Dataset Information

CRISPR/Cas9-mediated mutation on an insulin-like peptide encoding gene affects the growth of the ridgetail white prawn Exopalaemon carinicauda.

ABSTRACT: Insulin-like peptides (ILPs) play key roles in animal growth, metabolism and reproduction in vertebrates. In crustaceans, one type of ILPs, insulin-like androgenic gland hormone (IAG) had been reported to be related to the sex differentiations. However, the function of other types of ILPs is rarely reported. Here, we identified another type of ILPs in the ridgetail white prawn Exopalaemon carinicauda (EcILP), which is an ortholog of Drosophila melanogaster ILP7. Sequence characterization and expression analyses showed that EcILP is similar to vertebrate insulin/IGFs and insect ILPs in its heterodimeric structure and expression profile. Using CRISPR/Cas9 genome editing technology, we generated EcILP knockout (KO) prawns. EcILP-KO individuals have a significant higher growth-inhibitory trait and mortality than those in the normal group. In addition, knockdown of EcILP by RNA interference (RNAi) resulted in slower growth rate and higher mortality. These results indicated that EcILP was an important growth regulator in E. carinicauda.

SUBMITTER: Gao Y

PROVIDER: S-EPMC9556898 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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CRISPR/Cas9-mediated mutation on an insulin-like peptide encoding gene affects the growth of the ridgetail white prawn Exopalaemon carinicauda.

Gao Yi Y Zhang Xiaojun X Yuan Jianbo J Zhang Chengsong C Li Shihao S Li Fuhua F

Frontiers in endocrinology 20220929

Insulin-like peptides (ILPs) play key roles in animal growth, metabolism and reproduction in vertebrates. In crustaceans, one type of ILPs, insulin-like androgenic gland hormone (IAG) had been reported to be related to the sex differentiations. However, the function of other types of ILPs is rarely reported. Here, we identified another type of ILPs in the ridgetail white prawn Exopalaemon carinicauda (EcILP), which is an ortholog of Drosophila melanogaster ILP7. Sequence cha ...[more]

PMID: 36246877

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Project description:BackgroundMetschnikowia bicuspidata is a pathogenic yesst that can cause disease in many different economic aquatic animal species. In recent years, there was a new disease outbreak in ridgetail white prawn (Exopalaemon carinicauda) in coastal areas of Jiangsu Province China that was referred to as zombie disease by local farmers. The pathogen was first isolated and identified as M. bicuspidata. Although the pathogenicity and pathogenesis of this pathogen in other animals have been reported in some previous studies, research on its molecular mechanisms is still very limited. Therefore, a genome-wide study is necessary to better understand the physiological and pathogenic mechanisms of M. bicuspidata.ResultIn this study, we obtained a pathogenic strain, MQ2101, of M. bicuspidata from diseased E. carinicauda and sequenced its whole genome. The size of the whole genome was 15.98 Mb, and it was assembled into 5 scaffolds. The genome contained 3934 coding genes, among which 3899 genes with biological functions were annotated in multiple underlying databases. In KOG database, 2627 genes were annotated, which were categorized into 25 classes including general function prediction only, posttranslational modification, protein turnover, chaperones, and signal transduction mechanisms. In KEGG database, 2493 genes were annotated, which were categorized into five classes, including cellular processes, environmental information processing, genetic information processing, metabolism and organismal systems. In GO database, 2893 genes were annotated, which were mainly classified in cell, cell part, cellular processes and metabolic processes. There were 1055 genes annotated in the PHI database, accounting for 26.81% of the total genome, among which 5 genes were directly related to pathogenicity (identity ≥ 50%), including hsp90, PacC, and PHO84. There were also some genes related to the activity of the yeast itself that could be targeted by antiyeast drugs. Analysis based on the DFVF database showed that strain MQ2101 contained 235 potential virulence genes. BLAST searches in the CAZy database showed that strain MQ2101 may have a more complex carbohydrate metabolism system than other yeasts of the same family. In addition, two gene clusters and 168 putative secretory proteins were predicted in strain MQ2101, and functional analysis showed that some of the secretory proteins may be directly involved in the pathogenesis of the strain. Gene family analysis with five other yeasts revealed that strain MQ2101 has 245 unique gene families, including 274 genes involved in pathogenicity that could serve as potential targets.ConclusionGenome-wide analysis elucidated the pathogenicity-associated genes of M. bicuspidate while also revealing a complex metabolic mechanism and providing putative targets of action for the development of antiyeast drugs for this pathogen. The obtained whole-genome sequencing data provide an important theoretical basis for transcriptomic, proteomic and metabolic studies of M. bicuspidata and lay a foundation for defining its specific mechanism of host infestation.

Project description:As an important pattern-recognition receptor (PRR), C-type lectins (CTLs) play significant roles in recognizing microbes and battle against pathogenic microorganism in innate immunity. In this study, two tandem threonine containing CTLs (designated as EcThr-LecA and EcThr-LecB) were identified from Exopalaemon carinicauda. The full-length cDNA of EcThr-LecA and EcThr-LecB consisted of 1521 and 1518 bp with 1251 and 1242 bp open reading frame encoding a protein with 412 and 413 amino acids, respectively. The genome structure of EcThr-LecA included 10 exons and 9 introns, and the sequences of intron6 and intron7 were variable. The nucleotide sequence of intron2 in EcThr-LecB was specific and different with that of EcThr-LecA. EcThr-LecA and EcThr-LecB proteins were predicted to have a signal peptide, two conserved carbohydrate recognition domain (CRD), and tandem threonine region. The expression levels of EcThr-LecA and EcThr-LecB in the intestine were significantly up-regulated after Vibrio parahaemolyticus and white spot syndrome virus (WSSV) challenge. RNA interference (RNAi) was used to explore the effects of EcThr-LecB silencing on the mRNA expression of anti-lipopolysaccharide factor (ALF), crustin (CRU), and lysozyme (LYSO). Knock down of EcThr-LecB could evidently down-regulate the expression of eight different antibacterial peptides (AMPs), including EcALF2, EcCRU1, EcCRU3, EcCRU4, EcLYSO1, EcLYSO2, EcLYSO3, and EcLYSO4, whereas make no effect on the transcription of EcALF1, EcALF3, EcCRU2, and EcLYSO5. The recombinant two CRD domains and tandem threonine region (RLecB) of EcThr-LecB could bind diverse bacteria, lipopolysaccharide, and peptidoglycans in vitro. In addition, RLecB could accelerate the clearance of V. parahaemolyticus in vivo. The present data indicated that new-found tandem threonine containing CTLs in E. carinicauda may act as PRR to participate in the innate immune defense against pathogens by the recognition of non-self, regulation of AMPs, and clearance of invaders.

Dataset Information

CRISPR/Cas9-mediated mutation on an insulin-like peptide encoding gene affects the growth of the ridgetail white prawn Exopalaemon carinicauda.

Publications

CRISPR/Cas9-mediated mutation on an insulin-like peptide encoding gene affects the growth of the ridgetail white prawn <i>Exopalaemon carinicauda</i>.

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