms and dinoflagellates and have been detected in seafood worldwide. Our present knowledge of their metabolism or biosynthesis is limited. In this study, the production of BMAA and DAB as a function of time was monitored in five strains representing four species of diatoms, i.e. Phaeodactylum tricornutum, Thalassiosira weissflogii, Thalassiosira pseudonana and Navicula pelliculosa, previously identified as BMAA and DAB producers. Subsequently, three strains were selected and exposed to three nitrogen treatments – starvation, control (the standard concentration in f/2 medium) and enrichment, because BMAA metabolism has been suggested to be closely associated with cellular nitrogen metabolism in both cyanobacteria and diatoms. Chlorophyll a and total protein concentrations were also determined. Our results indicate that BMAA and DAB production in diatoms is species- and strain-specific. However, production might also be affected by stress, particularly as related to nitrogen starvation and cell density. Furthermore, this study shows a significant correlation between the production of the two neurotoxins which might further suggest common steps in the metabolic pathways.     

Metabolism of mutagenic polycyclic aromatic hydrocarbons by photosynthetic algal species

Polycyclic aromatic hydrocarbons (PAH) known to produce carcinogenic and mutagenic effects have been shown to contaminate waters, sediments and soils. While it is accepted that metabolites of these compounds are responsible for most of their biological effects in mammals, their metabolism, and to a large extent their bioactivity, in aquatic plants have not been explored. Cultures of photosynthetic algal species were assayed for their ability to metabolize benzo[a]pyrene (BaP), a carcinogenic PAH under conditions which either permitted (white light) or disallowed (gold light) photooxidation of the compound. Growth of Selenastrum capricornutum, a fresh-water green alga, was completely inhibited when incubated in white light with 160 micrograms BaP/l medium. By contrast concentrations at the upper limit of BaP solubility in aqueous medium had no effect on algal growth when gold light was used. BaP quinones and phenol derivatives were found to inhibit growth of Selenastrum under white light incubation. BaP phototoxicity and metabolism were observed to be species-specific. All 3 tested species of the order Chlorococcales were growth-inhibited by BaP in white light whereas neither the green alga Chlamydomonas reinhardtii nor a blue-green, a yellow-green or an euglenoid alga responded in this fashion. Assays of radiolabeled BaP metabolism in Selenastrum showed that the majority of radioactivity associated with BaP was found in media as opposed to algal cell pellets, that the extent of metabolism was BaP concentration dependent, and that the proportion of various metabolites detected was a function of the light source. After gold light incubation, BaP diols predominated while after white light treatment at equal BaP concentrations, the 3,6-quinone was found in the highest concentration. Extracted material from algal cell pellets and from media was tested for mutagenicity in a forward mutation suspension assay in Salmonella typhimurium using resistance to 8-azaguanine for selection. Direct-acting mutagens were detected in extracted media from incubation of Selenastrum with 400 micrograms BaP/l for 1 day in gold light. Extracts of media from algae incubated in gold light from 1 to 4 days with 1200 micrograms BaP/l were found to have direct-acting mutagens as well as those requiring further metabolism. Media extracts from white light incubations of BaP were mutagenic upon addition of rat liver homogenates. Activity of these materials from white light treatment are largely attributable to unmetabolized BaP.     

Review on proteomic analyses of benzo[a]pyrene toxicity

Benzo[a]pyrene (BaP), a five-ring polycyclic aromatic hydrocarbon, is a well-recognized environmental pollutant. Coal-processing waste products, petroleum sludge, asphalt, creosote, and tobacco smoke, all contain high levels of BaP. Exposure to BaP elicits many adverse biological effects, including tumor formation, immunosuppression, teratogenicity, and hormonal effects. In addition to the genetic damage caused by BaP exposure, several studies have indicated the disruption of protein-protein signaling pathways. However, contrary to the large number of studies on BaP-induced DNA damage, only few data have been gathered on its effects at the protein level. This review highlights all proteomic studies to date used for assessing the toxicity of BaP and its metabolites in various organ systems. It will also give an overview on the role proteomics may play to elucidate the mechanisms underlying BaP toxicity.     

