Project description:The proteome of the marine bacterium Photobacterium angustum S14 was exposed to UVB and analyzed by the implementation of both the post-digest ICPL labeling method and 2D-DIGE technique using exponentially growing cells. A total of 40 and 23 proteins were quantified in all replicates using either the ICPL or 2D-DIGE methods, respectively. By combining both datasets from 8 biological replicates (4 biological replicates for each proteomics technique), 55 proteins were found to respond significantly to UVB radiation in P. angustum. A total of 8 UVB biomarkers of P. angustum were quantified in all replicates using both methods. Among them, the protein found to present the highest increase in abundance (almost a 3-fold change) was RecA, which is known to play a crucial role in the so-called recombinational repair process. We also observed a high number of antioxidants, transport proteins, metabolism-related proteins, transcription/translation regulators, chaperonins and proteases. We also discuss and compare the UVB response and global protein expression profiles obtained for two different marine bacteria with trophic lifestyles: the copiotroph P. angustum and oligotroph Sphingopyxis alaskensis.
Project description:A type of photoheterotrophic bacteria contain a transmembrane light-driven proton pump called proteorhodopsins (PRs). Due to the prevalence of these organisms in the upper water column of the World's Ocean, and their potential for light-driven ATP generation, they have been suggested to significantly influence energy and matter flows in the biosphere. To date, evidence for the significance of the light-driven metabolism of PR-containing prokaryotes has been obtained by comparing growth in batch culture, under light versus dark conditions, and it appears that responses to light are linked to unfavorable conditions, which so far have not been well parameterized. We studied light responses to carbon yields of the PR-containing Photobacterium angustum S14 using continuous culture conditions and light-dark cycles. We observed significant effects of light-dark cycles compared to dark controls, as well as significant differences between samples after 12 h illumination versus 12 h darkness. However, these effects were only observed under higher cell counts and lower pH associated with higher substrate concentrations. Under these substrate levels Pirt's maintenance coefficient was higher when compared to lower substrate dark controls, and decreased under light-dark cycles. It appears that light responses by P. angustum S14 are induced by the energetic status of the cells rather than by low substrate concentrations.
Project description:UVB oxidizes proteins through the generation of reactive oxygen species. One consequence of UVB irradiation is carbonylation, the irreversible formation of a carbonyl group on proline, lysine, arginine or threonine residues. In this study, redox proteomics was performed to identify carbonylated proteins in the UVB resistant marine bacterium Photobacterium angustum. Mass-spectrometry was performed with either biotin-labeled or dinitrophenylhydrazide (DNPH) derivatized proteins. The DNPH redox proteomics method enabled the identification of 62 carbonylated proteins (5% of 1221 identified proteins) in cells exposed to UVB or darkness. Eleven carbonylated proteins were quantified and the UVB/dark abundance ratio was determined at both the protein and peptide levels. As a result we determined which functional classes of proteins were carbonylated, which residues were preferentially modified, and what the implications of the carbonylation were for protein function. As the first large scale, shotgun redox proteomics analysis examining carbonylation to be performed on bacteria, our study provides a new level of understanding about the effects of UVB on cellular proteins, and provides a methodology for advancing studies in other biological systems.
Project description:The molecular mechanisms underlying the physiological and cellular response to starvation are still not fully understood. We have used quantitative proteomics and RNA-seq to examine the temporal responses to starvation in the multicellular organism C. elegans, comparing the response in both wild type animals and in animals lacking the transcription factor HLH-30. Our findings show that starvation alters the abundance of hundreds of proteins and mRNAs in a temporal manner, many of which are involved in central metabolic pathways including lipoprotein metabolism. We show that hlh-30 animals die prematurely when starved, which can be prevented by knockdown of either vit-1 or vit-5, encoding two different lipoproteins. We show that the size and number of intestinal lipid droplets under starvation are altered in hlh-30 animals, that can be rescued by knockdown of vit-1, indicating that rescue of survival of hlh-30 animals under starvation conditions is closely linked to the size and number of intestinal lipid droplets.
