Diversity analysis of sea anemone peptide toxins in different tissues of Heteractis crispa based on transcriptomics

Abstract

Peptide toxins found in sea anemones venom have diverse properties that make them important research subjects in the fields of pharmacology, neuroscience and biotechnology. This study used high-throughput sequencing technology to systematically analyze the venom components of the tentacles, column, and mesenterial filaments of sea anemone Heteractis crispa, revealing the diversity and complexity of sea anemone toxins in different tissues. A total of 1049 transcripts were identified and categorized into 60 families, of which 91.0% were proteins and 9.0% were peptides. Of those 1049 transcripts, 416, 291, and 307 putative proteins and peptide precursors were identified from tentacles, column, and mesenterial filaments respectively, while 428 were identified when the datasets were combined. Of these putative toxin sequences, 42 were detected in all three tissues, including 33 proteins and 9 peptides, with the majority of peptides being ShKT domain, β-defensin, and Kunitz-type. In addition, this study applied bioinformatics approaches to predict the family classification, 3D structures, and functional annotation of these representative peptides, as well as the evolutionary relationships between peptides, laying the foundation for the next step of peptide pharmacological activity research.

Figure 1

Cluster heatmap and genes annotated by GO analysis in three different tissues. (a) Cluster heatmap of three different tissues. (b) Bar chart of the number of genes in tentacles. (c) Bar chart of the number of genes in the column. (d) Bar chart of the number of genes in the mesenterial filaments.

Figure 2

Families of putative protein and peptide toxins in H. crispa transcriptome. Based on their amino acid sequences and cysteine scaffolds, the 956 protein sequences and 93 peptide sequences with significant BLAST hits to manually curated lists of animal toxins in UniProt (www.uniprot.org/program/Toxins) were assigned to distinct toxin families.

Figure 3

Transcripts of protein and peptide toxins from several H. crispa tissues are compared. (a) Correlation between datasets of putative protein and neurotoxic peptide detected from H. crispa combine, tentacles, column, and mesenterial filaments. (b) 42 putative protein and peptide transcripts from various H. crispa tissues. (c) The ten most greatly expressed protein and peptide transcripts from different H. crispa tissues.

Figure 4

Classification of peptide toxic cysteine patterns from H. crispa.

Figure 5

Sea anemone ShKT domain mature peptide sequences. (a–b) The conserved cysteine residues are highlighted with green text on yellow background. T, C, F, and M respectively represent tentacles, column, mesenterial filaments, and combine, highlighted in blue, orange, green, and yellow. (c) Homology modeling and ShK prediction of sea anemone mature peptides HC-36, HC-37, and HC-43 (PDB 4LFQ).

Figure 6

β-defensin-like sea anemone mature peptide sequences. (a–b) The conserved cysteine residues are highlighted with green text on yellow background. T, C, F, and M respectively represent tentacles, column, mesenterial filaments, and combine, highlighted in blue, orange, green, and yellow. (c) Homology modeling prediction of several mature peptides from sea anemones with CgNa (PDB 2H9X), BDS I (PDB 1BDS), and APETx2 (PDB 2MUB).

Figure 7

Representative mature peptide sequences from sea anemones containing Kunitz-type peptides. (a) The conserved cysteine residues are highlighted with green text on yellow background. T, C, F, and M respectively represent tentacles, column, mesenterial filaments, and combine, highlighted in blue, orange, green, and yellow. (b) Homology modeling prediction of mature peptides sea anemone HC-50 and HC-47 with SHPI-1 (PDB 3M7Q).

Figure 8

Representative Sea anemone mature peptide sequences in other families. (a) The conserved cysteine residues are highlighted with green text on yellow background. T, C, F, and M respectively represent tentacles, column, mesenterial filaments, and combine, highlighted in blue, orange, green, and yellow. (b) Homology modeling prediction of several representative sea anemone mature peptides with human EGF (PDB 7SZ1), PI-actitoxin-Avd5a (PDB 1Y1B).

Figure 9

Phylogenetic tree from sequenced 93 peptide sequences and reported sequences in the Blast database. The tree was established by the NJ approach. Sequences with the same background color indicate peptides from the same family.