Bioremediation of high molecular weight polycyclic aromatic hydrocarbons: a review of the microbial degradation of benzo[a]pyrene

Over the past 30 years, research on the microbial degradation of polycyclic aromatic hydrocarbons (PAHs) has resulted in the isolation of numerous genera of bacteria, fungi and algae capable of degrading low molecular weight PAHs (compounds containing three or less fused benzene rings). High molecular weight PAHs (compounds containing four or more fused benzene rings) are generally recalcitrant to microbial attack, although some fungi and algae are capable of transforming these compounds. Until recently, only a few genera of bacteria have been isolated with the ability to utilise four-ring PAHs as sole carbon and energy sources while cometabolism of five-ring compounds has been reported. The focuss of this review is on the high molecular weight PAH benzo[a]pyrene (BaP). There is concern about the presence of BaP in the environment because of its carcinogenicity, teratogenicity and toxicity. BaP has been observed to accumulate in marine organisms and plants which could indirectly cause human exposure through food consumption. This review provides an outline of the occurrence of BaP in the environment and the ability of bacteria, fungi and algae to degrade the compound, including pathways for BaP degradation by these organisms. In addition, approaches for improving microbial degradation of BaP are discussed.     

Poly(ADP-Ribose) Glycohydrolase (PARG) Silencing Suppresses Benzo(a)pyrene Induced Cell Transformation

Benzo(a)pyrene (BaP) is a ubiquitously distributed environmental pollutant and known carcinogen, which can induce malignant transformation in rodent and human cells. Poly(ADP-ribose) glycohydrolase (PARG), the primary enzyme that catalyzes the degradation of poly(ADP-ribose) (PAR), has been known to play an important role in regulating DNA damage repair and maintaining genomic stability. Although PARG has been shown to be a downstream effector of BaP, the role of PARG in BaP induced carcinogenesis remains unclear. In this study, we used the PARG-deficient human bronchial epithelial cell line (shPARG) as a model to examine how PARG contributed to the carcinogenesis induced by chronic BaP exposure under various concentrations (0, 10, 20 and 40 μM). Our results showed that PARG silencing dramatically reduced DNA damages, chromosome abnormalities, and micronuclei formations in the PARG-deficient human bronchial epithelial cells compared to the control cells (16HBE cells). Meanwhile, the wound healing assay showed that PARG silencing significantly inhibited BaP-induced cell migration. Furthermore, silencing of PARG significantly reduced the volume and weight of tumors in Balb/c nude mice injected with BaP induced transformed human bronchial epithelial cells. This was the first study that reported evidences to support an oncogenic role of PARG in BaP induced carcinogenesis, which provided a new perspective for our understanding in BaP exposure induced cancer.     

Silicon uptake and metabolism of the marine diatom Thalassiosira pseudonana: Solid-state 29Si NMR and fluorescence microscopic studies

Uptake and metabolism of silicon by diatoms are studied by the combined use of solid-state 29Si NMR spectroscopy and confocal laser fluorescence microscopy especially with respect to the presence and nature of an intracellular silicon-storage pool. Cells of the marine diatom Thalassiosira pseudonana were synchronized by silicon starvation and frozen without any freeze-drying or chemical treatment in order to analyze integer and unmodified diatoms. The frozen samples were investigated by solid-state 29Si NMR spectroscopy to identify potential silica precursors. The developmental state of the cell culture and the formation of new siliceous girdle bands and valves were monitored by laser fluorescence microscopic studies. A comparison of fluorescence microscopic and NMR data allows the assignment of NMR spectra to the various developmental stages of the dividing diatom cells. A detailed analysis of solid-state 29Si NMR spectra suggests that the silicon-storage pool-if present-consists of four-coordinated, condensed silicon; possibly a silica sol.     

The use of diatoms in monitoring the development of created wetlands at a sandmining site in Western Australia

Former sand-mining pits at Capel, 200 km south of Perth in Western Australia, have been rehabilitated into artificial wetlands since 1975–1979. A chain of fresh water lakes was created as a potential waterbird refuge and an area for passive recreation. Initially, the lakes had low pH, high ammonium, iron and manganese levels and low phosphorus concentration. The lakes were characterised by low diversity of diatoms dominated by acidophilous species. Following an increase in pH in the effluent water discharged into the lake from the mining process plant and landscaping of the lakes since 1988, the diversity of diatoms gradually increased. The system is now dominated by periphytic diatom communities, preferring high conductivity. There has been a marked transition in the diatom community from acidophilous to alkaliphilous species. Planktonic diatom blooms replaced dinoflagellate blooms. Concomitantly, there has been a dramatic increase in the diversity of invertebrates and waterbirds in these lakes. The value of diatoms in assessing the progressive development of created wetlands as self-sustaining ecosystems at sand mines in Australia is discussed.     

