Abstract
The Pacific oyster (Crassostrea gigas) is one of the most important aquaculture species worldwide. Glycogen contributes greatly to the special taste and creamy white color of oysters. Previous genome-wide association studies (GWAS) identified several single nucleotide polymorphism (SNP) sites that were strongly related to glycogen content. Genes within 100 kb upstream and downstream of the associated SNPs were screened. One gene annotated as protein phosphatase 1 regulatory subunit 3B (PPP1R3B), which can promote glycogen synthesis together with protein phosphatase 1 catalytic subunit (PPP1C) in mammals, was selected as a candidate gene in this study. First, full-length CgPPP1R3B was cloned and its function was characterized. The gene expression profiles of CgPPP1R3B in different tissues and seasons showed a close relationship to glycogen content. RNA interference (RNAi) experiments of this gene in vivo showed that decreased CgPPP1R3B levels resulted in lower glycogen contents in the experimental group than in the control group. Co-immunoprecipitation (Co-IP) and yeast two-hybrid (Y2H) assays indicated that CgPPP1R3B can interact with CgPPP1C, glycogen synthase (CgGS) and glycogen phosphorylase (CgGP), thus participating in glycogen metabolism. Co-sedimentation analysis in vitro demonstrated that the CgPPP1R3B protein can bind to glycogen molecules directly, and these results indicated the conserved function of the CgPPP1R3B protein compared to that of mammals. In addition, thirteen SNPs were precisely mapped in this gene. Ten of the thirteen SNPs were confirmed to be significantly (p < 0.05) related to glycogen content in an independent wild population (n = 288). The CgPPP1R3B levels in oysters with high glycogen content were significantly higher than those of oysters with low glycogen content, and gene expression levels were significantly associated with various genotypes of four associated SNPs (p < 0.05). The data indicated that the associated SNPs may control glycogen content by regulating CgPPP1R3B expression. These results suggest that CgPPP1R3B is an important gene for glycogen metabolic regulation and that the associated SNPs of this gene are potential markers for oyster molecular breeding for increased glycogen content.
Highlights
The Pacific oyster (Crassostrea gigas) is one of the most vital aquaculture species worldwide
We focused on functional studies of the CgPPP1R3B gene in regulating glycogen metabolism as well as genetic variations in or near CgPPP1R3B and their potential for improving oyster glycogen content
Gonad development stage were determined based on experience and seawater temperature record according to (Lango-Reynoso et al, 2006), oysters used in each experiment were in the same gonad developmental stage
Summary
The Pacific oyster (Crassostrea gigas) is one of the most vital aquaculture species worldwide. The heritability of glycogen content in oyster was 0.29 ± 0.02, indicating that it is genetically controlled (Liu et al, 2019), glycogen content is a quantitative trait with high variation among individuals and may affected by many genes and pathways. Genetic improvement of glycogen content is necessary and feasible. Glycogen content is a carcass trait that cannot be measured without killing the animal. Indirect selective breeding for glycogen content by morphological traits seems impossible because of the weak correlation between these characteristics, as shown in several studies (Li et al, 2017; Li C. et al, 2018). Molecular breeding is indispensable for the genetic improvement of glycogen content, which requires elucidation of the underlying genetic basis and the identification of genetic variations
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