Genome sequences of two Synechococcus ecotypes inhabiting the octopus Spring microbial mat in Yellowstone National Park revealed the presence of all genes required for nitrogenase biosynthesis. We demonstrate that nif genes of the Synechococcus ecotypes are expressed in situ in a region of the mat that varies in temperature from 53.5 degrees C to 63.4 degrees C (average 60 degrees C) transcripts are only detected at the end of the day when the mat becomes anoxic. Nitrogenase activity in mat samples was also detected in the evening. Hitherto, N-2 fixation in hot spring mats was attributed either to filamentous cyanobacteria (not present at > 50 degrees C in these mats) or to heterotrophic bacteria. To explore how energy-generating processes of the Synechococcus ecotypes track natural light and O-2 conditions, we evaluated accumulation of transcripts encoding proteins involved in photosynthesis, respiration, and fermentation. Transcripts from photosynthesis (cpcF, cpcE, psaB, and psbB) and respiration (coxA and cydA) genes declined in the evening. In contrast, transcripts encoding enzymes that may participate in fermentation fell into two categories; some (Idh, pdhB, ald, and ackA) decreased in the evening, whereas others (PflB, pflA, adhE, and acs) increased at the end of the day and remained high into the night. Energy required for N-2 fixation during the night may be derived from fermentation pathways that become prominent as the mat becomes anoxic. In a broader context, our data suggest that there are critical regulatory switches in situ that are linked to the diel cycle and that these switches alter many metabolic processes within the microbial mat.

Polyketide synthases (PKSs) occur in many bacteria, fungi and plants. They are highly versatile enzymes involved in the biosynthesis of a large variety of compounds including antimicrobial agents, polymers associated with bacterial cell walls and plant pigments. While harmful algae are known to produce polyketide toxins, sequences of the genomes of non-toxic algae, including those of many green algal species, have surprisingly revealed the presence of genes encoding type I PKSs. The genome of the model alga Chlamydomonas reinhardtii (Chlorophyta) contains a single type I PKS gene, designated PKS1 (Cre10.g449750), which encodes a giant PKS with a predicted mass of 2.3 MDa. Here, we show that PKS1 is induced in 2-day-old zygotes and is required for their development into zygospores, the dormant stage of the zygote. Wild-type zygospores contain knob-like structures (similar to 50 nm diameter) that form at the cell surface and develop a central cell wall layer; both of these structures are absent from homozygous pks1 mutants. Additionally, in contrast to wild-type zygotes, chlorophyll degradation is delayed in homozygous pks1 mutant zygotes, indicating a disruption in zygospore development. In agreement with the role of the PKS in the formation of the highly resistant zygospore wall, mutant zygotes have lost the formidable desiccation tolerance of wild-type zygotes. Together, our results represent functional analyses of a PKS mutant in a photosynthetic eukaryotic microorganism, revealing a central function for polyketides in the sexual cycle and survival under stressful environmental conditions.

Phytoplankton in nature must acclimate to a wide range of light conditions resulting from diel light cycles, ocean circulation and mixing, cloud cover, and the variable bio-optical characteristics of the water column. In this study, we used whole-genome cDNA microarrays to investigate the effects of a gradually fluctuating daily light cycle on gene expression in the cyanobacterium Synechocystis sp. strain PCC6803. From these data, we developed a conceptual framework depicting the diel regulation of metabolic pathways in the cell. The framework is focused on potential photoacclimation responses, including the regulation of the photosystems, cell division, and DNA replication. The mRNA abundance of genes involved in many metabolic pathways, and particularly those encoding proteins that function in photosynthesis and DNA replication, changed markedly over the course of the day. The levels of mRNA encoding polypeptides important for the formation of the light-harvesting apparatus, photosystems I and II, and cell division were found in high concentrations during the day. The transcript levels of many genes encoding enzymes involved in anabolic processes also increased considerably during the day. In contrast, transposon transcripts and mRNAs encoding proteins involved in DNA replication, cell wall synthesis, and respiratory activity were not found in high concentrations during the day. Although gradually varying light exposure induced significant changes in transcript accumulation within Synechocystis, the direction of these changes differed between our study and previous studies in which there was an abrupt transition between irradiances.

Metabolic exchange between cnidarians and their symbiotic dinoflagellates is central to maintaining their mutualistic relationship. Sugars are translocated to the host, while ammonium and nitrate are utilized by the dinoflagellates (Symbiodinium spp.). We investigated membrane protein sequences of each partner to identify potential transporter proteins that move sugars into cnidarian cells and nitrogen products into Symbiodinium cells. We examined the facilitated glucose transporters (GLUT), sodium/glucose cotransporters (SGLT), and aquaporin (AQP) channels in the cnidarian host as mechanisms for sugar uptake, and the ammonium and high-affinity nitrate transporters (AMT and NRT2, respectively) in the algal symbiont as mechanisms for nitrogen uptake. Homologous protein sequences were used for phylogenetic analysis and tertiary structure deductions. In cnidarians, we identified putative glucose transporters of the GLUT family and glycerol transporting AQP proteins, as well as sodium monocarboxylate transporters and sodium myo-inositol cotransporters homologous to SGLT proteins. We hypothesize that cnidarians use GLUT proteins as the primary mechanism for glucose uptake, while glycerol moves into cells by passive diffusion. We also identified putative AMT proteins in several Symbiodinium clades and putative NRT2 proteins only in a single clade. We further observed an upregulation of expressed putative AMT proteins in Symbiodinium, which may have emerged as an adaptation to conditions experienced inside the host cell. This study is the first to identify transporter sequences from a diversity of cnidarian species and Symbiodinium clades, which will be useful for future experimental analyses of the host-symbiont proteome and the nutritional exchange of Symbiodinium cells in hospite.

