Transient Receptor Potential (TRP) Channels in the Pacific Oyster (Crassostrea gigas): Genome-Wide Identification and Expression Profiling after Heat Stress between C. gigas and C. angulata

Abstract

Transmembrane proteins are involved in an array of stress responses, particularly in thermo-sensation and thermo-regulation. In this study, we performed a genome-wide identification and characterization of the Transient Receptor Potential (TRP) genes in the Pacific oyster (Crassostrea gigas) and investigated their expression profiles after heat stress to identify critical TRPs potentially associated with thermal regulation. A total of 66 TRP genes were identified in the C. gigas, which showed significant gene expansion and tandem duplication. Meta-analysis of the available RNA-Seq data generated from samples after acute heat stress revealed a set of heat-inducible TRPs. Further examination of their expression profiles under chronic heat stress, and comparison between C. gigas and C. angulata, two oyster species with different tolerance levels to heat stress, led to the identification of TRPC3.6, TRPC3.7, and TRPV4.7 as important TRPs involved in thermal regulation in oysters. This work provided valuable information for future studies on the molecular mechanism of TRP mediated thermal tolerance, and identification of diagnostic biomarker for thermal stress in the oysters.

Keywords: Crassostrea gigas; TRPs; gene expression; phylogeny; thermal stress.

Figure 1

Identification of TRP genes and their distribution across chromosomes of C. gigas. The scale on the right is in million bases (Mb). Chromosome numbers are shown at the top of each vertical bar. Genomic locations of TRP genes are marked with the black lines.

Figure 2

Phylogenetic analysis of TRP genes in C. gigas. The maximum-likelihood tree was inferred using the edge-linked partition model in IQ-TREE program. Numbers around the nodes correspond to bootstrap support values in percentages. Accession numbers for all sequences are provided in Table S1. Abbreviations: Hsa, Homo sapiens; Mmu, Mus musculus; Dre, Danio rerio; Dme, Drosophila melanogaster, Cvi, Crassostrea virginica and Cgi, Crassostrea gigas. The C. gigas TRPs are indicated by stars.

Figure 3

Syntenic analyses of (A) TRPA1, (B) TRPV1, and (C) TRPV4 in C. gigas. The TRP genes were highlighted in solid red box, and their neighboring genes were indicated by different color boxes with gene symbols. The syntenic information of TRPs for other species was from Genomicus (v96.01).

Figure 4

Chronic heat stress experiment and survival analysis. (A) Schematic presentation of the chronic heat stress experiment; (B) Mortality comparison between two oyster species. A number of 200 juvenile oysters per group were used for the experiment. ** indicates statistical significance (p < 0.01) (ANOVA).

Figure 5

Relative expression of TRP genes between C. gigas and C. angulata after chronic heat stress. The Y-axis represents relative expression as normalized expression levels of the elongation factor 1 alpha (EF1α) gene. Each value of relative expression was shown as mean ± S.E. (n = 3). Asterisks indicate statistical significance (* indicates p < 0.05 and ** indicates p < 0.01) (ANOVA).

Figure 6

(A) The anatomy of the oyster; (B) Tissue distribution of selected TRP genes in oysters. The Y-axis represents relative expression as normalized expression levels of the elongation factor 1 alpha (EF1α) gene. Each value of relative expression was shown as mean ± S.E. (n = 3). Values in bars that share the same superscript letter are not significantly different (p > 0.05, Tukey’s test).

Figure 7

Effect of chronic heat stress on expression of (A) HIF-1α, (B) CAT, (C) SOD, (D) CAS in gill of two oyster species. Values were expressed as mean ± S.E (n = 3). Asterisks indicate statistical significance (* indicates p < 0.05 and ** indicates p < 0.01) (ANOVA). HIF-1α, hypoxia inducible factor-1α; CAT, catalase; SOD, superoxide dismutase; CAS, caspase.