Transcriptomic changes with increasing algal symbiont reveal the detailed process underlying establishment of coral-algal symbiosis

To clarify the establishment process of coral-algal symbiotic relationships, coral transcriptome changes during increasing algal symbiont densities were examined in juvenile corals following inoculation with the algae Symbiodinium goreaui (clade C) and S. trenchii (clade D), and comparison of their transcriptomes with aposymbiotic corals by RNA-sequencing. Since Symbiodinium clades C and D showed very different rates of density increase, comparisons were made of early onsets of both symbionts, revealing that the host behaved differently for each. RNA-sequencing showed that the number of differentially-expressed genes in corals colonized by clade D increased ca. two-fold from 10 to 20 days, whereas corals with clade C showed unremarkable changes consistent with a slow rate of density increase. The data revealed dynamic metabolic changes in symbiotic corals. In addition, the endocytosis pathway was also upregulated, while lysosomal digestive enzymes and the immune system tended to be downregulated as the density of clade D algae increased. The present dataset provides an enormous number of candidate symbiosis-related molecules that exhibit the detailed process by which coral-algal endosymbiosis is established.


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Supplementary Table   Table. S1.   The contigs containing candidate coding sequences were BLASTX aligned against the NCBI nr protein database to evaluate sequence similarity to genes in other species with an E-value cutoff of 1e-04. To identify enriched functional gene groups and pathways, GO enrichment analysis was performed using the database for annotation, visualization, and integrated discovery (DAVID). Significant processes were selected based on a corrected P-value using the method of Benjamini with a cutoff of 0.05. Pathway enrichment analysis was also conducted using KEGG.  The distribution of GC contents of Acropora , Symbiodinium reference gene dataset (A), and GC contents of our assembled contigs from Illumina reads (B). Pink bars shows GC contents of Acropora data and blue r bars shows GC contents of Symbiodinium reference data. Symbiodinium dataset have a higher GC-contents in contrast to the Acropora dataset. Up&regulated"genes" "in"symbio6c"corals" Down&regulated"genes" "in"symbio6c"corals" Clade"C" Symbiosis" Day"10" Clade"D" Symbiosis" Day"10" Clade"D" Symbiosis" Day"20" Clade"C" Symbiosis" Day"20" Clade"C" Symbiosis" Day"10" Clade"D" Symbiosis" Day"10" Clade"D" Symbiosis" Day"20" Clade"C" Symbiosis" Day"20" Heat map shows the expression patterns of genes related to Regulation of immune system process in aposymbiotic corals, corals associated with clade C (C-coral) or clade D (D-corals) in 10 days and 20 days after inoculation of each Symbiodinium. There are selected notable treatment-specific genes (FDR P < 0.05).

Fig. S a. Heat map of the expression patterns of genes related to endocytosis.
Heat map shows the expression patterns of genes related to endocytosis in aposymbiotic corals, corals associated with clade C (C-coral) or clade D (D-corals) in 10 days and 20 days after inoculation of each Symbiodinium. Gnes hose expression were down-regulated were shown. There are selected notable treatment-specific genes (FDR P < 0.05).

Fig. S b. Heat map of the expression patterns of genes related to endocytosis.
Heat map shows the expression patterns of genes related to endocytosis in each endosymbiotic Symbiodinium in 10 days and 20 days after inoculation to corals. There are selected notable treatmentspecific genes (FDR P < 0.05).

Fig. S7. Heat map of the expression patterns of genes related to lysosomal acid hydrolases.
Heat map shows the expression patterns of genes related to lysosomal acid hydrolases in aposymbiotic corals, corals associated with clade C (C-coral) or clade D (D-corals) in 10 days and 20 days after inoculation of each Symbiodinium. Genes hose expression were down-regulated were shown. There are selected notable treatment-specific genes (FDR P < 0.05).

Fig. S8. Heat map of the expression patterns of genes related to RNAi.
Heat map shows the expression patterns of genes related to RNAi in each endosymbiotic Symbiodinium in 10 days and 20 days after inoculation to corals. There are selected notable treatment-specific genes (FDR P < 0.05).

Fig. S9. Heat map of the expression patterns genes related to Glucose, Fluctose metabolic process.
Heat map shows the expression patterns genes in in aposymbiotic corals, corals associated with clade C (C-coral) or clade D (D-corals) in 10 days and 20 days after inoculation of each Symbiodinium. There are selected notable treatment-specific genes (FDR P < 0.05).  Heat map shows the expression patterns genes related to carbon fixation in aposymbiotic corals, corals associated with clade C (C-coral) or clade D (D-corals) in 10 days and 20 days after inoculation of each Symbiodinium. There are selected notable treatment-specific genes (FDR P < 0.05).

Fig. S10. Heat map of the expression patterns genes related to biosynthesis of amino acids.
Heat map shows the expression patterns genes related to biosynthesis of amino acids in aposymbiotic corals, corals associated with clade C (C-coral) or clade D (D-corals) in 10 days and 20 days after inoculation of each Symbiodinium. There are selected notable treatment-specific genes (FDR P < 0.05).

Fig. S13. Heat map of the expression patterns genes related to Insulin signaling.
Insulin signaling in aposymbiotic corals, corals associated with clade C (C-coral) or clade D (D-corals) in 10 days and 20 days after inoculation of each Symbiodinium. And expression data of Insulin-related genes of Symbiodinium are also shown. There are selected notable treatment-specific genes (FDR P < 0.05).