We have carefully characterized and reexamined the motility and phototactic responses of Synechoeystis sp. adenylyl cyclase (Cya1) and catabolite activator protein (SYCRP1) mutants to different light regimens, glucose, 3-(3,4-dichlorophenyl)-1,1-dimethylurea, and cyclic AMP. We find that contrary to earlier reports, cya1 and sycrp1 mutants are motile and phototactic but are impaired in one particular phase of phototaxis in comparison with wild-type Synechoeystis sp.

In the mid-20th century, the unicellular and genetically tractable green alga Chlamydomonas reinhardtii was first developed as a model organism to elucidate fundamental cellular processes such as photosynthesis, light perception and the structure, function and biogenesis of cilia. Various studies of C. reinhardtii have profoundly advanced plant and cell biology, and have also impacted algal biotechnology and our understanding of human disease. However, the 'real' life of C. reinhardtii in the natural environment has largely been neglected. To extend our understanding of the biology of C. reinhardtii, it will be rewarding to explore its behavior in its natural habitats, learning more about its abundance and life cycle, its genetic and physiological diversity, and its biotic and abiotic interactions.

The hliA gene of the cyanobacterium Synechococcus elongatus PCC 7942 is known to be upregulated by high-intensity light through the activity of the NblS sensor kinase. In this work it was found that, within the hliA upstream region, changes to the sequence around -30 to -25 (relative to the transcriptional start site) resulted in elevated hliA expression, implicating this region in negative regulation of the gene. Electrophoretic mobility shift assays performed were consistent with a protein binding this region that acts to keep the gene off in lower light. A reduction in gene dosage of nblS in vivo resulted in enhanced hliA expression, suggesting that negative control of hliA is mediated through NblS. An extended version of the high light regulatory 1 (HLR1) motif (previously described in Synechocystis PCC 6803) was identified within the sequence surrounding -30 to -25 of hliA. The extended HLR1 sequence was found upstream of other NblS-controlled genes from S. elongatus and Synechocystis PCC 6803 and upstream of hli genes from a variety of cyanobacterial and related genomes. These results point to the evolutionary conservation of the HLR1 element and its importance in NblS-mediated signaling and yield new insight into NblS-mediated control of gene expression.

The genes encoding the alpha and beta subunits of allophycocyanin, phycocyanin and phycoerythrin from the red alga Aglaothamnion neglectum were isolated and characterized. While the operons containing the different phycobiliprotein genes are dispersed on the plastid genome, the genes encoding the alpha and beta subunits for each phycobiliprotein are contiguous. The beta subunit gene is 5' for both the phycocyanin and phycoerythrin operons, while the alpha subunit gene is 5' for the allophycocyanin operon. The amino acid sequences of A. neglectum phycobiliproteins, as deduced from the nucleotide sequences of the genes, are 65-85% identical to analogous proteins from other red algae and cyanobacteria. The conserved nature of the plastid-encoded red algal and cyanobacterial phycobiliprotein genes supports the proposed origin of red algal plastids from cyanobacterial endosymbionts.

We have designed a microfluidic device in which we can manipulate, lyse, label, separate, and quantify the protein contents of a single cell using single-molecule fluorescence counting. Generic labeling of proteins is achieved through fluorescent-antibody binding. The use of cylindrical optics enables high-efficiency (approximate to 60%) counting of molecules in micrometer-sized channels. We used this microfluidic device to quantify beta(2) adrenergic receptors expressed in insect cells (SF9). We also analyzed phycobiliprotein contents in individual cyanobacterial cells ( Synechococcus sp. PCC 7942) and observed marked differences in the levels of specific complexes in cell populations that were grown under nitrogen-depleted conditions.

Metabolite exchange is fundamental to the viability of the cnidarian-Symbiodiniaceae symbiosis and survival of coral reefs. Coral holobiont tolerance to environmental change might be achieved through changes in Symbiodiniaceae species composition, but differences in the metabolites supplied by different Symbiodiniaceae species could influence holobiont fitness. Using C-13 stable-isotope labelling coupled to gas chromatography-mass spectrometry, we characterized newly fixed carbon fate in the model cnidarian Exaiptasia pallida (Aiptasia) when experimentally colonized with either native Breviolum minutum or non-native Durusdinium trenchii Relative to anemones containing B. minutum, D. trenchii-colonized hosts exhibited a 4.5-fold reduction in C-13-labelled glucose and reduced abundance and diversity of C-13-labelled carbohydrates and lipogenesis precursors, indicating symbiont species-specific modifications to carbohydrate availability and lipid storage. Mapping carbon fate also revealed significant alterations to host molecular signalling pathways. In particular, D. trenchii-colonized hosts exhibited a 40-fold reduction in C-13-labelled scyllo-inositol, a potential interpartner signalling molecule in symbiosis specificity. C-13-labelling also highlighted differential antioxidant- and ammonium-producing pathway activities, suggesting physiological responses to different symbiont species. Such differences in symbiont metabolite contribution and host utilization may limit the proliferation of stress-driven symbioses; this contributes valuable information towards future scenarios that select in favour of less-competent symbionts in response to environmental change.