Project description:Background Coral reefs belong to the most ecologically and economically important ecosystems on our planet. Yet, they are under steady decline worldwide due to rising sea surface temperatures, disease, and pollution. Understanding the molecular impact of these stressors on different coral species is imperative in order to predict how coral populations will respond to this continued disturbance. The use of molecular tools such as microarrays has provided deep insight into the molecular stress response of corals. Here, we have performed comparative genomic hybridizations (CGH) with different coral species to an Acropora palmata microarray platform containing 13,546 cDNA clones in order to identify potentially rapidly evolving genes and to determine the suitability of existing microarray platforms for use in gene expression studies (via heterologous hybridization). Results Our results showed that the current microarray platform for A. palmata is able to provide biological relevant information for a wide variety of coral species covering both the complex clade as well the robust clade. Analysis of the fraction of highly diverged genes showed a significantly higher amount of genes without annotation corroborating previous findings that point towards a higher rate of divergence for taxonomically restricted genes. Among the genes with annotation, we found many mitochondrial genes to be highly diverged in M. faveolata when compared to A. palmata, while the majority of nuclear encoded genes maintained an average divergence rate. Conclusions The use of present microarray platforms for transcriptional analyses in different coral species will greatly enhance the understanding of the molecular basis of stress and health and highlight evolutionary differences between scleractinian coral species. On a genomic basis, we show that cDNA arrays can be used to identify patterns of divergence. Mitochondrion-encoded genes seem to have diverged faster than nuclear encoded genes in robust corals. Accordingly, this needs to be taken into account when using mitochondrial markers for scleractinian phylogenies.

Project description:Background Coral reefs belong to the most ecologically and economically important ecosystems on our planet. Yet, they are under steady decline worldwide due to rising sea surface temperatures, disease, and pollution. Understanding the molecular impact of these stressors on different coral species is imperative in order to predict how coral populations will respond to this continued disturbance. The use of molecular tools such as microarrays has provided deep insight into the molecular stress response of corals. Here, we have performed comparative genomic hybridizations (CGH) with different coral species to an Acropora palmata microarray platform containing 13,546 cDNA clones in order to identify potentially rapidly evolving genes and to determine the suitability of existing microarray platforms for use in gene expression studies (via heterologous hybridization). Results Our results showed that the current microarray platform for A. palmata is able to provide biological relevant information for a wide variety of coral species covering both the complex clade as well the robust clade. Analysis of the fraction of highly diverged genes showed a significantly higher amount of genes without annotation corroborating previous findings that point towards a higher rate of divergence for taxonomically restricted genes. Among the genes with annotation, we found many mitochondrial genes to be highly diverged in M. faveolata when compared to A. palmata, while the majority of nuclear encoded genes maintained an average divergence rate. Conclusions The use of present microarray platforms for transcriptional analyses in different coral species will greatly enhance the understanding of the molecular basis of stress and health and highlight evolutionary differences between scleractinian coral species. On a genomic basis, we show that cDNA arrays can be used to identify patterns of divergence. Mitochondrion-encoded genes seem to have diverged faster than nuclear encoded genes in robust corals. Accordingly, this needs to be taken into account when using mitochondrial markers for scleractinian phylogenies. Microarray experiments were performed as follows. Appropriate Cy3 and Cy5 labeled DNAs were mixed together in a hybridization buffer containing 0.25% SDS, 25 mM HEPES and 3 × SSC, resulting in a final volume of 70?l. The hybridization mixtures were boiled for 2 min at 99°C and allowed to cool at room temperature for 5 min. The cooled hybridization mixtures were pipetted under an mSeries Lifterslip (Erie Scientific), and hybridization took place in Corning hybridization chambers overnight at 55°C. Microarrays were washed once in 2 × SSC, 0.03% SDS heated to 55 °C for 5 min. followed by one wash in 1 x SSC and another wash in 0.2 x SSC for 5 min each. The slides were kept in 0.2 × SSC until analysis. Slides were dried via centrifugation and scanned using an Axon 4000B scanner. The experimental setup followed a reference design, i.e., all samples were hybridized against the same pool of labeled A. palmata DNA. For each species, a total of four hybridizations were performed, including two dye swap hybridizations in order to account for potential dye bias i.e. two hybridizations with Cy3 labeled M. faveolata DNA against a Cy5 labeled A. palmata reference and two hybridizations with Cy5 labeled M. faveolata DNA against a Cy3 labeled Cy3 A. palmata reference were performed. The same hybridization scheme was used for A. cervicornis and S. radians.

Project description:Purpose: Corals are major sources of dimethylsulphoniopropionate (DMSP), a compound that plays a central role in the global sulphur cycle. While DMSP biosynthesis pathways have been investigated in plants and algae, the molecular basis for its production by corals is unknown. Given its potential role as an osmolyte, the effect of salinity stress on levels of DMSP was investigated in both adults and juveniles (lacking photosynthetic symbionts) of the coral Acropora millepora. This study used transcriptomic data to analyse the effects of salinity over the coral A. millepora and to identify coral genes likely to be involved in DMSP biosynthesis. Methods: Adults coral transcriptomic libraries were constructed from samples exposed during 1 and 24 hours of salinity treatment (25 PSU) and control (35 PSU) conditions (n=5 per condition). Juveniles coral transcriptomic libraries were constructed from samples exposed to 24 and 48 hours of salinity treatment (28 PSU) and control (35 PSU) conditions (n=6 per condition). All libraries were sequenced by 100 bp paired-end in a HiSeq 2000. Reads were mapped onto the Acropora millepora genome using TopHat2 to produce a count data gene expression matrix for subsequent gene expression analysis using DESeq2 package. Results: In adult coral samples, 5.5 - 10.2 million RNAseq reads were obtained for each treatment sampling time while 3.4 - 8.8 million reads were obtained for each juvenile coral sample. The count matrix of the 26,622 A. millepora gene predictions were generated using htseq-count workflow. BlastP analysis of the A. millepora gene predictions led to the identification of coral members of gene families implicated in DMSP biosynthesis in other organisms, while RNA-seq data was used to identify the differentially expressed ones in response to hyposaline stress and on this basis were considered to be candidates for roles in DMSP biosynthesis in corals. Conclusions: Hyposaline stress increased DMSP production in both adults and aposymbiotic juvenile corals, and transcriptomic analyses highlighted the potential involvement of specific candidate genes in the production of DMSP via an alga-like pathway. The biochemistry of DMSP production is not well established for any eukaryotic system and, as the first animals in which it has been demonstrated, this is particularly true in the case of corals. Our RNA-seq results enabled the identification of candidates for roles in DMSP biosynthesis in corals but, given its critical roles in diverse biological processes, a thorough investigation of the molecular mechanisms leading to its production by corals is required.

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Evolutionary insights into scleractinian corals using comparative genomic hybridizations

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