Quantitative proteomic comparison of salt stress in Chlamydomonas reinhardtii and the snow alga Chlamydomonas nivalis reveals mechanisms for salt-triggered fatty acid accumulation via reallocation of carbon resources.

BACKGROUND: Chlamydomonas reinhardtii is a model green alga strain for molecular studies; its fully sequenced genome has enabled omic-based analyses that have been applied to better understand its metabolic responses to stress. Here, we characterised physiological and proteomic changes between a low-starch C. reinhardtii strain and the snow alga Chlamydomonas nivalis, to reveal insights into their contrasting responses to salinity stress. RESULTS: Each strain was grown in conditions tailored to their growth requirements to encourage maximal fatty acid (as a proxy measure of lipid) production, with internal controls to allow comparison points. In 0.2 M NaCl, C. nivalis accumulates carbohydrates up to 10.4% DCW at 80 h, and fatty acids up to 52.0% dry cell weight (DCW) over 12 days, however, C. reinhardtii does not show fatty acid accumulation over time, and shows limited carbohydrate accumulation up to 5.5% DCW. Analysis of the C. nivalis fatty acid profiles showed that salt stress improved the biofuel qualities over time. Photosynthesis and respiration rates are reduced in C. reinhardtii relative to C. nivalis in response to 0.2 M NaCl. De novo sequencing and homology matching was used in conjunction with iTRAQ-based quantitative analysis to identify and relatively quantify proteomic alterations in cells exposed to salt stress. There were abundance differences in proteins associated with stress, photosynthesis, carbohydrate and lipid metabolism proteins. In terms of lipid synthesis, salt stress induced an increase in dihydrolipoyl dehydrogenase in C. nivalis (1.1-fold change), whilst levels in C. reinhardtii remained unaffected; this enzyme is involved in acetyl CoA production and has been linked to TAG accumulation in microalgae. In salt-stressed C. nivalis there were decreases in the abundance of UDP-sulfoquinovose (- 1.77-fold change), which is involved in sulfoquinovosyl diacylglycerol metabolism, and in citrate synthase (- 2.7-fold change), also involved in the TCA cycle. Decreases in these enzymes have been shown to lead to increased TAG production as fatty acid biosynthesis is favoured. Data are available via ProteomeXchange with identifier PXD018148. CONCLUSIONS: These differences in protein abundance have given greater understanding of the mechanism by which salt stress promotes fatty acid accumulation in the un-sequenced microalga C. nivalis as it switches to a non-growth state, whereas C. reinhardtii does not have this response.

Change of carbon source causes dramatic effects in the phospho-proteome of the archaeon Sulfolobus solfataricus.

Protein phosphorylation is known to occur in Archaea. However, knowledge of phosphorylation in the third domain of life is rather scarce. Homology-based searches of archaeal genome sequences reveals the absence of two-component systems in crenarchaeal genomes but the presence of eukaryotic-like protein kinases and protein phosphatases. Here, the influence of the offered carbon source (glucose versus tryptone) on the phospho-proteome of Sulfolobus solfataricus P2 was studied by precursor acquisition independent from ion count (PAcIFIC). In comparison to previous phospho-proteome studies, a high number of phosphorylation sites (1318) located on 690 phospho-peptides from 540 unique phospho-proteins were detected, thus increasing the number of currently known archaeal phospho-proteins from 80 to 621. Furthermore, a 25.8/20.6/53.6 Ser/Thr/Tyr percentage ratio with an unexpectedly high predominance of tyrosine phosphorylation was detected. Phospho-proteins in most functional classes (21 out of 26 arCOGs) were identified, suggesting an important regulatory role in S. solfataricus. Focusing on the central carbohydrate metabolism in response to the offered carbon source, significant changes were observed. The observed complex phosphorylation pattern hints at an important physiological function of protein phosphorylation in control of the central carbohydrate metabolism, which might particularly operate in channeling carbon flux into the respective metabolic pathways.

A review of current proteomics technologies with a survey on their widespread use in reproductive biology investigations.

