Annotated Genome Sequence of the High-Biomass-Producing Yellow-Green Alga Tribonema minus.

Here, we report the annotated genome sequence for a heterokont alga from the class Xanthophyceae. This high-biomass-producing strain, Tribonema minus UTEX B 3156, was isolated from a wastewater treatment plant in California. It is stable in outdoor raceway ponds and is a promising industrial feedstock for biofuels and bioproducts.

Characterization of a novel strain of Tribonema minus demonstrating high biomass productivity in outdoor raceway ponds.

Photosynthetic algae represent a large, diverse bioresource potential. Yellow-green algae of the genus Tribonema are candidates for production of biofuels and other bioproducts. We report on a filamentous isolate from an outdoor raceway polyculture growing on municipal reclaimed wastewater which we classified as T. minus. Over one year of cultivation in 3.5 m2 raceway ponds fed by reclaimed municipal wastewater, T. minus cultures were more productive than the native algal polycultures, with annual average productivities of 15.9 ± 0.3 and 13.4 ± 0.4 g/m2/day, respectively. The biochemical composition of T. minus biomass grown outdoors was constant year-round, with 28.3 ± 0.4% carbohydrates, 37.6 ± 0.7% proteins, and 6.1 ± 0.3% fatty acids (measured as methyl esters), with up to 4.0% of the valuable omega-3 eicosapentaenoic acid, on an ash-free dry-weight basis. In summary, T. minus was more productive, easier to harvest and produced higher quality biomass than the native polycultures.

Reconstruction and analysis of a carbon-core metabolic network for Dunaliella salina.

BACKGROUND: The green microalga Dunaliella salina accumulates a high proportion of ß-carotene during abiotic stress conditions. To better understand the intracellular flux distribution leading to carotenoid accumulation, this work aimed at reconstructing a carbon core metabolic network for D. salina CCAP 19/18 based on the recently published nuclear genome and its validation with experimental observations and literature data. RESULTS: The reconstruction resulted in a network model with 221 reactions and 212 metabolites within three compartments: cytosol, chloroplast and mitochondrion. The network was implemented in the MATLAB toolbox CellNetAnalyzer and checked for feasibility. Furthermore, a flux balance analysis was carried out for different light and nutrient uptake rates. The comparison of the experimental knowledge with the model prediction revealed that the results of the stoichiometric network analysis are plausible and in good agreement with the observed behavior. Accordingly, our model provides an excellent tool for investigating the carbon core metabolism of D. salina. CONCLUSIONS: The reconstructed metabolic network of D. salina presented in this work is able to predict the biological behavior under light and nutrient stress and will lead to an improved process understanding for the optimized production of high-value products in microalgae.

Carbon partitioning in green algae (chlorophyta) and the enolase enzyme.

The exact mechanisms underlying the distribution of fixed carbon within photoautotrophic cells, also referred to as carbon partitioning, and the subcellular localization of many enzymes involved in carbon metabolism are still unknown. In contrast to the majority of investigated green algae, higher plants have multiple isoforms of the glycolytic enolase enzyme, which are differentially regulated in higher plants. Here we report on the number of gene copies coding for the enolase in several genomes of species spanning the major classes of green algae. Our genomic analysis of several green algae revealed the presence of only one gene coding for a glycolytic enolase [EC 4.2.1.11]. Our predicted cytosolic localization would require export of organic carbon from the plastid to provide substrate for the enolase and subsequent re-import of organic carbon back into the plastids. Further, our comparative sequence study of the enolase and its 3D-structure prediction may suggest that the N-terminal extension found in green algal enolases could be involved in regulation of the enolase activity. In summary, we propose that the enolase represents one of the crucial regulatory bottlenecks in carbon partitioning in green algae.

Isoprenoid biosynthesis in eukaryotic phototrophs: a spotlight on algae.

