Comparative Genomic and Transcriptomic Analyses of CHHs and Their Putative Receptors in Scylla paramamosain, Portunus trituberculatus, and Eriocheir sinensis

Abstract

Crustacean hyperglycemic hormone superfamily neuropeptides (CHHs) are typical crustacean eyestalk hormones that include the crustacean hyperglycemic hormone (CHH), moult-inhibiting hormone (MIH), vitellogenesis/gonad-inhibiting hormone (VIH/GIH) and mandibular organ-inhibiting hormone (MOIH), which are divided into two subfamilies: type I CHH (included CHH) and type II CHH (consisting of MIH, VIH/GIH, and MOIH). They are involved in various biological activities, such as metabolism, molting, reproduction, and osmotic regulation. Discovery of the ion transport peptide (ITP) in insects expanded the members of CHHs and revealed that CHHs are not restricted to crustaceans. In this study, we focused on three economically important crabs: the mud crab, Scylla paramamosain, the swimming crab, Portunus trituberculatus, and the Chinese mitten crab, Eriocheir sinensis. Their genomes, Pacbio full-length transcriptomic data as well as comparative RNA-seq data were obtained and used to analyze the genomic structures and expression patterns of CHHs and their putative receptors through bioinformatic methods. Two type I CHH members (CHH1 and CHH2) were identified, of which CHH1 had two splice variants, CHH1-v1 and CHH1-v2. One copy of type II CHH (MIH) was found in P. trituberculatus and E. sinensis. While most decapods, including S. paramamosain, have two copies of type II CHHs (MIH/VIH), these MIH/VIHs are adjacent to each other on the same chromosome. Besides type I and II CHH, ITP-like peptides have also been found in the three crabs, and they are mainly expressed in the eyestalk. Four, five, and three G protein-coupled receptors (GPCRs) were identified in S. paramamosain, P. trituberculatus, and E. sinensis, respectively, which might be putative CHH receptors. These GPCRs were divided into three groups. One group was composed of two contiguous genomic position GPCRs, and they were mainly expressed in the hepatopancreas. These findings provide a basis for further studies on CHHs receptor binding tests and on CHHs/GPCRs signaling pathways.

In silico analysis of crustacean hyperglycemic hormone family G protein-coupled receptor candidates

“… 2 Scylla paramamosain . From ( 22 ). Sp-GPCR-A36 and -A37 appear to be isoforms generated by alternative splicing.…”

“…In C. maenas , only CFRC-A24β1 and CFRC-A34β1 were present in the YO transcriptome [ Table 3 ( 23 )]. Although differential tissue expression of CFRCs is reported for E. sinensis , S. paramamosain , and P. trituberculatus , expression in the YO was not included in the analysis ( 22 ).…”

“…Reference sequences for B. mori A2, A24, and A34 ITP receptors were obtained from (18) (GenBank accessions NP_001127737.1, NP_001127722.1, NP_001127750.1, respectively). Other CFRC reference sequences were sourced from the NCBI GenBank database, with a particular focus on hexapod sequences that were classified as potential ITP receptor candidates, related GPCRs, and sequences that were previously identified from crustaceans (17,19,(21)(22)(23). These sequences served as the input for iterative NCBI-BLAST (blastp 2.13.0+) searches with the intent of retrieving a sufficiently broad array of sequences for phylogenetic inference.…”

“…Phylogenetic analysis showed that Pc-GPCR-A9 clustered with BNGR-A24 and Pc-GPCR-A52, -A53, and -A63 clustered with BNGR-A34 (19). Subsequently, putative CHH family receptors were identified in transcriptomes from green shore crab, Carcinus maenas; spiny lobsters, Sagmariasus verreauxi and Panulirus argus; blackback land crab, Gecarcinus lateralis; mud crab, Scylla paramamosain; swimming crab, Portunus trituberculatus; Chinese mitten crab, Eriocheir sinensis; American lobster, Homarus americanus; Norway lobster, Nephrops norvegicus; and blue crab, Callinectes sapidus (17,(21)(22)(23)(24)(25).…”

“…Any effort to identify the MIH receptor must start with a comprehensive search for ITP receptor homologs in crustacean transcriptomes, particularly those detected in the YO. Previous efforts using sequence homology, though partially successful, were hampered by fragmented and siloed databases, representing a relatively small number of species and taxonomic groups (17,19,(21)(22)(23)(24)(25). CHH/MIH/GIH/MOIH peptides probably arose after the Hexapoda-Malacostraca split approximately 515 million years ago (39,40).…”