The oyster genome reveals stress adaptation and complexity of shell formation

Comparison with seven other sequenced genomes identified 8,654 oyster-specific genes (Supplementary Text E3.1) that are probably important in the evolution and adaptation of oysters and other molluscs. With oysters being the only representative, these genes could be shared by other molluscs. Among these genes, gene ontology terms related to protein binding, apoptosis, cytokine activity and inflammatory response are highly enriched (P<0.0001; Supplementary Text E2 and Supplementary Table 17), indicating over-representation of some host-defence genes against biotic and abiotic stress. Manual examination shows that several gene families related to defence pathways, including protein folding, oxidation and anti-oxidation, apoptosis and immune responses, are expanded in C. gigas (Fig. 3a and Supplementary Table 18). The oyster genome contains 88 heat shock protein 70 (HSP70) genes, which have crucial roles in protecting cells against heat and other stresses, compared with ~17 in humans and 39 in sea urchins. Phylogenetic analysis finds clustering of 71 oyster HSP70 genes to themselves, suggesting that the expansion is specific to the oyster (Supplementary Fig. 19). Also expanded are cytochrome P450 (Supplementary Fig. 20) and multi-copper oxidase gene families, which are important in the biotransformation of endobiotic and xenobiotic chemicals26, and extracellular superoxide dismutases, which are important in defence against oxidative stress. The oyster genome has 48 genes coding for inhibitor of apoptosis proteins (IAPs), compared with 8 in humans and 7 in sea urchins, indicating a powerful anti-apoptosis system in oysters. Genes encoding lectin-like proteins, including C-type lectin, fibrinogen-related proteins and C1q domain-containing proteins (C1QDCs), are highly over-represented in the oyster genome (P<0.0001; Supplementary Table 18); these genes have important roles in the innate immune response in invertebrates27, 28, 29. Interestingly, many immune-related genes, including genes coding for Gram-negative bacteria-binding proteins, peptidoglycan-recognition proteins, defensin, C-type-lectin-domain-containing proteins and C1QDCs, are highly expressed in the digestive gland (Supplementary Fig. 21), indicating that the digestive system of this filter feeder is an important first-line defence organ against pathogens.

a, Expansion and expression of key genes in major stress-response pathways in C. gigas. Genes include HSPs and HSF in the heat-shock response; GRP78, CRT, CNX, GRP94, PERK, IRE1 and EIF2a in the endoplasmic reticulum unfolded-protein response (UPRER); IAPs, BCL2 like, BAG, BI1, caspases, FADD and TNFR in apoptotic pathways; CYP450 and MO in oxidation; and SOD, GPX, PRX and CAT in anti-oxidation. Boxes with bold black borders indicate gene families (HSPs, IAPs and SODs) expanded in C. gigas, and the filled colours correspond to their degree of upregulation in RPKMtreatment/RPKMcontrol by stress, found in 61 transcriptomes from oysters challenged with 9 types of stressors (Supplementary Text G2 and Supplementary Table 23). b, Venn diagram of common and unique genes expressed in response to temperature, salinity, air exposure and heavy-metal stress (zinc, cadmium, copper, lead and mercury), showing overlap of responses. c, Number of genes with and without detectable paralogues differentially expressed under stress and normal conditions, showing that genes responding to stress are more likely to have paralogues (P<11010; 2 test). Green sections of the pie chart represent 1,442, 809, 358, 550 and 7,938 paralogues for air exposure, metal, temperature, salinity and normal conditions, respectively.

To investigate genome-wide responses to stress, we sequenced 61 transcriptomes from C. gigas subjected to nine stressors, including temperature, salinity, air exposure and heavy metals (Supplementary Text G1 and Supplementary Tables 19 and 20). We found that 5,844 genes were differentially expressed under at least one stressor, and genes responding to different stressors showed significant overlap (Fig. 3b and Supplementary Fig. 23a). Air exposure induced a response from the largest number of genes (4,420), indicating that air exposure is a major stressor and that oysters have evolved an extensive gene set in defence. Genes differentially expressed in response to stress are more likely to have paralogues (Fig. 3c), suggesting that expansion and selective retention of duplicated defence-related genes are probably important to oyster adaptation. Under most stressors, genes coding for HSPs, histones, IAPs and protein biogenesis were upregulated, and those for protein degradation downregulated, pointing to concerted responses to maintain cellular homeostasis30 (Supplementary Text G3 and Supplementary Table 21). Genes involved in the unfolded protein response to cellular stress in the endoplasmic reticulum (coding for calreticulin, calnexin, 78- and 94-kDa glucose-regulated proteins) were upregulated, indicating that protein quality control is critical in cellular homeostasis under stress.

Air exposure induced up to 67-fold upregulation of five highly expressed IAPs (Supplementary Fig. 24a). Other inhibitors of apoptosis were also upregulated: BCL2 up to fourfold and BAG up to 12-fold (Supplementary Fig. 24b). These apoptosis inhibitors were also highly upregulated under heat and low salinity stress. These findings, along with the expansion of IAPs, suggest that a powerful anti-apoptosis system exists and may be critical for the amazing endurance of oysters to air exposure and other stresses. The existence of an intrinsic apoptosis pathway in invertebrates has been controversial, and parts of the pathways have only recently been demonstrated for two lophotrochozoans31, 32. The finding of key genes belonging to both intrinsic (BAX, BAK, BAG, BCL2, BI1 and procaspase) and extrinsic (TNFR and caspase 8) apoptosis pathways indicates that oysters have advanced apoptosis systems. Powerful inhibition of apoptosis as shown by genomic and transcriptomic analyses may be central to the ability of oysters to tolerate prolonged air exposure and other stresses.

Heat stress induced a ~2,000-fold increase in expression of five highly inducible HSP70 genes or a 13.9-fold increase in average expression of all HSP70 genes, amounting to 4.2% of all transcripts (Supplementary Figs 24c and 25). The genomic expansion and massive upregulation of HSP genes help to explain why C. gigas can tolerate temperatures as high as 49C when exposed to summer sun at low tide33. HSP genes were also upregulated under other stressors and may be central to the oyster defence against all stresses (Supplementary Fig. 25). HSP genes may also inhibit apoptosis by binding to effector caspases34.

Genes involved in signal transduction, including genes coding for G-protein-coupled receptors and Ras GTPase, were also activated by stressors (Supplementary Fig. 24f) and over-represented in the oyster genome (Supplementary Table 11). These regulators may have a role in orchestrating stress responses, which seem to be well coordinated (Fig. 3a and Supplementary Fig. 25). The expansion of key defence genes and the strong, complex transcriptomic response to stress highlight the sophisticated genomic adaptations of the oyster to sessile life in a highly stressful environment.

View post:
The oyster genome reveals stress adaptation and complexity of shell formation