Isoprenoids are one of the largest groups of natural compounds and have a variety of important functions in the primary metabolism of land plants and algae. In recent years, our understanding of the numerous facets of isoprenoid metabolism in land plants has been rapidly increasing, while knowledge on the metabolic network of isoprenoids in algae still lags behind. Here, current views on the biochemistry and genetics of the core isoprenoid metabolism in land plants and in the major algal phyla are compared and some of the most pressing open questions are highlighted. Based on the different evolutionary histories of the various groups of eukaryotic phototrophs, we discuss the distribution and regulation of the mevalonate (MVA) and the methylerythritol phosphate (MEP) pathways in land plants and algae and the potential consequences of the loss of the MVA pathway in groups such as the green algae. For the prenyltransferases, serving as gatekeepers to the various branches of terpenoid biosynthesis in land plants and algae, we explore the minimal inventory necessary for the formation of primary isoprenoids and present a preliminary analysis of their occurrence and phylogeny in algae with primary and secondary plastids. The review concludes with some perspectives on genetic engineering of the isoprenoid metabolism in algae.

The Dunaliella salina organelle genomes: large sequences, inflated with intronic and intergenic DNA.

BACKGROUND: Dunaliella salina Teodoresco, a unicellular, halophilic green alga belonging to the Chlorophyceae, is among the most industrially important microalgae. This is because D. salina can produce massive amounts of beta-carotene, which can be collected for commercial purposes, and because of its potential as a feedstock for biofuels production. Although the biochemistry and physiology of D. salina have been studied in great detail, virtually nothing is known about the genomes it carries, especially those within its mitochondrion and plastid. This study presents the complete mitochondrial and plastid genome sequences of D. salina and compares them with those of the model green algae Chlamydomonas reinhardtii and Volvox carteri. RESULTS: The D. salina organelle genomes are large, circular-mapping molecules with approximately 60% noncoding DNA, placing them among the most inflated organelle DNAs sampled from the Chlorophyta. In fact, the D. salina plastid genome, at 269 kb, is the largest complete plastid DNA (ptDNA) sequence currently deposited in GenBank, and both the mitochondrial and plastid genomes have unprecedentedly high intron densities for organelle DNA: approximately 1.5 and approximately 0.4 introns per gene, respectively. Moreover, what appear to be the relics of genes, introns, and intronic open reading frames are found scattered throughout the intergenic ptDNA regions -- a trait without parallel in other characterized organelle genomes and one that gives insight into the mechanisms and modes of expansion of the D. salina ptDNA. CONCLUSIONS: These findings confirm the notion that chlamydomonadalean algae have some of the most extreme organelle genomes of all eukaryotes. They also suggest that the events giving rise to the expanded ptDNA architecture of D. salina and other Chlamydomonadales may have occurred early in the evolution of this lineage. Although interesting from a genome evolution standpoint, the D. salina organelle DNA sequences will aid in the development of a viable plastid transformation system for this model alga, and they will complement the forthcoming D. salina nuclear genome sequence, placing D. salina in a group of a select few photosynthetic eukaryotes for which complete genome sequences from all three genetic compartments are available.

Identification and characterization of a new strain of the unicellular green alga Dunaliella salina (Teod.) from Korea.

The unicellular green alga Dunaliella salina is a halotolerant eukaryotic organism. Its halophytic properties provide an important advantage for open pond mass cultivation, since D. salina can be grown selectively. D. salina was originally described by E. C. Teodoresco in 1905. Since that time, numerous isolates of D. salina have been identified from hypersaline environments on different continents. The new Dunaliella strain used for this study was isolated from the salt farm area of the west coastal side of South Korea. Cells of the new strain were approximately oval- or pear-shaped (approximately 16-24 microm long and 10-15 microm wide), and contained one pyrenoid, cytoplasmatic granules, and no visible eyespot. Although levels of beta-carotene per cell were relatively low in cells grown at salinities between 0.5 to 2.5 M NaCl, cells grown at 4.5 M NaCl contained about a ten-fold increase in cellular levels of beta-carotene, which demonstrated that cells of the new Korean strain of Dunaliella can overaccumulate beta- carotene in response to salt stress. Analysis of the ITS1 and ITS2 regions of the new Korean isolate showed that it is in the same clade as D. salina. Consequently, based on comparative cell morphology, biochemistry, and molecular phylogeny, the new Dunaliella isolate from South Korea was classified as D. salina KCTC10654BP.