Project description:How animals coordinate gene expression in response to starvation is an outstanding problem closely linked to aging, obesity, and cancer. Newly hatched Caenorhabditis elegans respond to food deprivation by halting development and promoting long-term survival (L1 diapause), thereby providing an excellent model to study starvation response. Through a genetic search, we have discovered that the tumor suppressor Rb critically promotes survival during L1 diapause and likely does so by regulating the expression of genes in both insulin-IGF-1 signaling (IIS)-dependent and -independent pathways mainly in neurons and the intestine. Global gene expression analyses suggested that Rb maintains the “starvation-induced transcriptome” and represses the “re-feeding induced transcriptome”, including the repression of many pathogen/toxin/oxidative stress-inducible and metabolic genes, as well as the activation of many other stress-resistant genes, mitochondrial respiratory chain genes, and potential IIS receptor antagonists. Notably, the majority of genes dysregulated in starved L1 Rb(-) animals were not found to be dysregulated in fed conditions. Together, these findings identify Rb as a critical regulator of the starvation response and suggest a link between functions of tumor suppressors and starvation survival. These results may provide mechanistic insights into why cancer cells are often hypersensitive to starvation treatment.
Project description:Aspergillus fumigatus has to cope with a combination of several stress types after colonisng the human body, and the interplay between the different stress responses can significantly influence the survival of this human pathogen during the invasion of the host organism. In this study, we examined how the H2O2 induced oxidative stress response depends on iron availability. Surprisingly, the applied H2O2 treatment, which induced only a negligible stress response in iron fed cultures, deleteriously affected the fungus under iron starvation and the majority of stress responses on the transcript level were characteristic only for the combined H2O2-iron starvation stress treatments. Our data suggest that the survival of the fungus highly depends on fragile balances, e.g. between siderophore and ergosterol productions or between economization on iron and production of essential iron containing proteins. The applied stress conditions also affected several processes related to virulence or drug susceptibility including secondary metabolism, zinc acquisition or antifungal drug transport. Our[It would be interesting to mention the antioxidative enzymes which are upregulated/increase in abundance under iron-depleting conditions] data clearly demonstrate that studying stress responses under single stress treatments is not sufficient to understand how fungal pathogens survive in a complex habitat and support the view that the evolutionary success of A. fumigatus as an opportunistic human pathogen is not the mere consequence of the productions of certain virulence factors. Importantly, this fungal pathogen is able to mount and coordinate high-complexity and outstandingly efficient responses to multiple and superpositioning stresses in various harsh habitats like the human body.
Project description:Cellular quiescence is a reversible differentiation state when cells are changing the gene expression programme to reduce metabolic functions and adapt to a new cellular environment. The epigenetic changes that accompany these alterations are not so well understood. Here we investigate the role of Leo1, a subunit of the conserved Paf1 (RNA polymerase-associated factor 1) complex, in the quiescence process using fission yeast as a model organism. Fission yeast cells enter the G0 phase of the cell cycle when exposed to nitrogen starvation and the heterochromatin regions become very dynamic. The reduction of heterochromatin in early G0 correlates with the reduced activity of target of rapamycin, TORC2, signalling. Cells lacking the Leo1 show reduced survival in G0. In these cells heterochromatin regions including subtelomeres are stabilized and many genes, including membrane transport genes, fail to be expressed. Our results suggest that Leo1 is essential for dynamic regulation of heterochromatin and gene expression during cellular quiescence.
Project description:How animals coordinate gene expression in response to starvation is an outstanding problem closely linked to aging, obesity, and cancer. Newly hatched Caenorhabditis elegans respond to food deprivation by halting development and promoting long-term survival (L1 diapause), thereby providing an excellent model to study starvation response. Through a genetic search, we have discovered that the tumor suppressor Rb critically promotes survival during L1 diapause and likely does so by regulating the expression of genes in both insulin-IGF-1 signaling (IIS)-dependent and -independent pathways mainly in neurons and the intestine. Global gene expression analyses suggested that Rb maintains the M-bM-^@M-^\starvation-induced transcriptomeM-bM-^@M-^] and represses the M-bM-^@M-^\re-feeding induced transcriptomeM-bM-^@M-^], including the repression of many pathogen/toxin/oxidative stress-inducible and metabolic genes, as well as the activation of many other stress-resistant genes, mitochondrial respiratory chain genes, and potential IIS receptor antagonists. Notably, the majority of genes dysregulated in starved L1 Rb(-) animals were not found to be dysregulated in fed conditions. Together, these findings identify Rb as a critical regulator of the starvation response and suggest a link between functions of tumor suppressors and starvation survival. These results may provide mechanistic insights into why cancer cells are often hypersensitive to starvation treatment. Gravid C. elegans adult animals were bleached with hypochlorite and sodium hydroxide. The resulting eggs were hatched in 8M-bM-^@M-^S10 mL S-basal without cholesterol in 15-mL conical tubes, which were placed on an end-over-end rocker (VWR) at 20 M-BM-0C. 30 hours later, starved L1s were collected for RNA preparations. Wild type and lin-35/Rb mutant were profiled in triplicate. All replicates were biological replicates.