Generation of Myostatin b Knockout Yellow Catfish (Tachysurus Fulvidraco) Using Transcription Activator-Like Effector Nucleases

Abstract

Myostatin (Mstn), a member of the transforming growth factor b superfamily, plays an inhibiting role in mammalian muscle growth. Mammals like human, cattle, mouse, sheep, and dog carrying null alleles of Mstn display a double-muscle phenotype. Mstn is conserved in fish; however, little is known whether the fish with mutated mstn display a similar phenotype to mammals because of the lack of mutant fish with mstn null alleles. Previously, we knocked out one of the duplicated copies of myostatin gene (mstna) in yellow catfish using zincfinger nucleases. In this study, we report the identification of the second myostatin gene (mstnb) and knockout of mstnb in yellow catfish. The gene comprises three exons. It is predicted to encode 373 amino acid residues. The predicted protein exhibits 59.3% identity with yellow catfish Mstna and 57.3% identity with human MSTN. Employing TALEN (transcription activator-like effector nucleases) technology, we obtained two founders (from four randomly selected founders) of yellow catfish carrying the mutated mstnb gene in their germ cells. Totally, six mutated alleles of mstnb were obtained from the founders. Among the six alleles, four are nonframeshift and two are frameshift mutation. The frameshift mutated alleles include mstnb nju22 , an 8 bp deletion, and mstnb nju24 , a complex type of mutation comprising a 7 bp deletion and a 12 bp insertion. They are predicted to encode function null Mstnb. Our results will help to understand the roles of mstn genes in fish growth.

Muscle Hyperplasia in Japanese Quail by Single Amino Acid Deletion in MSTN Propeptide

“…Interestingly, C42del mutation had haploinsufficiency in terms of increased body weight and muscle mass in the male and female, showing similar body weight and muscle mass between WT/C42del and WT/WT quail. This haploinsufficiency of MSTN mutation in body weight was also observed in zebrafish and catfish [12,13], which might be partially due to a unique mechanism of nutrient supply to the fetus in fishes and quail.…”

“…MSTN is mainly expressed in skeletal muscle and negatively regulates the growth of muscle [2]. This inhibitory effect on muscle growth has been further confirmed by MSTN mutation resulting in increased muscle mass in humans and animals including cattle, dogs, pigs, goats, sheep, rabbits, and fishes [3][4][5][6][7][8][9][10][11][12][13].…”

“…Function of MSTN in negative regulation of muscle growth is conserved across species and natural or genetically engineered disruption of the MSTN gene gives increased muscle mass in mice, cattle, dogs, pigs, goats, sheep, rabbits, fishes, and humans [2][3][4][5][6][7][8][9][10][11][12][13]. However, the mechanism of enhancing muscle mass by MSTN mutation is somewhat different across species.…”

Zebrafish foxc1a Plays a Crucial Role in Early Somitogenesis by Restricting the Expression of aldh1a2 Directly

“…The mutant zebrafish were screened using the method as we reported recently (22). Briefly, the sexual mature founders were allowed to mate ad libitum, producing F1 embryos.…”

Status and prospects of gene editing and transgenic in fishes

“…The MSTN gene as negative growth hormone regulator via its mutation or disruption depicted dramatic muscle growth in cattle, buffalo, sheep, pig, and dogs (Yamada 2012). Recently, MSTN gene was characterized and targeted in other fish species, such as yellow catfish (Dong et al 2014), bay scallop (Guo et al 2012) and rainbow trout (Phelps et al 2013). MSTN gene consists of various isoforms across species (Gong et al 2003).…”