Xanthophyll biosynthetic mutants of Arabidopsis thaliana: altered nonphotochemical quenching of chlorophyll fluorescence is due to changes in Photosystem II antenna size and stability.

Xanthophylls (oxygen derivatives of carotenes) are essential components of the plant photosynthetic apparatus. Lutein, the most abundant xanthophyll, is attached primarily to the bulk antenna complex, light-harvesting complex (LHC) II. We have used mutations in Arabidopsis thaliana that selectively eliminate (and substitute) specific xanthophylls in order to study their function(s) in vivo. These include two lutein-deficient mutants, lut1 and lut2, the epoxy xanthophyll-deficient aba1 mutant and the lut2aba1 double mutant. Photosystem stoichiometry, antenna sizes and xanthophyll cycle activity have been related to alterations in nonphotochemical quenching of chlorophyll fluorescence (NPQ). Nondenaturing polyacrylamide gel electrophoresis indicates reduced stability of trimeric LHC II in the absence of lutein (and/or epoxy xanthophylls). Photosystem (antenna) size and stoichiometry is altered in all mutants relative to wild type (WT). Maximal DeltapH-dependent NPQ (qE) is reduced in the following order: WT>aba1>lut1 approximately lut2>lut2aba1, paralleling reduction in Photosystem (PS) II antenna size. Finally, light-activation of NPQ shows that zeaxanthin and antheraxanthin present constitutively in lut mutants are not qE active, and hence, the same can be inferred of the lutein they replace. Thus, a direct involvement of lutein in the mechanism of qE is unlikely. Rather, altered NPQ in xanthophyll biosynthetic mutants is explained by disturbed macro-organization of LHC II and reduced PS II-antenna size in the absence of the optimal, wild-type xanthophyll composition. These data suggest the evolutionary conservation of lutein content in plants was selected for due to its unique ability to optimize antenna structure, stability and macro-organization for efficient regulation of light-harvesting under natural environmental conditions.

Biosynthesis and distribution of chlorophyll among the photosystems during recovery of the green alga Dunaliella salina from irradiance stress.

To elucidate the mechanism of an irradiance-dependent adjustment in the chlorophyll (Chl) antenna size of Dunaliella salina, we investigated the regulation of expression of the Chl a oxygenase (CAO) and light-harvesting complex b (Lhcb) genes as a function of Chl availability in the photosynthetic apparatus. After a high-light to low-light shift of the cultures, levels of both CAO and Lhcb transcripts were rapidly induced by about 6-fold and reached a high steady-state level within 1.5 h of the shift. This was accompanied by repair of photodamaged photosystem II (PSII) reaction centers, accumulation of Chl a and Chl b (4:1 ratio), photosystem I (PSI), light-harvesting complex, and by enlargement of the Chl antenna size of both photosystems. In gabaculine-treated cells, induction of CAO and Lhcb transcripts was not affected despite substantial inhibition in de novo Chl biosynthesis. However, cells were able to synthesize and accumulate some Chl a and Chl b (1:1 ratio), resulting in a marked lowering of the Chl a to Chl b ratio in the presence of this inhibitor. Assembly incorporation of light-harvesting complex and a corresponding Chl antenna size increase, mostly for the existing photosystems, was noted in the presence of gabaculine. Repair of photodamaged PSII was not affected by gabaculine. However, assembly accumulation of new PSI was limited under such conditions. These results suggest a coordinate regulation of CAO and Lhcb gene transcription by irradiance, independent of Chl availability. The results are discussed in terms of different signal transduction pathways for the regulation of the photosynthetic apparatus organization by irradiance.