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1.  The quantitative proteomic response of Synechocystis sp. PCC6803 to phosphate acclimation 
Aquatic Biosystems  2013;9:5.
Inorganic phosphate (Pi) is a critical nutrient for all life and is periodically limiting in marine and freshwater provinces, yet little is understood how organisms acclimate to fluctuations in Pi within their environment. To investigate whole cell adaptation, we grew Synechocystis sp. PCC6803, a model freshwater cyanobacterium, in 3%, and 0.3% inorganic phosphate (Pi) media. The cells were allowed to acclimate over 60 days, and cells were harvested for quantitative high throughput mass spectrometry-based proteomics using the iTRAQ™ labelling technology.
In total, 120 proteins were identified, and 52 proteins were considered differentially abundant compared to the control. Alkaline phosphatase (APase) activities correlated significantly (p < 0.05) with observed relative PhoA abundances. PstS1 and PstS2 were both observed, yet PstS1 was not differentially more abundant than the control. Phycobilisome protein abundances appeared to be coordinated, and are significantly less abundant in 0.3% Pi than 3% Pi cultures. Also, the central metabolic cell function appears to have shifted towards the production of (NADPH) reducing energy and nucleotide sugars.
This acclimation response bears strong similarity to the previously reported response to nitrogen deprivation within Synechocystis sp. PCC 6803. However, it also demonstrates some characteristics of desiccation stress, such as the regulation of fatty acids and increased abundance of rehydrin in the 3% Pi culture.
PMCID: PMC3600050  PMID: 23442353
Cyanobacteria; iTRAQ; Phosphate acclimation; Phycobilisome; Pentose phosphate pathway; Ribose sugars; Synechocystis
2.  Influence of fermentation conditions on the surface properties and adhesion of Lactobacillus rhamnosus GG 
The surface properties of probiotic bacteria influence to a large extent their interactions within the gut ecosystem. There is limited amount of information on the effect of the production process on the surface properties of probiotic lactobacilli in relation to the mechanisms of their adhesion to the gastrointestinal mucosa. The aim of this work was to investigate the effect of the fermentation pH and temperature on the surface properties and adhesion ability to Caco-2 cells of the probiotic strain Lactobacillus rhamnosus GG.
The cells were grown at pH 5, 5.5, 6 (temperature 37°C) and at pH 6.5 (temperature 25°C, 30°C and 37°C), and their surfaces analysed by X-ray photoelectron spectrometry (XPS), Fourier transform infrared spectroscopy (FT-IR) and gel-based proteomics. The results indicated that for all the fermentation conditions, with the exception of pH 5, a higher nitrogen to carbon ratio and a lower phosphate content was observed at the surface of the bacteria, which resulted in a lower surface hydrophobicity and reduced adhesion levels to Caco-2 cells as compared to the control fermentation (pH 6.5, 37°C). A number of adhesive proteins, which have been suggested in previous published works to take part in the adhesion of bacteria to the human gastrointestinal tract, were identified by proteomic analysis, with no significant differences between samples however.
The temperature and the pH of the fermentation influenced the surface composition, hydrophobicity and the levels of adhesion of L. rhamnosus GG to Caco-2 cells. It was deduced from the data that a protein rich surface reduced the adhesion ability of the cells.
PMCID: PMC3441878  PMID: 22931558
3.  Comparative quantitative proteomics of prochlorococcus ecotypes to a decrease in environmental phosphate concentrations 
Aquatic Biosystems  2012;8:7.
The well-lit surface waters of oligotrophic gyres significantly contribute to global primary production. Marine cyanobacteria of the genus Prochlorococcus are a major fraction of photosynthetic organisms within these areas. Labile phosphate is considered a limiting nutrient in some oligotrophic regions such as the Caribbean Sea, and as such it is crucial to understand the physiological response of primary producers such as Prochlorococcus to fluctuations in the availability of this critical nutrient.
Prochlorococcus strains representing both high light (HL) (MIT9312) and low light (LL) (NATL2A and SS120) ecotypes were grown identically in phosphate depleted media (10 μM Pi). The three strains displayed marked differences in cellular protein expression, as determined by high throughput large scale quantitative proteomic analysis. The only strain to demonstrate a significantly different growth rate under reduced phosphate conditions was MIT9312. Additionally, there was a significant increase in phosphate-related proteins such as PhoE (> 15 fold increase) and a depression of the Rubisco protein RbcL abundance in this strain, whereas there appeared to be no significant change within the LL strain SS120.
This differential response between ecotypes highlights the relative importance of phosphate availability to each strain and from these results we draw the conclusion that the expression of phosphate acquisition mechanisms are activated at strain specific phosphate concentrations.
PMCID: PMC3349580  PMID: 22480396
Prochlorococcus; PstS; PhoA; PhoE; Growth; Phosphate
4.  A systems biology approach to investigate the response of Synechocystis sp. PCC6803 to a high salt environment 
Saline Systems  2009;5:8.
