Unfolding the secrets of coral-algal symbiosis

Abstract

Dinoflagellates from the genus Symbiodinium form a mutualistic symbiotic relationship with reef-building corals. Here we applied massively parallel Illumina sequencing to assess genetic similarity and diversity among four phylogenetically diverse dinoflagellate clades (A, B, C and D) that are commonly associated with corals. We obtained more than 30,000 predicted genes for each Symbiodinium clade, with a majority of the aligned transcripts corresponding to sequence data sets of symbiotic dinoflagellates and <2% of sequences having bacterial or other foreign origin. We report 1053 genes, orthologous among four Symbiodinium clades, that share a high level of sequence identity to known proteins from the SwissProt (SP) database. Approximately 80% of the transcripts aligning to the 1053 SP genes were unique to Symbiodinium species and did not align to other dinoflagellates and unrelated eukaryotic transcriptomes/genomes. Six pathways were common to all four Symbiodinium clades including the phosphatidylinositol signaling system and inositol phosphate metabolism pathways. The list of Symbiodinium transcripts common to all four clades included conserved genes such as heat shock proteins (Hsp70 and Hsp90), calmodulin, actin and tubulin, several ribosomal, photosynthetic and cytochrome genes and chloroplast-based heme-containing cytochrome P450, involved in the biosynthesis of xanthophylls. Antioxidant genes, which are important in stress responses, were also preserved, as were a number of calcium-dependent and calcium/calmodulin-dependent protein kinases that may play a role in the establishment of symbiosis. Our findings disclose new knowledge about the genetic uniqueness of symbiotic dinoflagellates and provide a list of homologous genes important for the foundation of coral-algal symbiosis.

Figure 1

Biological processes (BP) enriched in all four Symbiodinium clades, using DAVID enrichment analyses of nonredundant transcripts ⩾300 nt with BLASTx hits to SP, E-value <10⁻¹⁵.

Figure 2

The Venn diagram of proposed Symbiodinium transcripts from each clade (⩾300 nt in size) aligned to the SP database, showing the number of genes unique to each clade as well as those shared among clades. Sequence homology was inferred when the expectation value (E-value) was ⩽10⁻¹⁵.

Figure 3

Diagram of the CCaMK structure (Swulius and Waxham, 2008), showing the conserved catalytic domain (black box) required for kinase activity; autoinhibitory domain (gray box) and the C-terminal regulatory domain (white box) holding Ca²⁺-binding EF-hands (a). The multiple sequence alignment (MSA) of the conserved CCaMK region from four Symbiodinium clades including representatives of different isoforms from other species: Karenia brevis (CO064068); Lilium longiflorum (2113422A); Medicago truncatula (Q6RET7); CCaMK of Malus domestica (Q07250); Acropora digitata (adi_v1.00159); and Homo sapiens (Q14012) (b). The identical residues in all sequences are indicated by white letters with a black background (amino acids conserved in 100% of the sequences), white letters with a gray background (80% conserved) and black letters with a gray background (60% conserved). The MSA was constructed using clustalX (ftp://ftp.ebi.ac.uk/pub/software/clustalw2). Phylogenetic analyses of deduced amino acid sequences of Symbiodinium and other representative species were done using sequences from MSA (c). The phylogenetic tree was tested using a 1000-replicated bootstrap analysis (Felsenstein, 1989) and bootstrap values >50% are indicated at each node. A distance method using maximum likelihood estimates was based on the Dayhoff PAM matrix. The scale for the branch length (0.1 substitutions per site) is presented under the tree.