Proteomics is very much a technology-driven field. The ambition is to identify, quantify and to assess the state of posttranslational modification and interaction partners for every protein in the cell. The proteome is in a state of flux and is thus extremely complex. Analysis of the proteome is exacerbated by the huge dynamic concentration range of proteins in the cellular environment. The impact that mass spectrometry-based proteomics has had on the field of biology has heavily depended on dramatic improvements in mass spectrometry that have been made in recent years. We examined 1541 reports indexed in PubMed relating to proteomics and reproduction to identify trends in the field and to make some broad observations for future work. To set the scene, in the first part of the report, we give a comprehensive overview of proteomics and associated techniques and technologies (such as separations and mass spectrometry). The second part examines the field in light of these techniques and suggests some opportunities for application of these tools in the area of reproduction.

Aligning conservation priorities across taxa in Madagascar with high-resolution planning tools.

Globally, priority areas for biodiversity are relatively well known, yet few detailed plans exist to direct conservation action within them, despite urgent need. Madagascar, like other globally recognized biodiversity hot spots, has complex spatial patterns of endemism that differ among taxonomic groups, creating challenges for the selection of within-country priorities. We show, in an analysis of wide taxonomic and geographic breadth and high spatial resolution, that multitaxonomic rather than single-taxon approaches are critical for identifying areas likely to promote the persistence of most species. Our conservation prioritization, facilitated by newly available techniques, identifies optimal expansion sites for the Madagascar government's current goal of tripling the land area under protection. Our findings further suggest that high-resolution multitaxonomic approaches to prioritization may be necessary to ensure protection for biodiversity in other global hot spots.

Carbon substrate utilisation profile of a high concentration effluent degrading microbial consortium.

This paper presents the carbon substrate utilisation profile of a group of microorganisms responsible for the biodegradation of a highly concentrated industrial effluent. A 1 litre bioreactor was used to study this consortium's biodegradation potential, with the chemical oxygen demand (COD) of the waste being reduced by 90% from 24000 mg l(-1) within 456 hours. This study also demonstrates that the consortium is capable of degrading organic solvents, such as isopropanol, at concentrations of 260 mg l(-1). The population distribution and biochemical behaviour were also characterised using denaturing gradient gel electrophoresis (DGGE) and Biolog Eco Plates at various stages of bioreactor operation. The DGGE results indicated that the dominant bands of the microbial population profile were stable at various operational stages, and that only a few bands varied with time. Moreover, four Biolog Eco plates were inoculated with samples drawn from the bioreactor at 0, 24, 72 and 120 hours after inoculation. Based on this Biolog Eco Plate profiling, a carbon source utilisation analysis was conducted to group the substrates according to their colour development patterns. Patterns were quantified via measurement of well optical density. Subsequently, cross-correlation statistical techniques were used to establish the existence of recurrent behavioural responses from each carbon source to the various wastewater samples. From the cross correlation, an attempt was made to classify the metabolic potential for future biodegradation processes. Carbohydrates, amino acids and carboxylic acids were the most predominant groups of sole carbon substrates showing similar growth behaviour in the consortium.

Metabolic flux distribution analysis by 13C-tracer experiments using the Markov chain-Monte Carlo method.

Metabolic flux analysis using 13C-tracer experiments is an important tool in metabolic engineering since intracellular fluxes are non-measurable quantities in vivo. Current metabolic flux analysis approaches are fully based on stoichiometric constraints and carbon atom balances, where the over-determined system is iteratively solved by a parameter estimation approach. However, the unavoidable measurement noises involved in the fractional enrichment data obtained by 13C-enrichment experiment and the possible existence of unknown pathways prevent a simple parameter estimation method for intracellular flux quantification. The MCMC (Markov chain-Monte Carlo) method, which obtains intracellular flux distributions through delicately constructed Markov chains, is shown to be an effective approach for deep understanding of the intracellular metabolic network. Its application is illustrated through the simulation of an example metabolic network.

Application of a solvent-tolerant microbial consortium for biofiltration of extremely high concentration gaseous solvent streams.

