Project description:Coral bleaching has devastating effects on coral survival and reef ecosystem function, but many of the fundamental cellular effects of thermal stress on cnidarian physiology are unclear. We used label-free liquid chromatography-tandem mass spectrometry to assess the effects of high temperatures on the proteome of the model symbiotic anemone Aiptasia sp. Anemones were acclimated to elevated temperatures (30 °C and 33.5 °C) for two weeks or exposed to short-term thermal shock (33.5 °C, 24 hours) without acclimation. We identified 2,137 protein clusters in Aiptasia, 136 of which were differentially abundant between treatments. There were minimal differences (nine proteins) in protein abundances between the control (25 °C) and acclimated high-temperature (30 °C and 33.5 °C) treatments, indicating that thermal acclimation in symbiotic cnidarians is not primarily regulated at the level of protein expression. Heat shock resulted in significant changes in the abundance of 104 proteins, including those involved in protein folding and synthesis, redox homeostasis, and central metabolism. Nineteen highly abundant cytoskeletal and structural proteins showed particularly reduced abundance (approximately 50%), demonstrating proteostasis disruption and inhibition of protein synthesis. Heat shock induced proteins involved in multiple mechanisms for stabilizing nascent proteins, preventing protein aggregation and degrading damaged proteins, indicative of endoplasmic reticulum stress. Antioxidant mechanisms and metabolic enzymes necessary for redox homeostasis were also upregulated. Disruption of host proteostasis occurred before either bleaching or symbiont photoinhibition was detected, strongly suggesting endogenous reactive oxygen species production as the proximate cause of thermal damage. The effects of thermal shock were most pronounced at the endoplasmic reticulum, and proteostasis maintenance and protein turnover mechanisms may be essential in the response to severe thermal stress in symbiotic cnidarians.

Project description:Background Regulation of transcription is essential for any organism and Rhizobium etli (a multi-replicon, nitrogen-fixing symbiotic bacterium) is no exception. This bacterium is commonly found in the rhizosphere (free-living) or inside of root-nodules of the common bean (Phaseolus vulgaris) in a symbiotic relationship. Abiotic stresses, such as high soil temperatures and salinity, compromise the genetic stability of R. etli and therefore its symbiotic interaction with P. vulgaris. However, it is still unclear which genes are up- or down-regulated to cope with these stress conditions. The aim of this study was to identify the genes and non-coding RNAs (ncRNAs) that are differentially expressed under heat and saline shock, as well as the promoter regions of the up-regulated loci. Results Analysing the heat and saline shock responses of R. etli CE3 through RNA-Seq, we identified 756 and 392 differentially expressed genes, respectively, and 106 were up-regulated under both conditions. Notably, the set of genes over-expressed under either condition was preferentially encoded on plasmids, although this observation was more significant for the heat shock response. In contrast, during either saline shock or heat shock, the down-regulated genes were principally chromosomally encoded. Our functional analysis shows that genes encoding chaperone proteins were up-regulated during the heat shock response, whereas genes involved in the metabolism of compatible solutes were up-regulated following saline shock. Furthermore, we identified thirteen and nine ncRNAs that were differentially expressed under heat and saline shock, respectively, as well as eleven ncRNAs that had not been previously identified. Finally, using an in silico analysis, we studied the promoter motifs in all of the non-coding regions associated with the genes and ncRNAs up-regulated under both conditions. Conclusions Our data suggest that the replicon contribution is different for different stress responses and that the heat shock response is more complex than the saline shock response. In general, this work exemplifies how strategies that not only consider differentially regulated genes but also regulatory elements of the stress response provide a more comprehensive view of bacterial gene regulation.

Project description:Background Regulation of transcription is essential for any organism and Rhizobium etli (a multi-replicon, nitrogen-fixing symbiotic bacterium) is no exception. This bacterium is commonly found in the rhizosphere (free-living) or inside of root-nodules of the common bean (Phaseolus vulgaris) in a symbiotic relationship. Abiotic stresses, such as high soil temperatures and salinity, compromise the genetic stability of R. etli and therefore its symbiotic interaction with P. vulgaris. However, it is still unclear which genes are up- or down-regulated to cope with these stress conditions. The aim of this study was to identify the genes and non-coding RNAs (ncRNAs) that are differentially expressed under heat and saline shock, as well as the promoter regions of the up-regulated loci. Results Analysing the heat and saline shock responses of R. etli CE3 through RNA-Seq, we identified 756 and 392 differentially expressed genes, respectively, and 106 were up-regulated under both conditions. Notably, the set of genes over-expressed under either condition was preferentially encoded on plasmids, although this observation was more significant for the heat shock response. In contrast, during either saline shock or heat shock, the down-regulated genes were principally chromosomally encoded. Our functional analysis shows that genes encoding chaperone proteins were up-regulated during the heat shock response, whereas genes involved in the metabolism of compatible solutes were up-regulated following saline shock. Furthermore, we identified thirteen and nine ncRNAs that were differentially expressed under heat and saline shock, respectively, as well as eleven ncRNAs that had not been previously identified. Finally, using an in silico analysis, we studied the promoter motifs in all of the non-coding regions associated with the genes and ncRNAs up-regulated under both conditions. Conclusions Our data suggest that the replicon contribution is different for different stress responses and that the heat shock response is more complex than the saline shock response. In general, this work exemplifies how strategies that not only consider differentially regulated genes but also regulatory elements of the stress response provide a more comprehensive view of bacterial gene regulation. mRNA of nine independent cultures of wild type strain Rhizobium etli CE3 were sequenced using Illumina GAIIx. Our parameters were: all cultures were taken from exponential phase, at 30M-BM-0C for 30min in control condition; 42M-BM-0C for 30min for the culture subject to heat -shock, and 30min at 30M-BM-0C in supplementary PY medium with 80mM NaCl for the saline shock condition.

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