Project description:We investigated the gene expression responses of Candidatus Pelagibacter ubique cultures to iron limitation. Differential expression was observed for genes in iron acquisition and incorporation operons. SfuC in particular was 16 times higher in iron-limited cultures and encodes a periplasmic iron-binding protein. Six natural seawater cultures were amended with minimal nutrients and inoculated with P. ubique. Close to maximum cell density, all carboys were supplemented with 100 nM ferrichrome (an iron-chelating siderophore) and three carboys were additionally supplemented with 1 µM FeCl3. Each of the six carboys was sampled for microarray analyses one, two, and eleven days after the ferrichrome addition.
Project description:Eukaryotic cytoplasm organizes itself via membrane-bound organelles and membrane-less biomolecular condensates (BMCs). Known BMCs exhibit liquid-like properties and are typically visualized on the scale of ~1 µm. They have been studied mostly by microscopy, examining select individual proteins. Here, we investigate the global organization of native cytoplasm with quantitative proteomics, using differential pressure filtration and dilution experiments. These assays revealed that BMCs form predominantly at the mesoscale of ~100 nm, which we validated via microscopy. Our data indicate that at least 18% of the proteome is organized via such mesoscale BMCs, suggesting that cells widely employ dynamic liquid-like clustering to organize their cytoplasm, at surprisingly small length scales.
Project description:Diatoms are single celled photosynthetic bloom-forming algae that are responsible for at least 20% of global primary production. Nevertheless, more than 30% of the oceans are considered “ocean deserts” due to iron limitation. We used the diatom Phaeodactylum tricornutum as a model system to explore diatom’s response to iron limitation and its interplay with susceptibility to oxidative stress. By analyzing physiological parameters and proteome profiling, we defined two distinct phases: short-term (< 3 days, phase I) and chronic (> 5 days, phase II) iron limitation. While at phase I no changes in physiological parameters were observed, molecular markers for iron starvation, such as ISIP and flavodoxin, were highly upregulated. At phase II, down regulation of numerous iron-containing proteins was detected in parallel to reduction in growth rate, chlorophyll content, photosynthetic activity, respiration rate and antioxidant capacity. Intriguingly, while application of oxidative stress to phase I and II iron limited cells similarly oxidized the GSH pool, phase II iron limitation exhibited transient resistance to oxidative stress, despite the down regulation of many antioxidant proteins. By comparing proteomic profiles of P. tricornutum under iron limitation and metatranscriptomic data of an iron enrichment experiment conducted in the Pacific Ocean, we propose that iron limited cells in the natural environment resemble the phase II metabolic state. These results provide insights into the trade-off between maximal growth rate and susceptibility to oxidative stress as a possible key determinant in the response of diatoms to iron quota in the marine environment.