The aerobic biological oxidation of 2-propanol (isopropyl alcohol, IPA) at extremely high concentrations in air by an enriched solvent-tolerant microbial consortium operating at ambient temperature was evaluated for six months. Solvent-tolerant microbial cells were immobilised onto porous glass pall rings and fed with either IPA or its metabolic product acetone as sole carbon source. Successful biofiltration of solvent vapour at a concentration of 24 g m(-3) was achieved with oxidation of up to 100% total inlet carbon. The maximum IPA mass loading and IPA elimination capacity (EC) was 1700 g m(-3) h(-1). This performance exceeds all previous values published in the literature for similar processes. A slip feed experiment, using acetone, was also performed in order to assess the substrate specificity performance. The biofilter responded successfully to a switch from acetone to IPA as sole carbon source, displaying little reduction in overall organic carbon removal.

Anogenital gland secretions of Lemur catta and Propithecus verreauxi coquereli: a preliminary chemical examination.

Although prosimians are greatly olfaction-oriented, little is known about the specifics of how they use scent to communicate. In this preliminary study we attempted to delineate intra- and interspecific differences among the anogenital gland secretions of two lemur species (Lemur catta and Propithecus verreauxi coquereli) using gas chromatography-mass spectrometry (GC-MS). The results indicate that the two species are discernible through scent. Furthermore, we were able to identify reproductive status using this technique. The anogenital secretions of the different sexes in L. catta, though perhaps not P. v. coquereli, are chemically distinguishable. Given this information, it appears that at least some lemur species can use scent marks to determine species, sex, and reproductive status.

Culture of the marine cyanobacterium, Lyngbya majuscula (Oscillatoriaceae), for bioprocess intensified production of cyclic and linear lipopeptides.

Cyanobacteria are an ancient and diverse group of photosynthetic microorganisms, which inhabit many different and extreme environments. This indicates a high degree of biological adaptation, which has enabled these organisms to thrive and compete effectively in nature. The filamentous cyanobacterium, Lyngbya majuscula, produces several promising antifungal and cytotoxic agents, including laxaphycin A and B and curacin A. Samples of L. majuscula collected from Moorea Island, Tahiti (French Polynesia) and from the Culture Collection of Algae and Protozoa (CCAP 1446/4) were studied and adapted to large scale laboratory culture (5 l). This constitutes a 100-fold scale-up for the culture of this particular strain of L. majuscula. The effect of culture vessel configurations, growth conditions and media compositions on growth of L. majuscula was examined. Using optimised culture conditions, two strains of L. majuscula are currently being evaluated for their production of secondary metabolites. Results will be compared with those obtained from four environmental extracts. Comparisons were made by thin layer chromatography (TLC), high performance liquid chromatography (HPLC) and fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS). It was shown that varying the culture conditions under which L. majuscula was grown had the greatest effect on secondary metabolite production, thus providing potential for future bioprocess intensified production.

High-pressure adaptation by salt stress in a moderately halophilic bacterium obtained from open seawater.

High-pressure adaptation was examined using a moderately halophilic bacterium (Micrococcus roseus), which was isolated from open seawater and capable of growing in 15% w/v NaCl (optimum NaCl concentration: 3% w/v). After treatment at 207 MPa, colony-forming units (CFUs) significantly decreased; however, the loss of integral cells after pressure was only 30% when direct cell count was performed microscopically. In order to investigate the piezotolerance of M. roseus under high pressure without morphological change, the survival of cells was examined under pressure at 138 MPa for 2 h. M. roseus in 3% NaCl was still sensitive to pressure at 138 MPa. However, the cells in the third generations showed remarkably increased pressure resistance, and no significant loss of viability was confirmed. Furthermore, when M. roseus was cultured in 1, 3, 5, 10 and 15% NaCl, the survival ratio proportionally increased at increased NaCl concentration. M. roseus cultured in 15% NaCl was remarkably resistant (94.7% viability) to pressure at 138 MPa, even when suspended in lower concentration of NaCl. This suggests that NaCl concentrations in growth culture affect the piezotolerance of M. roseus and that this species has an ability to adapt to high pressure.

Identification of the green alga, Chlorella vulgaris (SDC1) using cyanobacteria derived 16S rDNA primers: targeting the chloroplast.

