Project description:Examining the impact of short term exposure to HDPE and PVC plastic leachates on gene transcription in Prochlorococcus cultures in vitro
Project description:RNA decay was measured in Prochlorococcus after inhibition of transcription by rifampicin using customized Affymetrix gene expression arrays. RNA turnover plays an important role in the gene regulation of microorganisms and influences their speed of acclimation to environmental changes. We investigated whole-genome RNA stability of Prochlorococcus, a relatively slow-growing marine cyanobacterium doubling approximately once a day, which is extremely abundant in the oceans. Using a combination of microarrays, quantitative RT-PCR and a new algorithm for determining RNA decay rates, we found a median half-life of 2.4 min and a median decay rate of 2.6 min for expressed genes â two-fold faster than that reported for any organism. The shortest transcript half-life (33 seconds) was for a gene of unknown function, while some of the longest (ca. 18 min) were for highly expressed genes. Genes organized in operons displayed intriguing mRNA decay patterns, such as increased stability, and delayed onset of decay with greater distance from the transcriptional start site. The same phenomenon was observed on a single probe resolution for genes greater than 2 kb. We hypothesize that the fast turnover relative to generation time in Prochlorococcus may enable a swift response to environmental changes through rapid recycling of nucleotides, which could be advantageous in nutrient poor oceans. Our growing understanding of RNA half-lives will inform on the modelling of cell processes and help interpret the growing bank of metatranscriptomic studies of wild populations of Prochlorococcus. The surprisingly complex decay patterns of large transcripts reported here, and the method developed to describe them, will open new avenues for the investigation and understanding of RNA decay for all organisms. Prochlorococcus cells were treated with rifampicin, which prevents initiation of new transcripts. Cells were harvested at 0 min (before rifampicin addition), 2.5 min, 5 min, 10 min, 20 min, 40 min and 60 min after rifampicin addition.
Project description:RNA decay was measured in Prochlorococcus after inhibition of transcription by rifampicin using customized Affymetrix gene expression arrays. RNA turnover plays an important role in the gene regulation of microorganisms and influences their speed of acclimation to environmental changes. We investigated whole-genome RNA stability of Prochlorococcus, a relatively slow-growing marine cyanobacterium doubling approximately once a day, which is extremely abundant in the oceans. Using a combination of microarrays, quantitative RT-PCR and a new algorithm for determining RNA decay rates, we found a median half-life of 2.4 min and a median decay rate of 2.6 min for expressed genes – two-fold faster than that reported for any organism. The shortest transcript half-life (33 seconds) was for a gene of unknown function, while some of the longest (ca. 18 min) were for highly expressed genes. Genes organized in operons displayed intriguing mRNA decay patterns, such as increased stability, and delayed onset of decay with greater distance from the transcriptional start site. The same phenomenon was observed on a single probe resolution for genes greater than 2 kb. We hypothesize that the fast turnover relative to generation time in Prochlorococcus may enable a swift response to environmental changes through rapid recycling of nucleotides, which could be advantageous in nutrient poor oceans. Our growing understanding of RNA half-lives will inform on the modelling of cell processes and help interpret the growing bank of metatranscriptomic studies of wild populations of Prochlorococcus. The surprisingly complex decay patterns of large transcripts reported here, and the method developed to describe them, will open new avenues for the investigation and understanding of RNA decay for all organisms.
Project description:Prochlorococcus is an obligate marine microorganism which are dominant autotroph in tropical and subtropical central oceans. However, what is the low salinity boundary and how Prochlorococcus would response to low salinity exposure is still unknown. In this study, we first tested the growing salinity range of two Prochlorococcus strains, NATL1A and MED4, and then compared the global transcriptome of their low salinity acclimated cells and cells growing in normal seawater salinity. We found that MED4 could be acclimated in the lowest salinity of 25% and NATL1A could be acclimated in the lowest salinity of 28%. Measurement of the effective quantum yield of PSII photochemistry (Fv/Fm) indicated that both strains were stressed when growing in salinity lower than 34%. The transcriptomic response of NATL1A and MED4 were approximately different, with much more genes having changed transcript abundance in NATL1A than in MED4. To cope with low salinity, NATL1A downregulated the transcript of most genes involved in translation, ribosomal structure and biogenesis, while MED4 upregulated those genes. Moreover, low salinity acclimated NATL1A cells suppressed ATP-producing genes and induced the expression of photosynthesis related genes, while low salinity acclimated MED4 upregulated ATP-producing genes and downregulated photosynthesis related genes. These results indicate that the response to low salinity stress of different Prochlorococcus strains could be distinct. The study provided the first glimpse into the growing salinity range of Prochlorococcus cells and their global gene expression changes due to low salinity stress.