Coexposure to benzo[a]pyrene plus UVA induced DNA double strand breaks: visualization of Ku assembly in the nucleus having DNA lesions

Benzo[a]pyrene (BaP) is a ubiquitous environmental pollutant with potential carcinogenicity. It has been shown that BaP, upon UVA irradiation, synergistically induced oxidative DNA damage, but other DNA damage was not confirmed. In this study, we examined whether coexposure to BaP plus UVA induces double strand breaks (DSBs) using xrs-5 cells, deficient in the repair of DSBs (Ku80 mutant), and whether Ku translocates involving the formation of DSBs. BaP plus UVA had a significant cytotoxic effect on CHO-K1 cells and an even more drastic effect on Ku80-deficient, xrs-5 cells, suggesting that the DSBs were generated by coexposure to BaP plus UVA. The DSBs were repaired in CHO-K1 cells within 30 min, but not in xrs-5 cells, indicating the involvement of a non-homologous end joining, which needs Ku proteins. Furthermore, we succeeded in visualizing that Ku80 rapidly assembled to the exposed region, in which DSBs might be generated, and clarified that the presence of both Ku70 and Ku80 was important for their accumulation.     

聚球藻硅质化作用初探

硅元素是全球生地化循环的重要组成成分之一,对海洋生态系统中以浮游植物主导的初级生产力和硅碳循环具有重要的意义。普遍认为硅藻主导着全球海洋的硅循环,成为海洋硅循环和碳循环交互作用的重要桥梁。海洋单细胞聚球藻对海洋食物网和能量流具有关键启动和支撑作用,是全球碳循环中固碳过程的主要贡献者,近年又被发现其具有重要的硅质化作用,为我们提供了一个在海洋中(特别是寡营养海域),除硅藻之外,连接硅碳循环交互作用的新视角,对硅藻在全球海洋硅碳循环的绝对地位具有重要的挑战意义。面对聚球藻在大洋中如此巨大的生物量,甚至高于硅藻,有必要弄清楚其碳沉降机制以及准确的模拟其硅循环,然而关于其在海洋硅循环的研究极少,硅质化作用的吸收和储存机理以及环境调节机制也不清楚;另外,其对世界海洋硅碳循环的调节作用也未见报道。为此,通过前人对海洋单细胞聚球藻硅质化作用研究的基础上进行有针对性的探讨,可望对海洋单细胞聚球藻硅质化作用及其对硅碳循环的调控机制有一个基本的认识,为深入研究聚球藻在全球海洋硅循环中的作用提供基础。     

The application of oligonucleotide probes and microarrays for the identification of freshwater diatoms

Diatoms are major contributors to global carbon fixation and constitute a significant portion of biofilms found in lotic ecosystems. Despite their widespread abundance and the fact that extensive studies have been performed on morphological features of frustules, molecular tools for the identification of diatoms are not commonly available. This study focuses on the development of oligonucleotide probes for the detection of diatom species relevant to water quality assessment. The selected panel of diatoms covers all the species found in water of varying quality from the rivers of central-East Apennine (Italy). Small subunit rRNA-targeted probes were applied to a microarray platform as well as to a new technique termed Primer–Probe, with the aim of obtaining a molecular tool suitable for accurate identification of both single and mixed species diatom populations. The Primer–Probe technique together with dot-blot assays proved to be ideal for the preliminary screening of a large set of DNA oligonucleotides designed by ARB software. It was shown that microarrays, as a promising technology for rapid and simultaneous detection of a wide range of species-specific genetic markers, can be adapted to monitor changes within a diatom community. It is suggested that microarrays will provide a molecular basis for microbial identification to support standard microscopy techniques used by ecologists and environmental scientists for monitoring water quality.     

硅提高植物抗旱性的生理机制研究进展

干旱作为限制作物产量和品质的主要非生物胁迫之一,对全球社会、经济和生态造成巨大损失。在全球气候变化背景下,提高植物抗旱性的重要性日益突显。硅能够提高植物的抗旱性：外源硅的施用可以影响气孔导度,改变蒸腾速率,改善植物水分状况;通过调节气孔动力学、合成光合色素,促进光化学反应,从而改善光合作用;此外硅可通过渗透调节以平衡植物对矿质元素的吸收,以及调节抗氧化防御系统,减轻植物在干旱胁迫中的氧化损伤。总结了硅对干旱胁迫下植物水分利用、光合作用、矿质元素吸收、抗氧化系统、植物激素代谢等方面的作用及相关生理机制。建议未来从复合逆境胁迫、低硅积累植物等方面进一步揭示硅提高植物抗旱性的作用机制,从而为农林生态系统合理利用硅素来提高生产效率提供科学依据和理论基础。     

The Probable Mechanism for Silicon Capture by Diatom Algae: Assimilation of Polycarbonic Acids with Diatoms—Is Endocytosis a Key Stage in Building of Siliceous Frustules?