We have tested a set of oligonucleotide primers originally developed for the specific amplification of 16S rRNA gene segments from cyanobacteria, in order to determine their versatility as an identification tool for phototrophic eucaryotes. Using web-based bioinformatics tools we determined that these primers not only targeted cyanobacterium sequences as previously described, but also 87% of sequences derived from phototrophic eucaryotes. In order to qualify our finding, a type culture and environmental strain from the freshwater unicellular, green algae genus Chlorella Beijerinck, were selected for further study. Subsequently, we sequenced a 578-bp fragment of the 16S rRNA gene, which proved to be present within the chloroplast genome, performed sequence analysis and positively identified our solvent-degrading environmental strain (SDC1) as Chlorella vulgaris.

Biodegradation of isopropanol in a three phase fixed bed bioreactor: start up and acclimation using a previously-enriched microbial culture.

The aerobic biodegradation of high liquid phase concentrations of 2-propanol (IPA) by a previously enriched solvent-tolerant bacterial consortium within a 1.9 l fed-batch three phase fixed bed bioreactor was investigated. Solvent concentrations of up to 7.9 g l(-1) were investigated. Previously enriched solvent-tolerant bacterial cells were immobilised onto porous glass cylinders as a means obioprocess intensification. Bioreactor start-up and acclimation was studied anacetone concentration tracked as an indicator of IPA utilization, as the sole carbon source within a minimal salts medium (MSM). The initial batch treatment of IPA exhibited a biodegradation rate of 0.11 g l(-1) h(-1) prior to biofilm formation Biofilm growth during the second batch treatment was consistent with an increase in metabolic activity and an IPA biodegradation rate of 0.34 g l(-1), followed by a reduction of biodegradation rate to a constant value of 0.078 g l(-1) h(-1) after 650 h. A maximum acetone generation rate of 1.3 g l(-1) h(-1) was obtained during the fourth IPA addition although the maximum acetone biodegradation rate of 0.38 g l(-1) h(-1) was observed during the initial IPA addition. It is proposed that the metabolic lag resulting from switching from alcohol dehydrogenase to acetone carboxylase is a major rate-limiting step in the deep oxidation of IPA to acetone. The results demonstrate the potential of a previously enriched solvent-tolerant bacterial consortium in fixed bed bioreactor systems, for the aerobic treatment of concentrated solvent-containing wastestreams.

Biodegradation of propanol and isopropanol by a mixed microbial consortium.

The aerobic biodegradation of high concentrations of 1-propanol and 2-propanol (IPA) by a mixed microbial consortium was investigated. Solvent concentrations were one order of magnitude greater than any previously reported in the literature. The consortium utilized these solvents as their sole carbon source to a maximum cell density of 2.4 x 10(9) cells ml(-1). Enrichment experiments with propanol or IPA as carbon sources were carried out in batch culture and maximum specific growth rates (mumax) calculated. At 20 degrees C, mumax values were calculated to be 0.0305 h(-1) and 0.1093 h(-1) on 1% (v/v) IPA and 1-propanol, respectively. Growth on propanol and IPA was carried out between temperatures of 10 degrees C and 45 degrees C. Temperature shock responses by the microbial consortium at temperatures above 45 degrees C were demonstrated by considerable cell flocculation. An increase in propanol substrate concentration from 1% (v/v) to 2% (v/v) decreased the mumax from 0.1093 h(-1) to 0.0715 h(-1). Maximum achievable biodegradation rates of propanol and IPA were 6.11 x 10(-3)% (v/v) h(-1) and 2.72 x 10(-3)% (v/v) h(-1), respectively. Generation of acetone during IPA biodegradation commenced at 264 h and reached a maximum concentration of 0.4% (v/v). The results demonstrate the potential of mixed microbial consortia in the bioremediation of solvent-containing waste streams.

Lemur traits and Madagascar ecology: coping with an island environment.