Project description:<p>We compared changes induced by the addition of 100 nM and 5 mM glucose on the proteome and metabolome complements in <em>Synechococcus</em> sp. strains WH8102, WH7803, and BL107 and <em>Prochlorococcus</em> sp. strains MED4, SS120, and MIT9313, grown either under standard light conditions or in darkness. Our results suggested that glucose is metabolized by these cyanobacteria, using primarily the oxidative pentoses and Calvin pathways, while no proof was found for the involvement of the Entner-Doudoroff pathway in this process. We observed differences in the effects of glucose availability, both between genera and between <em>Prochlorococcus</em> MED4 and SS120 strains, which might be related to their specific adaptations to the environment. We found evidence for fermentation in <em>Prochlorococcus</em> sp. strain SS120 and <em>Synechococcus</em> sp. strain WH8102 after 5 mM glucose addition. Our results additionally suggested that marine cyanobacteria can detect nanomolar glucose concentrations in the environment and that glucose might be used to sustain metabolism under darkness. Furthermore, the KaiB and KaiC proteins were also affected in <em>Synechococcus</em> sp. WH8102, pointing to a direct link between glucose assimilation and circadian rhythms in marine cyanobacteria. In conclusion, our study provides a wide overview on the metabolic effects induced by glucose availability in representative strains of the diverse marine picocyanobacteria, providing further evidence for the importance of mixotrophy in marine picocyanobacteria. The <em>Prochlorococcus sp.</em> strain PCC 9511 is genetically identical to MED4</p><p><strong>IMPORTANCE</strong> Glucose uptake by marine picocyanobacteria has been previously described and strongly suggests they are mixotrophic organisms (capable of using energy from the sun to make organic matter, but also to directly use organic matter from the environment when available). However, a detailed analysis of the effects of glucose addition on the proteome and metabolome of these microorganisms had not been carried out. Here, we analyzed three <em>Prochlorococcus</em> sp. and three <em>Synechococcus</em> sp. strains which were representative of several marine picocyanobacterial clades. We observed differential features in the effects of glucose availability, depending on both the genus and strain; our study illuminated the strategies utilized by these organisms to metabolize glucose and showed unexpected links to other pathways, such as circadian regulation. Furthermore, we found glucose addition had profound effects in the microbiome, favoring the growth of coexisting heterotrophic bacteria.</p>
Project description:Prochlorococcus is found throughout the euphotic zone in the oligotrophic open ocean. Deep mixing and sinking in aggregates or while attached to particles can, however, transport cells below this sunlit zone, depriving them of light for extended periods of time and influencing their circulation via ocean currents. Viability of these cells over extended periods of darkness could shape the ecology and evolution of the Prochlorococcus collective. We have shown that when co-cultured with a heterotrophic microbe and subjected to repeated periods of extended darkness, Prochlorococcus cells develop a heritable dark-tolerant phenotype – through an apparent epigenetic mechanism – such that they survive longer periods of darkness. Here we examine this adaptation at the level of physiology and metabolism in co-cultures of dark-tolerant and parent strains of Prochlorococcus, each grown with the heterotroph Alteromonas under diel light:dark conditions. The relative abundance of Alteromonas is higher in dark-tolerant than parental co-cultures, while dark tolerant Prochlorococcus cells are also larger, contain less chlorophyll, and are less synchronized to the light:dark cycle. Meta-transcriptome analysis of the cultures further suggests that dark-tolerant co-cultures undergo a coupled shift in which Prochlorococcus uses more organic carbon and less photosynthesis, and Alteromonas uses more organic acids and fewer sugars. Collectively, the data suggest that dark adaptation involves a loosening of the coupling between Prochlorococcus metabolism and the light:dark cycle and a strengthening of the coupling between the carbon metabolism of Prochlorococcus and Alteromonas.