Many organisms including unicellular (diatoms, radiolaria, and chrysophytes), higher plants (rice and horsetail) and animals (sponges) use silica as a main part of skeletons. The bioavailable form of silicon is silicic acid and the mechanism of silicic acid penetration into living cells is still an enigma. Macropinocytosis was assumed as a key stage of the silicon capture by diatoms but assimilation of monomeric silicic acid by this way requires enormous amounts of water to be passed through the cell. We hypothesized that silicon can be captured by diatoms via endocytosis in the form of partially condensed silicic acid (oligosilicates) whose formation on the diatom surface was supposed. Oligosilicates are negatively charged nanoparticles and similar to coils of poly(acrylic acid) (PAA). We have synthesized fluorescent tagged PAA as well as several neutral and positively charged polymers. Cultivation of the diatom Ulnaria ferefusiformis in the presence of these polymers showed that only PAA is able to penetrate into siliceous frustules. The presence of PAA in the frustules was confirmed with chromatography and PAA causes various aberrations of the valve morphology. Growth of U. ferefusiformis and two other diatoms in the presence of tri- and tetracarbonic fluorescent tagged acids points to the ability of diatoms to recognize substances that bear four acidic groups and to include them into siliceous frustules. Thus, partial condensation of silicic acid is a plausible first stage of silicon assimilation.     

苯并[α]芘对不同修复潜力羊茅属植物的根系分泌物中几种低分子量有机物的影响

根系分泌物是植物与土壤间进行物质交换和信息传递的重要载体, 是植物响应外界胁迫的重要途径, 也是构成根际微生态特征的关键因素。根系分泌物与有机污染物的植物修复密切相关, 研究胁迫条件下不同修复潜力植物间根系分泌物的释放特征有助于揭示植物修复的内在机制。该文借助根际袋土培试验研究了苯并[α]芘(BaP)胁迫下5种羊茅属(Festuca)植物根系不同生长期(30-70天)几种低分子量有机物的分泌特征。结果表明: 1) BaP浓度在10.25-161.74 mg·kg^-1范围内时, 待试植物能有效地促进土壤中BaP的去除, 其修复潜力依次为苇状羊茅(F. arundinacea) > 草原羊茅(F. chelungkiangnica) ≥ 毛稃羊茅(F. rubra subsp. arctica) ≥ 贫芒羊茅(F. sinomutica) > 细芒羊茅(F. stapfii)。2) BaP胁迫增强了植物根系对可溶性糖的分泌: 随着胁迫强度的增大、胁迫期的延长, 其分泌量变化呈“先升后降”趋势。3) BaP胁迫促进了植物根系低分子量有机酸的释放, 植物的修复潜力越大, 有机酸高峰值出现时的胁迫浓度越高; 组成成分较稳定, 草酸、乙酸、乳酸和苹果酸为主要组分(>97.34%), 在修复潜力较强植物的根系分泌物中检测出微量的反丁烯二酸。4) BaP胁迫对氨基酸种类影响不大, 但对分泌量影响较大。其中, 苏氨酸、丝氨酸、甘氨酸、丙氨酸的分泌量随BaP胁迫强度的增强而剧增; 脯氨酸、羟脯氨酸和天冬氨酸近乎以加和效应甚至协同效应的形式参与植物对BaP胁迫的应激反应: 参与应激组分的分泌量随胁迫强度的增强而剧增, 植物的修复潜力越强, 参与的组分越多。可见BaP胁迫下, 5种羊茅属植物根系分泌物中几种低分子量有机物的释放特征与植物自身的修复潜力有关: 修复潜力越强, 释放量越多且成分也越复杂, 并呈现出较强的环境适应性及生理可塑性。     

硅对干旱胁迫下玉米水分代谢的影响

利用盆栽试验研究了施硅(K2SiO3)对玉米植株水分代谢的影响.结果表明:施硅降低了干旱胁迫下玉米植株的气孔导度,降低了干旱胁迫早期到中期的蒸腾速率,保持了干旱胁迫后期较高的蒸腾速率,从而导致施硅玉米植株的叶片含水量和水势高于对照.由于植株的水分状况改善,施硅玉米植株生物量高于对照.硅增强玉米植株的抗旱性,而提高植株保水能力是硅提高抗旱性的重要原因.