The last decade's lemur research includes successes in discovering new living and extinct species and learning about the distribution, biogeography, physiology, behavior, and ecology of previously little-studied species. In addition, in both the dry forest and rain forest, long-term studies of lemur demography, life history, and reproduction, have been completed in conjunction with data on tree productivity, phenology, and climate. Lemurs contrast with anthropoids in several behavioral features, including female dominance, targeted female-female aggression, lack of sexual dimorphism regardless of mating system, sperm competition coupled with male-male aggression, high infant mortality, cathemerality, and strict seasonal breeding. Hypotheses to explain these traits include the "energy conservation hypothesis" (ECH) suggesting that harsh and unpredictable climate factors on the island of Madagascar have affected the evolution of female dominance, and the "evolutionary disequilibrium hypotheses" (EVDH) suggesting that the recent megafauna extinctions have influenced lemurs to become diurnal. These hypotheses are compared and contrasted in light of recent empirical data on climate, subfossils, and lemur behavior. New data on life histories of the rain forest lemurs at Ranomafana National Park give further support to the ECH. Birth seasons are synchronized within each species, but there is a 6-month distribution of births among species. Gestation and lactation lengths vary among sympatric lemurs, but all lemur species in the rain forest wean in synchrony at the season most likely to have abundant resources. Across-species weaning synchrony seen in Ranomafana corroborates data from the dry forest that late lactation and weaning is the life history event that is the primary focus of the annual schedule. Lemur adaptations may assure maximum offspring survival in this environment with an unpredictable food supply and heavy predation. In conclusion, a more comprehensive energy frugality hypothesis (EFH) is proposed, which postulates that the majority of lemur traits are either adaptations to conserve energy (e.g., low basal metabolic rate (BMR), torpor, sperm competition, small group size, seasonal breeding) or to maximize use of scarce resources (e.g., cathemerality, territoriality, female dominance, fibrous diet, weaning synchrony). Among primates, the isolated adaptive radiation of lemurs on Madagascar may have been uniquely characterized by selection toward efficiency to cope with the harsh and unpredictable island environment.

Diversity components of impending primate extinctions.

Many extant species are at risk to go extinct. This impending loss of species is likely to cause changes in future ecosystem functions. Ecological components of diversity, such as dietary or habitat specializations, can be used to estimate the impact of extinctions on ecosystem functions. As an approach to estimate the impact of future extinctions, we tested interdependency between ecological and taxonomic change based on current predictions of extinction rates in primates. We analyzed the ecological characteristics of extant primate faunas having species in various categories of endangerment of extinction and forecasted the future primate faunas as if they were paleontological faunas. Predicting future faunas combines the wealth of ecological information on living primates with large, fossil record-like changes in diversity. Predicted extinction patterns of living primates in Africa, Asia, Madagascar, and South America show that changes in ecology differ among the regions in ways that are not reducible to taxonomic measures. The ecological effects of primate extinctions are initially least severe in South America and larger in Asia and Africa. Disproportionately larger ecological changes are projected for Madagascar. The use of taxonomy as a proxy for ecology can mislead when estimating competence of future primate ecosystems.

A novel membrane protein is a mouse mammary tumor virus receptor.

Mouse mammary tumor virus (MMTV) infects a number of different cell types, including mammary gland and lymphoid cells, in vivo. To identify the cellular receptor for this virus, a mouse cDNA expression library was transfected into Cos-7 monkey kidney cells, and those transfected cells able to bind virus were selected by using antibody against the virus's cell surface envelope protein, gp52. One clone isolated from a library prepared from newborn thymus RNA, called MTVR, was able to confer virus binding to both monkey and human cells; this binding was blocked by anti-MTVR antibody. Moreover, transfection of MTVR into CV1 cells rendered them susceptible to infection by a murine leukemia virus-based retrovirus vector pseudotyped with the MMTV envelope protein. An epitope-tagged MTVR cofractionated with cellular membranes. Coimmunoprecipitation of the MMTV envelope protein and a MTVR-rabbit Fc fusion protein showed that these two proteins bound to each other. The MTVR sequence clone is unique, shows no homology to known membrane proteins, and is transcribed in many tissues.

A strategic approach to human resource planning in nursing.

In Britain, Canada and the USA, a chronic shortage of registered nurses has existed since World War II. This situation is predicted to become worse. Reviews both the general and nursing literature in human resource planning, and recommends a system which can be build up step by step and individualized for each nursing department. The core planning activities are a planning database, demand forecasting, supply forecasting, variance analysis and forecasting, and recruitment policies.