Project description:Copper-limiting growth conditions were thought to cause an induction of genes possibly involved in copper uptake and sorting. This rationale in mind, we performed microarray analyses on B. japonicum cells grown in three variations of the BVM minimal medium. Variant 1 contained 2 M-NM-<M CuSO4 (copper excess). Variant 2 was prepared in HCl-treated glassware without any copper added (copper starvation). The residual copper concentration in this copper-starvation medium was analyzed by GF-AAS and determined to be 5 nM. Variant 3 (extreme copper limitation) was prepared like variant 2 but with the addition of 10 M-NM-<M BCS and 1 mM ascorbic acid where BCS chelates Cu(I) selectively, and ascorbic acid reduces any Cu(II) to Cu(I). Changes in the transcription profiles were recorded by the pairwise comparison of cells grown in variant 2 vs. 1, and variant 3 vs. 2. Only a small set of genes were differentially up- or down-regulated when copper-starved cells were compared with cells grown in copper excess. Most notably, five genes located adjacent to each other on the B. japonicum genome displayed an increased expression: bll4882 to bll4878. The five genes were named pcuA, pcuB, pcuC, pcuD, and pcuE (mnemonic of proteins for Cu trafficking). The genes with decreased expression are either of unknown function or M-bM-^@M-^S not surprisingly M-bM-^@M-^S play a role in copper resistance. Extreme copper limitation (variant 3 vs. 2) did not further enhance the expression of the five pcu genes. Instead, another cluster of adjacent genes was strongly up-regulated: bll0889 to bll0883, which code for unidentified transport functions. Incidentally, the list also includes the copper chaperone ScoI. Taken together, copper-limiting growth conditions have led to the de-repression of genes potentially involved in copper acquisition. Microarray-based transcriptome analysis of B. japonicum 110spc4 wild-type cells grown under normal, copper-limiting and copper excess conditions

Project description:Chemical contamination is a common threat to biota thriving in estuarine and coastal ecosystems. In particular, trace metals tend to accumulate and exert deleterious effects on small invertebrates such as zooplankton, which are essential trophic links between phytoplankton and higher-level consumers in aquatic food webs. Beyond the direct effects of the contamination, we hypothesized that metal exposure could also affect the zooplankton microbiota, which in turn might further impair host fitness. To assess this assumption, copepods (Eurytemora affinis) were sampled in the oligo-mesohaline zone of the Seine estuary and exposed to dissolved copper (25 µg.L-1) over a 72-hour time period. Copepod response to copper treatment was assessed by determining transcriptomic changes in E. affinis along with shifts in its microbiota. Unexpectedly, very few genes were differentially expressed in copper-treated copepods compared to controls, with most of the reported differences involving genes upregulated in males compared to females. In contrast, copper increased the taxonomic diversity indices of the microbiota and resulted in substantial compositional changes at both the phyla and genus levels. Phylogenetic reconstruction of the microbiota further suggested that copper mitigated phylogenetic relatedness of taxa at the basal tree structure of the phylogeny, whereas it strengthened it at the terminal branches. Increased terminal phylogenetic clustering in copper-treated copepods concurred with higher proportions of bacterial genera previously identified as copper resistant (e.g., Pseudomonas, Acinetobacter, and Alkanindiges) and a higher relative abundance of the copA gene encoding a periplasmic inducible multi-copper oxidase. Overall, these results revealed very contrasting responses of E. affinis and its microbiota to copper exposure. The enrichment in micro-organisms likely to perform copper sequestration and/or enzymatic transformation processes, underlines here the need to follow the microbial component during the evaluation of the vulnerability of the zooplankton to the metallic stress.

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IlLycPhla

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