Salt overloading during agricultural processes is causing a decrease in crop productivity due to saline sensitivity. Salt tolerant cyanobacteria share many cellular characteristics with higher plants and therefore make ideal model systems for studying salinity stress. Here, the response of fully adapted Synechocystis sp. PCC6803 cells to the addition of 6% w/v NaCl was investigated using proteomics combined with targeted analysis of transcripts.
Isobaric mass tagging of peptides led to accurate relative quantitation and identification of 378 proteins, and approximately 40% of these were differentially expressed after incubation in BG-11 media supplemented with 6% salt for 9 days. Protein abundance changes were related to essential cellular functional alterations. Differentially expressed proteins involved in metabolic responses were also analysed using the probabilitistic tool Mixed Model on Graphs (MMG), where the role of energy conversion through glycolysis and reducing power through pentose phosphate pathway were highlighted. Temporal RT-qPCR experiments were also run to investigate protein expression changes at the transcript level, for 14 non-metabolic proteins. In 9 out of 14 cases the mRNA changes were in accordance with the proteins.
Synechocystis sp. PCC6803 has the ability to regulate essential metabolic processes to enable survival in high salt environments. This adaptation strategy is assisted by further regulation of proteins involved in non-metabolic cellular processes, supported by transcriptional and post-transcriptional control. This study demonstrates the effectiveness of using a systems biology approach in answering environmental, and in particular, salt adaptation questions in Synechocystis sp. PCC6803
PMCID: PMC2743698  PMID: 19735556
5.  Proteomics with a pinch of salt: A cyanobacterial perspective 
Saline Systems  2008;4:1.
Cyanobacteria are ancient life forms and have adapted to a variety of extreme environments, including high salinity. Biochemical, physiological and genetic studies have contributed to uncovering their underlying survival mechanisms, and as recent studies demonstrate, proteomics has the potential to increase our overall understanding further. To date, most salt-related cyanobacterial proteomic studies have utilised gel electrophoresis with the model organism Synechocystis sp. PCC6803. Moreover, focus has been on 2–4% w/v NaCl concentrations within different cellular compartments. Under these conditions, Synechocystis sp. PCC6803 was found to respond and adapt to salt stress through synthesis of general and specific stress proteins, altering the protein composition of extracellular layers, and re-directing control of complex central intermediary pathways. Post-transcriptional control was also predicted through non-correlating transcript level data and identification of protein isoforms.
In this paper, we also review technical developments with emphasis on improving the quality and quantity of proteomic data and overcoming the detrimental effects of salt on sample preparation and analysis. Developments in gel-free methods include protein and peptide fractionation workflows, which can increase coverage of the proteome (20% in Synechocystis sp. PCC6803). Quantitative techniques have also improved in accuracy, resulting in confidence in quantitation approaching or even surpassing that seen in transcriptomic techniques (better than 1.5-fold in differential expression). Furthermore, in vivo metabolic labelling and de novo protein sequencing software have improved the ability to apply proteomics to unsequenced environmental isolates. The example used in this review is a cyanobacterium isolated from a Saharan salt lake.
PMCID: PMC2386806  PMID: 18412952
6.  Macromolecular Fingerprinting of Sulfolobus Species in Biofilm: A Transcriptomic and Proteomic Approach Combined with Spectroscopic Analysis 
Journal of Proteome Research  2011;10(9):4105-4119.
Microorganisms in nature often live in surface-associated sessile communities, encased in a self-produced matrix, referred to as biofilms. Biofilms have been well studied in bacteria but in a limited way for archaea. We have recently characterized biofilm formation in three closely related hyperthermophilic crenarchaeotes: Sulfolobus acidocaldarius, S. solfataricus, and S. tokodaii. These strains form different communities ranging from simple carpet structures in S. solfataricus to high density tower-like structures in S. acidocaldarius under static condition. Here, we combine spectroscopic, proteomic, and transcriptomic analyses to describe physiological and regulatory features associated with biofilms. Spectroscopic analysis reveals that in comparison to planktonic life-style, biofilm life-style has distinctive influence on the physiology of each Sulfolobus spp. Proteomic and transcriptomic data show that biofilm-forming life-style is strain specific (eg ca. 15% of the S. acidocaldarius genes were differently expressed, S. solfataricus and S. tokodaii had ∼3.4 and ∼1%, respectively). The -omic data showed that regulated ORFs were widely distributed in basic cellular functions, including surface modifications. Several regulated genes are common to biofilm-forming cells in all three species. One of the most striking common response genes include putative Lrs14-like transcriptional regulators, indicating their possible roles as a key regulatory factor in biofilm development.
S. acidocaldarius, S. solfataricus, and S. tokodaii strains were grown independently as biofilms. Comparison between planktonic and biofilm cell popupations of all three strains was performed by spectroscopic analysis (FTIR and XPS), iTRAQ proteomics, and RNA microarrays. To highlight common features in biofilm formation among the Sulfolobus strains, the data is presented as a comparative analysis. One of the most striking common response genes include putative Lrs14-like transcriptional regulators, suggesting their roles as key regulatory factor in biofilm development.
PMCID: PMC3166137  PMID: 21761944
archaea; sulfolobus; biofilm; proteomics; transcriptomics; FTIR; thermophilic; acidophilic

Results 1-6 (6)